Factors that play a role in selecting a suitable HPLC column.
\\n\\n
Released this past November, the list is based on data collected from the Web of Science and highlights some of the world’s most influential scientific minds by naming the researchers whose publications over the previous decade have included a high number of Highly Cited Papers placing them among the top 1% most-cited.
\\n\\nWe wish to congratulate all of the researchers named and especially our authors on this amazing accomplishment! We are happy and proud to share in their success!
Note: Edited in March 2021
\\n"}]',published:!0,mainMedia:{caption:"Highly Cited",originalUrl:"/media/original/117"}},components:[{type:"htmlEditorComponent",content:'IntechOpen is proud to announce that 191 of our authors have made the Clarivate™ Highly Cited Researchers List for 2020, ranking them among the top 1% most-cited.
\n\nThroughout the years, the list has named a total of 261 IntechOpen authors as Highly Cited. Of those researchers, 69 have been featured on the list multiple times.
\n\n\n\nReleased this past November, the list is based on data collected from the Web of Science and highlights some of the world’s most influential scientific minds by naming the researchers whose publications over the previous decade have included a high number of Highly Cited Papers placing them among the top 1% most-cited.
\n\nWe wish to congratulate all of the researchers named and especially our authors on this amazing accomplishment! We are happy and proud to share in their success!
Note: Edited in March 2021
\n'}],latestNews:[{slug:"webinar-introduction-to-open-science-wednesday-18-may-1-pm-cest-20220518",title:"Webinar: Introduction to Open Science | Wednesday 18 May, 1 PM CEST"},{slug:"step-in-the-right-direction-intechopen-launches-a-portfolio-of-open-science-journals-20220414",title:"Step in the Right Direction: IntechOpen Launches a Portfolio of Open Science Journals"},{slug:"let-s-meet-at-london-book-fair-5-7-april-2022-olympia-london-20220321",title:"Let’s meet at London Book Fair, 5-7 April 2022, Olympia London"},{slug:"50-books-published-as-part-of-intechopen-and-knowledge-unlatched-ku-collaboration-20220316",title:"50 Books published as part of IntechOpen and Knowledge Unlatched (KU) Collaboration"},{slug:"intechopen-joins-the-united-nations-sustainable-development-goals-publishers-compact-20221702",title:"IntechOpen joins the United Nations Sustainable Development Goals Publishers Compact"},{slug:"intechopen-signs-exclusive-representation-agreement-with-lsr-libros-servicios-y-representaciones-s-a-de-c-v-20211123",title:"IntechOpen Signs Exclusive Representation Agreement with LSR Libros Servicios y Representaciones S.A. de C.V"},{slug:"intechopen-expands-partnership-with-research4life-20211110",title:"IntechOpen Expands Partnership with Research4Life"},{slug:"introducing-intechopen-book-series-a-new-publishing-format-for-oa-books-20210915",title:"Introducing IntechOpen Book Series - A New Publishing Format for OA Books"}]},book:{item:{type:"book",id:"10774",leadTitle:null,fullTitle:"Model Organisms in Plant Genetics",title:"Model Organisms in Plant Genetics",subtitle:null,reviewType:"peer-reviewed",abstract:"Model plants are required for research when targeted plant species are difficult to study or when research material is unavailable. Importantly, knowledge gained from model plants can be generally translated to other related plant species because many key cellular and molecular processes are conserved and regulated by ‘blueprint’ genes inherited from a common ancestor. Model Organisms in Plant Genetics addresses characteristics of model plants such as Arabidopsis, moss, soybean, maize, and cotton, highlighting their advantages and limitations as well as their importance in studies of plant development, plant genome polyploidization, adaptive selection, evolution, and domestication, as well as their importance in crop improvement.",isbn:"978-1-83969-750-0",printIsbn:"978-1-83969-749-4",pdfIsbn:"978-1-83969-751-7",doi:null,price:119,priceEur:129,priceUsd:155,slug:"model-organisms-in-plant-genetics",numberOfPages:112,isOpenForSubmission:!1,isInWos:null,isInBkci:!1,hash:"f6624b58571ac10c9b636c5d85ec5e54",bookSignature:"Ibrokhim Y. Abdurakhmonov",publishedDate:"June 23rd 2022",coverURL:"https://cdn.intechopen.com/books/images_new/10774.jpg",numberOfDownloads:628,numberOfWosCitations:0,numberOfCrossrefCitations:2,numberOfCrossrefCitationsByBook:null,numberOfDimensionsCitations:3,numberOfDimensionsCitationsByBook:null,hasAltmetrics:0,numberOfTotalCitations:5,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"March 26th 2021",dateEndSecondStepPublish:"May 27th 2021",dateEndThirdStepPublish:"July 26th 2021",dateEndFourthStepPublish:"October 14th 2021",dateEndFifthStepPublish:"December 13th 2021",currentStepOfPublishingProcess:5,indexedIn:"1,2,3,4,5,6",editedByType:"Edited by",kuFlag:!1,featuredMarkup:null,editors:[{id:"213344",title:"Prof.",name:"Ibrokhim Y.",middleName:null,surname:"Abdurakhmonov",slug:"ibrokhim-y.-abdurakhmonov",fullName:"Ibrokhim Y. Abdurakhmonov",profilePictureURL:"https://mts.intechopen.com/storage/users/213344/images/system/213344.jpg",biography:'Ibrokhim Y. Abdurakhmonov received a BS in Biotechnology from the National University, California, in 1997, an MS in Plant Breeding from Texas A&M University in 2001, and a Ph.D. in Molecular Genetics, DSc in Genetics, and a full professorship in Molecular Genetics and Molecular Biotechnology from the Academy of Sciences of Uzbekistan in 2002, 2009, and 2011, respectively. He founded the Center of Genomics and Bioinformatics of Uzbekistan in 2012. He received the 2010 prize from The World Academy of Sciences (TWAS) and \\"ICAC Cotton Researcher of the Year 2013\\" for his outstanding contribution to cotton genomics and biotechnology. He was elected as a fellow to TWAS in 2014 and as a member of the Academy of Sciences of Uzbekistan in 2017. In the same year, he was appointed Minister of Innovative Development of Uzbekistan.',institutionString:"Academy of Sciences of Uzbekistan",position:null,outsideEditionCount:0,totalCites:0,totalAuthoredChapters:"13",totalChapterViews:"0",totalEditedBooks:"13",institution:null}],equalEditorOne:null,equalEditorTwo:null,equalEditorThree:null,coeditorOne:null,coeditorTwo:null,coeditorThree:null,coeditorFour:null,coeditorFive:null,topics:[{id:"311",title:"Plant Genetics",slug:"agronomy-plant-genetics"}],chapters:[{id:"81301",title:"Introductory Chapter: Model Plants for Discovering the Key Biological Processes in Plant Research",doi:"10.5772/intechopen.103759",slug:"introductory-chapter-model-plants-for-discovering-the-key-biological-processes-in-plant-research",totalDownloads:2,totalCrossrefCites:0,totalDimensionsCites:0,hasAltmetrics:0,abstract:null,signatures:"Ibrokhim Y. Abdurakhmonov",downloadPdfUrl:"/chapter/pdf-download/81301",previewPdfUrl:"/chapter/pdf-preview/81301",authors:[{id:"213344",title:"Prof.",name:"Ibrokhim Y.",surname:"Abdurakhmonov",slug:"ibrokhim-y.-abdurakhmonov",fullName:"Ibrokhim Y. Abdurakhmonov"}],corrections:null},{id:"78295",title:"Overview of Arabidopsis as a Genetics Model System and Its Limitation, Leading to the Development of Emerging Plant Model Systems",doi:"10.5772/intechopen.99818",slug:"overview-of-em-arabidopsis-em-as-a-genetics-model-system-and-its-limitation-leading-to-the-developme",totalDownloads:202,totalCrossrefCites:0,totalDimensionsCites:0,hasAltmetrics:0,abstract:"Model plant systems make it easier to perform experiments with them. They help to understand and expand our knowledge about the genetic basis behind different plant process. Also, it is easier to design and perform genetic and genomic experiments using a model plant system. A. thaliana was initially chosen as the model plant system, and remains to this date, one of the most widely studied plant. With the advent of better molecular biology and sequencing tools and to understand the genetic basis for the unique processes in different plant species, there is emergence of several new model systems.",signatures:"Madhabendra Mohon Kar and Ayan Raichaudhuri",downloadPdfUrl:"/chapter/pdf-download/78295",previewPdfUrl:"/chapter/pdf-preview/78295",authors:[{id:"414526",title:"Dr.",name:"Ayan",surname:"Raichaudhuri",slug:"ayan-raichaudhuri",fullName:"Ayan Raichaudhuri"},{id:"428285",title:"Mr.",name:"Madhabendra Mohon",surname:"Kar",slug:"madhabendra-mohon-kar",fullName:"Madhabendra Mohon Kar"}],corrections:null},{id:"79173",title:"Mosses: Accessible Systems for Plant Development Studies",doi:"10.5772/intechopen.100535",slug:"mosses-accessible-systems-for-plant-development-studies",totalDownloads:140,totalCrossrefCites:0,totalDimensionsCites:1,hasAltmetrics:0,abstract:"Mosses are a cosmopolitan group of land plants, sister to vascular plants, with a high potential for molecular and cell biological research. The species Physcomitrium patens has helped gaining better understanding of the biological processes of the plant cell, and it has become a central system to understand water-to-land plant transition through 2D-to-3D growth transition, regulation of asymmetric cell division, shoot apical cell establishment and maintenance, phyllotaxis and regeneration. P. patens was the first fully sequenced moss in 2008, with the latest annotated release in 2018. It has been shown that many gene functions and networks are conserved in mosses when compared to angiosperms. Importantly, this model organism has a simplified and accessible body structure that facilitates close tracking in time and space with the support of live cell imaging set-ups and multiple reporter lines. This has become possible thanks to its fully established molecular toolkit, with highly efficient PEG-assisted, CRISPR/Cas9 and RNAi transformation and silencing protocols, among others. Here we provide examples on how mosses exhibit advantages over vascular plants to study several processes and their future potential to answer some other outstanding questions in plant cell biology.",signatures:"Jordi Floriach-Clark, Han Tang and Viola Willemsen",downloadPdfUrl:"/chapter/pdf-download/79173",previewPdfUrl:"/chapter/pdf-preview/79173",authors:[{id:"420613",title:"Assistant Prof.",name:"Viola",surname:"Willemsen",slug:"viola-willemsen",fullName:"Viola Willemsen"},{id:"420615",title:"Dr.",name:"Jordi",surname:"Floriach-Clark",slug:"jordi-floriach-clark",fullName:"Jordi Floriach-Clark"},{id:"420616",title:"Dr.",name:"Han",surname:"Tang",slug:"han-tang",fullName:"Han Tang"}],corrections:null},{id:"81949",title:"Maize (Zea mays L.) as a Model System for Plant Genetic, Genomic, and Applied Research",doi:"10.5772/intechopen.104658",slug:"maize-em-zea-mays-em-l-as-a-model-system-for-plant-genetic-genomic-and-applied-research",totalDownloads:6,totalCrossrefCites:0,totalDimensionsCites:0,hasAltmetrics:0,abstract:"Maize leads the world’s cereals after wheat and rice in terms of cultivated area, because of its economic importance for the production of both food purposes and raw materials for industry. The maize genus Zea L. belonging to the family of cereals (Poaceae or Graminaceae) includes six species. However, all cultivated maize belongs specifically to Zea mays L. subsp. mays (2n = 2× = 20) is the only cultivated species of the genus Zea L., and the remaining species of this genus are mostly wild herbaceous plants. In addition to meeting the nutritional needs of the world’s population, Zea mays L. is one of the classic model objects of genetic and physiological research, as well as in the field of breeding not only cereals but also other important agricultural plants. Especially, this model object has been used in genetic mapping of loci of quantitative traits and genes associated with economically valuable traits, such as yield, resistance to diseases and pests, grain quality, etc. in cereal crops.",signatures:"Fakhriddin N. Kushanov, Ozod S. Turaev, Oybek A. Muhammadiyev, Ramziddin F. Umarov, Nargiza M. Rakhimova and Noilabonu N. Mamadaliyeva",downloadPdfUrl:"/chapter/pdf-download/81949",previewPdfUrl:"/chapter/pdf-preview/81949",authors:[{id:"292067",title:"D.Sc.",name:"Fakhriddin N.",surname:"Kushanov",slug:"fakhriddin-n.-kushanov",fullName:"Fakhriddin N. Kushanov"},{id:"302522",title:"Ph.D.",name:"Ozod",surname:"Turaev",slug:"ozod-turaev",fullName:"Ozod Turaev"},{id:"459772",title:"MSc.",name:"Oybek A.",surname:"Muhammadiyev",slug:"oybek-a.-muhammadiyev",fullName:"Oybek A. Muhammadiyev"},{id:"459841",title:"MSc.",name:"Nargiza M.",surname:"Rakhimova",slug:"nargiza-m.-rakhimova",fullName:"Nargiza M. Rakhimova"},{id:"459842",title:"MSc.",name:"Ramziddin F.",surname:"Umarov",slug:"ramziddin-f.-umarov",fullName:"Ramziddin F. Umarov"},{id:"459843",title:"MSc.",name:"Noilabonu N.",surname:"Mamadaliyeva",slug:"noilabonu-n.-mamadaliyeva",fullName:"Noilabonu N. Mamadaliyeva"}],corrections:null},{id:"77917",title:"Cotton as a Model for Polyploidy and Fiber Development Study",doi:"10.5772/intechopen.99568",slug:"cotton-as-a-model-for-polyploidy-and-fiber-development-study",totalDownloads:152,totalCrossrefCites:1,totalDimensionsCites:1,hasAltmetrics:0,abstract:"Cotton is one of the most important crops in the world. The Gossypium genus is represented by 50 species, divided into two levels of ploidy: diploid (2n = 26) and tetraploid (2n = 52). This diversity of Gossypium species provides an ideal model for studying the evolution and domestication of polyploids. In this regard, studies of the origin and evolution of polyploid cotton species are crucial for understanding the ways and mechanisms of gene and genome evolution. In addition, studies of polyploidization of the cotton genome will allow to more accurately determine the localization of QTLs that determine fiber quality. In addition, due to the fact that cotton fibers are single trichomes originating from epidermal cells, they are one of the most favorable model systems for studying the molecular mechanisms of regulation of cell and cell wall elongation, as well as cellulose biosynthesis.",signatures:"Venera S. Kamburova, Ilkhom B. Salakhutdinov, Shukhrat E. Shermatov, Zabardast T. Buriev and Ibrokhim Y. Abdurakhmonov",downloadPdfUrl:"/chapter/pdf-download/77917",previewPdfUrl:"/chapter/pdf-preview/77917",authors:[{id:"213344",title:"Prof.",name:"Ibrokhim Y.",surname:"Abdurakhmonov",slug:"ibrokhim-y.-abdurakhmonov",fullName:"Ibrokhim Y. Abdurakhmonov"},{id:"187106",title:"Dr.",name:"Zabardast T.",surname:"Buriev",slug:"zabardast-t.-buriev",fullName:"Zabardast T. Buriev"},{id:"213348",title:"Dr.",name:"Shukhrat E.",surname:"Shermatov",slug:"shukhrat-e.-shermatov",fullName:"Shukhrat E. Shermatov"},{id:"328107",title:"Dr.",name:"Ilkhom B.",surname:"Salakhutdinov",slug:"ilkhom-b.-salakhutdinov",fullName:"Ilkhom B. Salakhutdinov"},{id:"328140",title:"Dr.",name:"Venera S.",surname:"Kamburova",slug:"venera-s.-kamburova",fullName:"Venera S. Kamburova"}],corrections:null},{id:"78282",title:"Soybean as a Model Crop to Study Plant Oil Genes: Mutations in FAD2 Gene Family",doi:"10.5772/intechopen.99752",slug:"soybean-as-a-model-crop-to-study-plant-oil-genes-mutations-in-fad2-gene-family",totalDownloads:129,totalCrossrefCites:1,totalDimensionsCites:1,hasAltmetrics:0,abstract:"Plants have numerous fatty acid desaturase (FAD) enzymes regulating the unsaturation of fatty acids, which are encoded by a FAD gene family. The FAD2 genes belong to such family and play a vital role in converting monounsaturated oleic acid to polyunsaturated linoleic acid. Oleic acid has the health benefits for humans, such as reduction in cholesterol level, antioxidation property, and industrial benefits like longer shelf life. The development of genotypes with high oleic acid content in seeds has become one of the primary goals in breeding oilseed plants. The identification and characterization of the FAD2 genes in plants have been an important step to better manipulate gene expression to improve the seed oil quality. The induction of mutations in FAD2 genes to reduce FAD2 enzyme activity has been an integral approach to generate genotypes with high oleic acid. This chapter will describe the FAD2 gene family in the model organism soybean and the correction of mutations in FAD2 genes with the increase of oleic acid content. Leveraging advanced research of FAD2 gene family in soybean promotes the study of FAD2 genes in other legume species, including peanut. The future perspectives and challenges associated with mutations in FAD2 genes will be discussed.",signatures:"Sy M. Traore and Guohao He",downloadPdfUrl:"/chapter/pdf-download/78282",previewPdfUrl:"/chapter/pdf-preview/78282",authors:[{id:"420094",title:"Prof.",name:"Guohao",surname:"He",slug:"guohao-he",fullName:"Guohao He"},{id:"428855",title:"Dr.",name:"Sy M.",surname:"Traore",slug:"sy-m.-traore",fullName:"Sy M. Traore"}],corrections:null}],productType:{id:"1",title:"Edited Volume",chapterContentType:"chapter",authoredCaption:"Edited by"},subseries:null,tags:null},relatedBooks:[{type:"book",id:"880",title:"Plant Breeding",subtitle:null,isOpenForSubmission:!1,hash:"00fb30196097697f0e1211ce27ba426d",slug:"plant-breeding",bookSignature:"Ibrokhim Y. Abdurakhmonov",coverURL:"https://cdn.intechopen.com/books/images_new/880.jpg",editedByType:"Edited by",editors:[{id:"213344",title:"Prof.",name:"Ibrokhim Y.",surname:"Abdurakhmonov",slug:"ibrokhim-y.-abdurakhmonov",fullName:"Ibrokhim Y. 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",isbn:"978-1-83768-439-7",printIsbn:"978-1-83768-438-0",pdfIsbn:"978-1-83768-440-3",doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!0,isSalesforceBook:!1,isNomenclature:!1,hash:"05102f198619f7cc1312eb39a352026c",bookSignature:"Associate Prof. Selim Sözen",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/12354.jpg",keywords:"Major Abdominal Trauma, Solid Organ Trauma, Bones Fracture, Pelvic Fracture, Spine Trauma, Sports Trauma, Lung Trauma, Facial Trauma, Orbital Trauma, Neck Trauma, Fractures, Brain Trauma",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:null,numberOfDimensionsCitations:null,numberOfTotalCitations:null,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"June 17th 2022",dateEndSecondStepPublish:"July 15th 2022",dateEndThirdStepPublish:"September 13th 2022",dateEndFourthStepPublish:"December 2nd 2022",dateEndFifthStepPublish:"January 31st 2023",dateConfirmationOfParticipation:null,remainingDaysToSecondStep:"17 days",secondStepPassed:!1,areRegistrationsClosed:!1,currentStepOfPublishingProcess:2,editedByType:null,kuFlag:!1,biosketch:"Dr. Selim Sözen is an expert in general surgery who received his medical degree from Ondokuz Mayıs University, Turkey. From 1999 to 2004, he was an assistant doctor at Ankara Atatürk Education and Research Hospital. He joined the Department of General Surgery, Medicine Faculty, Namık Kemal University, He is a member of the Turkish Surgical Association and a review board member for several journals. He has published 109 articles in scientific journals and presented 64 poster papers at scientific congresses.",coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"90616",title:"Associate Prof.",name:"Selim",middleName:null,surname:"Sözen",slug:"selim-sozen",fullName:"Selim Sözen",profilePictureURL:"https://mts.intechopen.com/storage/users/90616/images/system/90616.png",biography:"Dr. Selim Sözen is an expert in general surgery who received his medical degree from Ondokuz Mayıs University, Turkey, in 1998. From 1999 to 2004, he was an assistant doctor at Ankara Atatürk Education and Research Hospital, Turkey. From 2004 to 2013, he worked as a specialist at different government hospitals in Turkey. He joined the Department of General Surgery, Medicine Faculty, Namık Kemal University, Turkey, as an associate professor in 2013. He completed liver transplantation surgery at İnönü University, Turkey, in 2014–2015. Since 2016, Dr. Sözen has run his own surgery clinic in İstanbul, Turkey. He is a member of the Turkish Surgical Association and a review board member for several journals. He has published 109 articles in scientific journals and presented 64 poster papers at scientific congresses. 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Immunoglobulin G (IgG) is the most common. Eighty percent of all the antibodies present in the blood are IgG [2]. IgG is a relatively large molecule (approx. 150 kDa). It has four subclasses, which are IgG1, IgG2, IgG3, and IgG4 [3]. Monoclonal antibodies are antibodies derived from one unique B cell clone [4]. They have single antigenic determinant specificities [5]. Monoclonal antibodies are screened and isolated by special procedures, expressed, and purified [6]. Monoclonal antibodies, particularly IgG1, have tremendous application in biotherapeutics. Other subclasses such as IgG2 and IgG4 are also used as biotherapeutic, and interest in these two antibody classes is also increasing. As for today, IgG1 comprise most of the mAb biotherapeutic drugs in the market. Twenty-nine new mAbs are presently undergoing late-stage clinical trials, including human and humanized IgG1, IgG2, and IgG4 molecules [7]. Few IgG2 and IgG4 drugs are already available such as OKT3 (Muronomab-CD3), a murine IgG2a drug from Johnson & Johnson (1986), Bexxar (Tositumomab-I-131), and a murine IgG2a drug radiolabeled with I-131 from Corixa/GSK (2003). IgG4 antibodies are evolving as an important class of cancer immunotherapies [8].
\nSize exclusion chromatography (SEC) is a powerful analytical tool for the separation of monoclonal antibodies and other proteins [9]. SEC, as a strategy for the isolation and purification of antibodies, is not new; in 1989, high-resolution Superose 6 HR 10/30 fast protein liquid chromatography (FPLC) columns were used. [10]. Since then, many researchers used SEC for the purification of antibodies. The literature search for the number of “publications on the purification of monoclonal antibodies by size exclusion chromatography” [11] shows that between 1983 and 2003, there was a surge of research in this regard (Figure 1).
\nNumber of publications dealing with size exclusion chromatography over the years.
For the large-scale purification of monoclonal antibody biotherapeutics, Protein A is commonly used as the primary capture step. Following the use of Protein A chromatography, SEC is used to characterize the Protein A purified fractions. Size exclusion chromatography (SEC) is primarily used for the separation in analytical HPLC and for routine quality control analysis of the mAb. Detection wavelength of 280 nm is commonly used. SEC combined with multi-angle light scattering is one method for the characterization of the molar mass distribution of mAb, ADC, and other biomolecules [12].
\nIn this chapter, certain aspects of size exclusion chromatography and its use in the analytical purification or separation of monoclonal antibodies are discussed. Secondary interactions, effects of particle size and pore size on the SEC separation, particularly in resolving monomer peak from dimer and fragment peaks with a better resolution are also discussed. Selection of the right SEC column is critical to achieve the goal of separation. The calibration curve has a very important role in this regard. Since there are different kinds of proteins differing in structure and shape, many vendors provide calibration curves using globular protein standards, branched standards, and linear standards, so that the separation range can ideally be interpreted under the chromatographic conditions.
\nReproducible separation of a monomer from its dimer and other high molecular weight (HMW) impurities, fragments, and other low molecular weight (LMW) impurities is needed during the purification of the mAb biotherapeutic. Optimized particle chemistry and packing of SEC column help in this regard. mAb analysis using a mobile phase containing an appropriate amount of selected organic solvents such as isopropyl alcohol (IPA) may be needed to prevent alteration of peak retention time, poor peak shape, or resolution. Few examples are shown to elaborate this. Digestion with papain or pepsin is commonly applied to obtain antibody fragments without the loss of activity—this is discussed in the context of selecting right SEC columns from a variety of particle sizes, pore sizes, and dimensions. Forced degradation studies are needed to assess the stability of the protein, to understand the mechanism of degradation by oxidation, heat, light, or hydrolysis. Forced degradation study by SEC is separately discussed in Section 2.7. Interest in the accurate molecular weight analysis of intact monoclonal antibody IgG1 by SEC using MS-friendly mobile phases is increasing, and it is discussed in Section 4. Section 5 focuses mostly on the use of chromatographic methods which are orthogonal or complimentary size exclusion and useful to detect the protein heterogeneity. The use of ultra HPLC is needed for fast separation, and many methods already developed in HPLC need to be easily transferred to UHPLC. Section 6 briefly discusses the usefulness of a SEC column compatible to both HPLC and UHPLC instruments by easy method transfer. Section 7 is about desalting of mAb or any biopolymer solution, not by dialysis membrane or spin column but by using an analytical SEC HPLC column. The use of SEC column in hydrophilic liquid chromatography (HILIC) mode is interesting and can separate the nucleobases (Section 8). This chapter ends with few remarks about the tips and tricks for size exclusion chromatography.
\nSize exclusion chromatography uses a molecular sieving retention mechanism [13], based on differences in the hydrodynamic radii or differences in size of analytes such as proteins. Large sample molecules cannot penetrate or only partially penetrate the pores of the stationary phase. So, the larger molecules elute first and smaller molecules elute later, the order of elution being a function of the size.
\nSEC is the only mode of chromatography where theoretically there is no interaction of the analyte with a stationary phase. The whole process of partitioning or separating the different molecular species is due to the entropy factor and not due to adsorption, ΔH being equal to zero. Pure Silica particles are most commonly used as base material in this type of chromatography of biomolecules. Since the separation of the biomolecule by SEC will depend on its hydrodynamic radii, two proteins of the same molecular weight (such as 70 kDa) may elute at two different retention times, if there is a difference in their hydrodynamic radii (Figure 2). So, any factor at any stage of purification, affecting the shape of the protein, will affect the elution volume or retention time.
\nComparison of globular and rod-shaped proteins.
Unless otherwise mentioned, SEC analyses discussed in this chapter were carried out using the mobile phase 100 mM KH2PO4/Na2HPO4, pH 6.7, 100 mM Na2SO4, 0.05% NaN3. Agilent 1100, Agilent 1200 HPLC and Thermo Ultimate 3000 UHPLC systems and associated software were used for integration and peak analysis. Sodium azide (NaN3) was used as an antibacterial agent to prevent fouling of the phosphate buffer. Detection wavelength was 280 nm unless otherwise mentioned. Flow rates and injection volume of the sample are varied as needed. Reproducibility for calculation of % relative standard deviation (RSD) was based on 10 consecutive injections. Linearity of both monomer and dimer during loading study was calculated based on peak areas versus total material loaded during injection. Please refer to individual chromatograms for the respective chromatographic conditions.
\nAs mentioned earlier, among all modes of chromatography, it is only during the size exclusion chromatography where the analyst does not demonstrate any kind of interaction with the stationary phase. During all other chromatographic modes, an analyst demonstrates some kind of interaction between the protein and the stationary phase, followed by the elution using a stronger solvent. This is because SEC is purely a separation technique based on the permeability of the protein through the pores. Any factor which will prevent the free passage through the pores will result into a nonideal SEC condition. These interactions between proteins and the stationary phases are called secondary interactions. The secondary interactions are based on charge-related interactions or hydrophobicity-related interactions. The secondary interaction coming from the free silanol groups present on the silica surface of the stationary phase is often protected by a diol-bonded coating on the stationary phase which shields the silica surface from such action [14].
\nIn a real-world situation, any secondary interaction between the stationary phase and the proteins including monoclonal antibodies needs to be taken care of by selecting the right column where the stationary phase is effectively coated, and free silanol groups are inaccessible to proteins. Further method development by optimizing the chromatographic conditions may be needed to get the best separation. Ionic and hydrophobic interactions between the sample and the column packing material can be avoided by controlling the ionic strength. A general rule of thumb is that low ionic strength (<0.1 M) may induce charge-related secondary interactions and high ionic strength (>1.0 M) may lead to hydrophobicity-related secondary interactions, while the concept of low or high concentration may vary from mAb to mAb depending on the nature of the individual one. For each protein sample, there will be an optimal buffer type and salt concentration for the best separation that results in the highest resolution and recovery. This can be found out by trial and error approach only. If a sticky protein comes into contact with the stationary phase (dotted line in the figure below), it may undergo a conformational change; the binding constant of the conformationally changed protein can be so strong that it would not elute out of the column (Figure 3). The use of additives may be needed. Arginine prevents binding to the surface [15]. Similarly, a number of other additives may also be used to minimize the secondary interactions. The use of isopropyl alcohol (IPA) as another additive to avoid nonspecific interaction and the reproducibility of the analysis is somewhat discussed in the Section 2.5.
\nBinding of a protein to the stationary phase and the influence of arginine.
There is no universal protocol for working with additives to avoid nonspecific interactions for all proteins. Chromatographic conditions preventing secondary interactions need to be found out by trial-and-error type of experiments with a variety of additives.
\nPermeation of large molecules into pores depends on the pore size.
Relation between the particle and pore characteristics to the resolution and other peak parameters.
It is important to compromise between the particle sizes and pore volume in order to get the desired separation. Pore characteristics of the SEC column need to be optimized to have a high resolution of a mAb monomer from a dimer and higher order aggregates, as well as from fragments. For large biomolecules, such as therapeutic proteins and monoclonal antibodies (mAbs), a larger exclusion limit will yield better separation particularly of the dimer and higher order aggregates from the monomer.
\nIn the absence of any secondary or non-SEC retention mechanism, the calibration yields an S-shaped curve containing a linear portion in between the total exclusion and total inclusion limits. In Figure 6 [17], the calibration curve was generated using three different types of standards (globular, branched, and linear).
\nElution profile of mAb IgG obtained with TSKgel G2000SW, TSKgelG3000W, or TSKgel G4000SW columns.
The red line in Figure 6 represents the average molecular weight of the monoclonal antibody (mAb) IgG (150 kDa). If we follow the calibration curve generated by globular proteins (•), mAb is eluting very close to the total exclusion limit of TSKgel G2000SW, while it is eluting very close to the total inclusion limit of TSKgel G4000SW, resulting into poor separation of the monomer from its impurities in both the cases. But in the middle panel, as seen in the case of TSKgel G3000W, the monomer is eluting around in the middle of the linear range of the calibration curve, so the monomer peak can clearly be separated from its dimer and higher order aggregates and fragment impurities way better than with the other two columns. It is the pore volume between total exclusion and total inclusion volume which is important. The greater the pore volume per unit column volume, the better the separation. In other words, the shallower the calibration curve, the better the separation. All SEC columns with the same dimension, particle size, and pore size from different vendors otherwise may look identical, but the pore volume per unit column volume may not be the same. An analyst can gain advantage by selecting a column with larger pore volume per unit column volume.
\nDifferent SEC columns obtained from different vendors, having the same dimensions and particle size as labeled on the individual columns, may apparently look alike. The difference is in the pore volume per unit column volume due to the differences in particle size and pore size distributions, packing quality, and so on. In general, the better the pore volume per unit column volume, the better the separation and resolution of proteins from a SEC column. This is a very critical criterion when selecting a SEC column for better separation.
\nA chromatographer uses SEC primarily to separate monoclonal antibodies from its impurities or heterogeneities. The monomer IgG1 peak (150 kDa) needs to be purified from its dimer, trimer, or higher order aggregates popularly known as high molecular weight (HMW) species and the fragments which are known as the low molecular weight (LMW) species. SEC is widely accepted as a work horse for routine quality control with its purpose to monitor these HMW and LMW impurities from a mAb monomer. Protein aggregation of biotherapeutics is a common issue. Even a very small amount of aggregates may cause an immunogenic response in the human body and needs to be removed from the monoclonal antibody monomer. More than 99% purity is needed for medicinal purpose [17]. The formulation containing the pure monomer monoclonal antibody needs to be monitored for its stability. The purified protein in the formulation may also undergo aggregation over time. Protein aggregation can happen at any stage during expression and purification. Temperature, pH, ionic strength, concentration and many other factors can give rise to protein instability, leading to aggregation. Effect of ions on protein agitation and temperature-induced aggregation of mAbs has been reported in the literature [18]. Specific racemization of Heavy-Chain Cysteine-220 in the hinge region is identified as a possible cause of degradation during storage. Increased hydrophobicity can increase the likelihood of aggregate formation during manufacturing and storage [19]. Freeze-thaw cycles too can cause an aggregation of an already purified protein [20]. Formation of aggregates may be induced by light as well [21]. That is the reason why all registration applications for new molecular entities and associated drug products require photo-stability data [22]. Increased resolution between the monomer peak and other HMW and LMW impurities is the key for the purification of monoclonal antibodies. SEC can monitor the stability during storage. The removal of HMW and LMW impurities is equally important for application of newly emerging biotherapeutics such as ADCs, bi-specific antibodies, biosimilars, and bio betters as well. As an example from the recent literature report, aggregate and fragment levels were determined by SEC-HPLC for the characterization of bi-specific antibodies [23]. A single chain variable fragment (scFv), composed of the variable regions of the heavy chain (VH) and the light chain (VL), is gaining interest too as it retains the specificity of the original IgG. The scFv format is often used, but one problem that cannot be easily solved by purification is the fact that hybridomas can secrete different monoclonal antibodies [24]. Literature reports a few interesting articles about the concept of mAbs being a perfectly defined entity to researchers. The following is an excerpt from the commentary as shown here [25]. “most researchers consider monoclonal antibodies to be perfectly defined reagents with single specificities” but “Hybridomas frequently secrete more than one light and/or heavy chain.” So “the problem is probably best summarized thus: antibodies sold as different are often identical, while antibodies sold as identical are often different (thanks to Natalie de Souza (editor Nature Methods) for this pithy insightful observation), and the customer does not know which is which.” In another interesting article, the authors have discussed that “two kappa immunoglobulin light chains are secreted by an anti-DNA hybridoma” and its implications for isotypic exclusion are discussed [26]. Biotherapeutic scientist needs to be aware of these facts while analyzing mAb. The separation of scFvs from dimers and aggregates is also important; most of the affinity purified scFv fractions are monitored by SEC.
\nDifferent mAbs may have different amount of impurities under native conditions. Representative chromatograms of the separation of four different monoclonal antibodies (IgG1) at 0.75 mL/min using a 4 μm; 4.6 mm ID × 15 cm TSKgel SuperSW mAb HTP column are shown in Figure 7 [27]. This analysis clearly showed that under native conditions different monoclonal IgG1 antibodies had different extent of HMW and LMW species based on the individual % peak area analysis (data not shown here). The monomer peak eluted as fast as in 2 min.
\nSeparation of four different mAbs on a 4 μm; 4.6 mm ID × 15 cm TSKgel SuperSW mAb HTP column.
A chromatographer needs to choose a suitable column based on the separation criteria the chromatographer is looking for. Comparison of the analysis of mAb aggregates using 15 and 30 cm long TSKgel UP-SW3000, 2 μm columns using the same mobile phase and flow rate is shown below [28].
\nFast separation of the HMW and LMW species is important. The effect of the column length should be taken into account when selecting a column in this regard. The results indicate that the TSKgel UP-SW3000 column with a shorter length yielded a similar profile to the 30-cm column with 50% less run time and 50% lower backpressure at a typical flow rate of 0.35 mL/min (Figure 8). The resolution between dimer and monomer is still maintained within the acceptable range. Thus, a shorter column could be successfully used for the separation of the dimer and monomer, reducing the overall runtime by half but the resolution of the fragment on the LMW side of the monomer slightly decreased. The longer column yielded a better resolution. As long as the resolution is 1.5 and above yielding a baseline resolution of the two species, the method may remain acceptable. The selection of the correct length of the column should be based on the goal of the separation. The 15-cm column operated at the typical flow rate of 0.35 mL/min yielded a backpressure of 11 Mpa, which was well within its maximum operable pressure and thus could be used in both HPLC and UHPLC systems.
\nEffect of protein separation on the column length of SEC columns.
Factors that play a role in selecting a suitable HPLC column.
Two different sources of silica can be a factor in lot-to-lot reproducibility. Vendors always maintain a strict quality control passing criteria if the silica source is different.
Bonding chemistry developed on the same silica at different times can also be a factor in lot-to-lot reproducibility.
A robust method with a good SEC analytical column is critical for the analyst. A protein standard mixture can be used to confirm the lot-to-lot reproducibility of an SEC column. The protein standards are chosen to cover the whole range of calibration curve from total exclusion limit to total inclusion limit. A representative chromatogram of the analysis of a protein standard mixture using a TSKgel G3000SWXL, 5 μm, 7.8 mm ID ×30 cm column is shown in Figure 9. Thyroglobulin (700 kDa) is close to the total exclusion limit and para-amino-benzoic acid (PABA–137 Da) is near the total inclusion limit. Chromatographers use also vitamin B12 (1.4 kDa) in place of PABA. Vendors generally pass the packed SEC columns using a protein standard mixture and establish a QC pass criteria. For example, the specification for TSKgel G3000SWXL column passing QC is N (PABA) > 20,000 and As (PABA) = (0.7–1.6) where N represents the number of theoretical plates and As represents the peak asymmetry.
\nRetention time of five different proteins analyzed with a TSKgel G3000SWXL column was high reproducible. An overlay of five injections is shown.
Figure 9 [31] shows the reproducibility of five consecutive injections with low %RSD of all the peak parameters (data not shown here).
\nReproducibility of the analysis of a protein standard mixture with low %RSD of the peak parameters such as retention time, peak area, peak asymmetry and number of theoretical plates and passing the vendor-defined QC criteria can be used for monitoring the column health. Routine users purifying a particular mAb with a particualr SEC column may use the same mAb as their internal standard to monitor the column quality and lifetime over a number of injections. Nowadays, mAb standards from USP and NIST are available for similar purpose.
\nReproducibility of 15 consecutive injections during the analysis of a USP mAb using a 15-cm TSKgel UP-SW3000 column at 0.5 mL/min flow rate and phosphate buffer at pH 6.7 is shown in Figure 10.
\nReproducibility of 15 consecutive analytical injections of a USP mAb using a 15-cm TSKgel UP-SW3000 column at phosphate buffer pH 6.7. The overlay of 15 injections is shown.
The mAb monomer peak eluted at 2.717 min with good resolution between monomer and the dimer peaks as well as the fragments. Similar reproducibility is noticed in case of pH 6.2 (data not shown here).
\nReproducibility in the analysis of the USP mAb in pH 6.2 conditions with 250 mM KCl at 0.3 mL/min using a 30 cm column is shown below—the overlay of the 15 consecutive injections demonstrated consistency (Figure 11) (all USP Reference Standards are provided as delivered and specified by the US Pharmacopeia). The monomer peak elution time was 8.367 min. Similar reproducibility was obtained using a 15-cm column at pH 6.7 (data not shown here).
\nReproducibility of 15 consecutive analytical injections of a USP mAb using a 30-cm TSKgel UP-SW3000 column at pH 6.2. The overlay of 15 injections is shown.
Nonspecific absorption of antibodies onto the column gel matrix poses a challenge, and some newly engineered antibodies possess a high degree of hydrophobicity. The use of organic solvents such as isopropyl alcohol (IPA) or salts can decrease this interaction as reported by many scientists. However, the additives may alter the diffusion of these molecules, which results in retention time shift and poor peak resolution that did not occur in a typical aqueous buffer system, such as sodium phosphate buffer at neutral pH. The Figure 12 and Table 2 show that a TSKgel UP-SW3000, 2 μm SEC column was used for analyzing monoclonal antibodies (mAbs) with the addition of 15% IPA in sodium phosphate buffer, pH 6.7. As demonstrated, peak resolution and retention time shift were not impacted.
\nReproducibility of 14 consecutive analytical injections of a USP mAb using a 30-cm TSKgel UP-SW3000, 2 μm SEC column at pH 6.7 with isopropyl alcohol. The overlay of 14 injections is shown.
Retention time and peak areas of the monomer and dimer peak (Figure 12).
Similarly, Figure 13 shows the overlay of the 15 consecutive injections of USP mAb at pH 6.7 with 15% IPA using a 30-cm column (Figure 13 and Table 3). Improvement in the baseline was noticed after the first two injections. Overall, the analysis yielded excellent reproducibility. The monomer peak elution time was 8.338 min. As expected, there was no considerable difference here compared to the retention time obtained earlier at pH 6.2. The presence of IPA as additive obviously will yield a higher back pressure, and so long as the column is operated within its maximum operable pressure, this should not be an issue. The retention times of monomer, dimer, aggregates, and fragment peaks are nearly unchanged. Peak width and peak shape are very consistent from injection to injection. The baseline of the first injection (as shown in blue) indicate that the column takes only 1–2 injections to be stabilized. After that, all subsequent injections are overlaid perfectly.
\n(A) Reproducibility of 15 consecutive analytical injections of a USP mAb using a 30-cm TSKgel UP-SW3000, 2 μm SEC column at pH 6.7 with 15% IPA. The overlay of 15 injections is shown. (B) Chromatogram of 13 a zoomed in.
Retention time and peak areas of the monomer and dimer peak (Figure 13).
The overlay indicates the similarities of peak retention times, peak width and peak height of dimer, monomer, aggregates and fragment peaks between the two different conditions. An appropriate percentage of organic solvent such as isopropyl alcohol (IPA) did not alter the diffusion of mAb molecules using a TSKgel UP-SW3000 column. As demonstrated, this column can be successfully operated with the addition of 15% IPA. Data indicate that the column’s particle chemistry and packing are optimized so that with the addition of an appropriate amount of selected organic solvents, there is no alteration of peak retention time or poor peak resolution [32].
\nSample load, in both volumetric load and absolute load, may affect SEC separation. Mass overload takes place when sample molecules no longer have free access to diffuse into and out of the pores, thus bypassing part of the column and thereby effectively reducing the length of the column that remains to fractionate the sample. If the height equivalent theoretical plates (HETP) are plotted against the load amount, the HETP values should remain constant as long as the column efficiency is not compromised. The loading capacity is the maximum load beyond which the HETP value starts increasing, as shown in Figure 14 [33].
\nInfluence of sample load on height equivalent theoretical plates (HEPT) using three different columns (G2000SW, G3000SW, G4000SW).
To obtain the capacity of an analytical SEC column as often the chromatographers like to do, a loading study plot HETP vs. load amount is shown above (Figure 14). The loading capacity of a SEC column with defined dimensions depends on the sample. The loading capacity can be increased by increasing the column length or diameter. Increasing column length also increases resolution and retention time, leading to an additional separation time and amount of mobile phase.
\nBelow is the loading study of γ-globulin (150 kDa) using the TSKgel UP-SW3000 column. It is necessary to know the experimental range of loading where the retention time, peak shape, separation efficiency, etc., remain nearly unchanged over varying load concentrations. Please note that when a loading study is carried out, both volumetric loading and absolute loading amounts should be studied. In Figure 15, a volumetric loading study is shown. In any SEC analysis, by theory, the total volume injected should not be more than 3% of the column volume to avoid the effect of band broadening.
\n(A) Influence of peak resolution and retention time on the amount of γ-globulin loaded on a TSKgel UP-SW3000 column; (B) monomer retention time in dependence of the amount of sample.
Figure 15 shows that even at larger volumetric load containing up to 160 μg proteins, the monomer peak remains well resolved from its dimer. Retention time is remaining constant over the experimental range. Excellent linearity of both monomer peak areas and dimer peak areas versus total load were obtained (data not shown here). So if the primary interest of the analyst is to separate the monomer from the dimer, 160 μg loading in 40 μL volume can be used. Now if the monoclonal antibody concentration can be increased so that 160 μg can be loaded in lower volume (e.g., 10 μL), then the peak shape can further be improved, if needed.
\nPeak shape and efficiency were not affected when injecting 400 μg of a monoclonal antibody preparation during a loading study of a monoclonal antibody using a TSKgel G3000SWXL, 5 μm, 7.8 mm ID × 30 cm column (Figure 16). A 10-fold increase in total protein content did not affect the retention time, peak symmetry, or separation efficiency of the column [30]. Similar study using a TSKgel G2000SWXL, 5 μm, 7.8 mm ID × 30 cm analytical column even with a high load of Bovine Serum Albumin also yielded a well-resolved peak without any splitting [34].
\nInfluence of mAb loading on retention time, peak symmetry, or separation efficiency using a TSKgel G3000SWXL, 5 μm, 7.8 mm ID × 30 cm column.
As mentioned earlier, having an idea about the sample loading capacity will provide analyst the knowledge about the load range within which the desired sensitivity and resolution can be achieved. The loading study can be extended to a lower range of detection to get the limit of detection (LOD) and limit of quantitation (LOQ) values for the column in the analysis of mAbs (data not shown here). Similarly, aggregation pattern of mAbs as a function of concentration can be monitored using SEC columns if a mAb is susceptible to aggregation at higher concentration.
\nIgG is a relatively large molecule (approx. 150 kDa), and in order to improve the penetration to the tissue, fragmentation is carried out. Digestion with papain or pepsin is commonly applied to obtain antibody fragments without the loss of activity. When papain is used for the antibody digestion, 2 Fab (50 kDa each) and 1 Fc (50 kDa) are obtained from one antibody (Figure 17).
\nCleavage of an IgG with papain or pepsin.
When pepsin is used, a F(ab’)2 is obtained. SEC can be used to analyze the separation of these fragments. The scope of this analysis by SEC is taken as an opportunity to explain how to select a column with right particle size, pore size, column dimensions, and so on.
\nIn Figures 18 and 19, a set of four different SEC columns are compared during the separation of papain digestion products of a mAb to explain how to select the right SEC column for the right purpose [35].
\nCharacteristics of the SEC columns used in the analysis of
Separation of papain digestion products of a mAb using a set of four different SEC columns. Columns in panel A, B, and D are of 250 Å pore sizes, while column in panel D is of 300 Å pore size [
For analyzing monoclonal antibody and other biopolymers 250 Å pore size, 5 μm, 7.8 mm ID × 30 cm SEC columns are widely considered. For example, TSKgel G3000SWxl columns (Panel D) have a separation range for globular protein samples up to 500 kDa. The other SEC column is a 4-μm, 7.8 mm ID × 30 cm TSKgel SuperSW mAb HR SEC column (Panel B). It is smaller than the conventional 5 μm TSKgel G3000SWXL column. Smaller particle size and the optimized packing are expected to yield high-resolution analysis of mAb monomers, dimers, and fragments due to shallow calibration slope at the corresponding molar mass region of the mAb monomer (150 kDa). Monomer – dimer resolution increased from 1.63 to 2.02. Another SEC column (Panel A) is a 4-μm TSKgel SuperSW mAb HTP column which is smaller in dimensions, length, and ID (4.6 mm ID × 15 cm). This column offers high throughput analysis, separating the dimer and monomer in half the run time compared to all the other three columns in panels B, C, and D. Results are similar to the analysis of mAbs with a 5-μm conventional column (panel D). The fourth column (Panel C) discussed here is of even smaller particle size (3 μm) with higher molar mass exclusion limit (2500 kDa, globular proteins) than all other three columns (500 kDa, globular proteins). Due to the higher exclusion limit, this TSKgel UltraSW aggregate column (Panel C) is expected to yield higher resolution of mAb multimers and aggregates. Please see further discussion about this in Section 2.7.
\nStability of the biotherapeutic proteins in formulation is very critical for candidate selection, characterization of the biotherapeutic, formulation and assay development, and so on. A forced degradation study, popularly known as stress testing, is considerably a faster way to monitor the stability of the therapeutic proteins. Stress is provided by increasing the temperature, changing the pH or a combination of both. Depending on the nature, individual mAbs can be susceptible to light, freeze–thaw conditions, mechanical stress, oxidation and so on. So, monitoring the stability from time to time is important. Literature reports: “Whereas stability-testing requirements are defined in regulatory guidelines, standard procedures for forced degradation of therapeutic proteins are largely unavailable, except for photo stability” [36]. Stress conditions induce unfolding of the native protein structure. As a result, the exposed hydrophobic patches may be able to interact with each other, leading to aggregation [37]. Protein degradation can also be measured using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE), and the protein structure can be characterized by other techniques using Fourier transform infrared spectrometry and circular dichroism
Analysis of forced degradation of IgG after acid and heat denaturation using a SEC TSKgel UltraSW aggregate, 3 μm, 30 nm, 7.8 mm ID × 30 cm column.
The analysis of a heat denatured, large hydrophobic metalloprotein, Apo ferritin, analyzed using a TSKgel UltraSW Aggregate column yielded high resolution between the monomer (450 kDa) and dimer (900 kDa) (Figure 21). The trimer (1350 kDa), tetramer (1800 kDa), and higher order aggregates of Apo ferritin were well separated. Tetramer of Apo ferritin is approximately equivalent to 13 mer of a mAb. Larger exclusion limit yielded better resolution of the higher order aggregates. [40].
\nAnalysis of Apo ferritin after heat denaturation using a TSKgel UltraSW aggregate column.
The use of organic solvents such as isopropyl alcohol (IPA) or salts can decrease the secondary interaction as reported by many scientists and mentioned earlier briefly in this chapter in Section 2.5. Peak resolution and retention time shift were not impacted with the use of 15% IPA as demonstrated in the two examples (Figure 12 and Figure 13). It is necessary to evaluate the individual SEC column regarding the impact of the additives since a stationary phase with minimum impact is always a favorable choice. After important mobile phase parameters have been set, such as pH, stationary phase, and ionic strength, significant improvements can, in fact, be made to separate mAb monomers from aggregates and fragments. There are no universal additives which can be applied for every mAb or protein purification. Other common additives are methanol and ethanol. Use of sodium perchlorate may also improve the separation and resolution. By switching from 0.2 mol/L sodium chloride to 0.2 mol/L of the more chaotropic sodium perchlorate salt, together with a twofold reduction in the buffer concentration, less peak tailing and distinct peaks for the dimer and trimer could be noticed [41]. Sodium dodecyl sulfate (SDS), urea, guanidine hydrochloride, etc., are sometimes used when the proteins need to be solubilized, leading to denaturation of the protein and breakage of the noncovalent bonds. The use of additives should be considered only when needed. In many cases, the performance of the column is irreversibly changed when a column is subjected to particular additives. Retention time may shift when the additive is added and the resolution may change, expectedly to a better resolution but these values should remain constant and the analysis should be reproducible. It is always better to dedicate the SEC column if the column is subjected to additives, since we still have no clear idea how the pore characteristic of the stationary phase may behave with and without additives. Analysts should check the operational and conditions (OCS) sheet for the column as provided by the vendor to make sure that additives are compatible with the stationary phase. If compatible the analyst should be aware of the percentage of organic solvent the column is compatible with. Generally, when the organic solvent is used, the column may need a slower ramping rate for proper equilibration of the column with the mobile phase containing the additive, generally by using gradual solvent changes using a shallow gradient at low flow rate.
\nThe use of mass spectrometry is becoming increasingly popular for scientists dealing with biomolecule separation to identify the individual peaks by molecular weight. The liquid chromatography-MS (LC–MS) system available nowadays is very robust and useful for routine mass determination. Reversed phase LC–MS or SEC-MS using organic solvents such as acetonitrile can be used for the mass spectrometric characterization of mAbs. But mAbs get denatured under these conditions.
\nThere is a growing interest in the analysis of mAbs by online-SEC-MS under native conditions. Conventional SEC analysis of mAbs use phosphate buffers at pH 6.7—for example, the most common one is composed of 100 mmol/L phosphates (monobasic + dibasic) as buffering salts +100 mM Na2SO4 as neutral salt to adjust the ionic strength +0.05% NaN3 (as antibacterial agent). Both the buffering salts (phosphates) and neutral salt (sodium sulfate) are helpful in preventing secondary interaction of the proteins with the stationary phase. The concentration of these salts may need further optimization depending on the individual properties of the mAbs. But phosphate buffer is not suitable for the mass spectrometer and yields substantial noise and damage the MS system. So online SEC-MS is not possible in the presence of phosphate and other non-volatile salts. Use of volatile salts at lower concentration, which do not interfere with the MS system, can be applied and the method needs to be optimized as well. SEC columns should not exhibit particle shedding which will interfere with the MS signal.
\nThe data below illustrate the effective use of MS-friendly mobile phase compositions in the online SEC-MS analysis of a monoclonal IgG1, IgG2 antibody, ADC and Bi-specific mAb using volatile salt environments (Figure 22).
\nSeparation of an ADC, two IgG1 and IgG2 mAbs, and the corresponding bi-specific mAb using a TSKgel UP-SW3000 column (2 μm, 4.6 mm ID x 30 cm).
The online LC–MS compatible chromatographic conditions used for the analysis of IgG1, IgG2, ADC, and a Bi-specific mAb is shown below.
\nFollowing the development of an optimized separation, liquid chromatography mass spectrometry (LC–MS) analysis was performed using a Q Exactive Plus mass spectrometer (ThermoFisher Scientific) coupled to a Shimadzu Nexera XR UHPLC system. Samples were injected onto a TSKgel UP-SW3000 column (2 μm, 4.6 mm ID x 30 cm) and isocratically separated at 0.350 ml/min for 15 min with a mobile phase comprising 20 mM ammonium acetate and 10 mM ammonium bicarbonate, pH 7.2. A 15-min blank isocratic gradient was run between sample injections. No carryover was observed in the blank runs. Eluted proteins were analyzed by the mass spectrometer set to repetitively scan m/z from 800 to 6000 in a positive ion mode. The full MS scan was collected at 17,500 resolution, with spray voltage 4 kV, S-Lens RF 75, and in-source CID 80 eV. Protein mass deconvolution was performed using ProMass (Novatia). The (1) total ion chromatogram, (2) mass spectrum, and (3) deconvoluted mass spectrum of one mAb was evaluated. A main peak can be seen at m/z 149,264; adjacent peaks at m/z 149,426 and 149,592 correspond to different glycoforms.
\nHere we report the use of a TSKgel® UP-SW3000, 2 μm column for the separation of a bispecific antibody and the two parent mAbs (IgG1) followed by MS analysis. The Bispecific T cell Engager (BiTE®) technology was used in this study. BiTE is a fusion protein consisting of two single-chain variable fragments (scFvs)–CD19, a biomarker for normal and neoplastic B cells and CD3 (on T cells) – recombinantly linked by a nonimmunogenic five-amino-acid chain (Figure 23). BiTE is approximately 55 kDa in size. SEC/MS analysis was performed by the Wistar Proteomics and Metabolomics Facility (Philadelphia, PA) using a Nexera® XR UHPLC system (Shimadzu) coupled to a Q Exactive™ Plus mass spectrometer (Thermo Fisher Scientific) (Figures 24–26).
\nScheme of a BiTE and corresponding original mAb 1 and mAb 2.
SEC/MS analysis of the CD19 X CD3 BiTE antibody.
SEC/MS analysis of the original IgG1 mAb1.
Analysis of blank injections in order to assess column particle shedding using the TSKgel UP-SW3000 column.
Prior to analysis, a blank injection was run in order to assess column particle shedding. The total ion chromatogram of a blank injection was run on a new TSKgel UP-SW3000 column. MS data indicate that there is no shedding from the TSKgel UP-SW3000 column prior to sample injection. Additionally a blank injection was run between each of the sample injections in order to monitor sample carryover.
\nEach mAb is different, and a method with the use of volatile salts needs to be optimized for reproducibility. There was a difference between the retention time of mAb1 under the isocratic mobile phase 20 mM ammonium acetate and 10 mM ammonium bicarbonate, pH 7.2 compared to 100 mM phosphate buffer containing 100 mM Na2SO4 and 0.05% NaN3 pH 6.8. In an attempt to look for the condition where a MS compatible buffer yields a retention time similar to phosphate buffer, a comparison of elution profiles under 100 mM phosphate buffer and 100 mM ammonium acetate buffer both at pH 6.8 is shown below (Figure 27 and Table 4).
\nComparison of elution profiles of IgG1 under 100 mM phosphate buffer and 100 mM ammonium acetate buffer, pH 6.8 using a TSKgel UP-SW3000 column.
Analysis of retention time, peak area, peak height, as (peak asymmetry) and N (theoretical plates) of the monomers. The average (Avg), standard deviation (Std) and relative standard deviation (RSD) are also shown.
Monomer peak areas remain constant under both conditions with high reproducibility of all the peak parameters. % RSD deviations of all the peak parameters were low. Mass spectrometric analysis under this chromatographic condition will be reported elsewhere.
\nTwo chromatographic modes are considered orthogonal techniques if the selectivity of the two modes are significantly different. Under ideal conditions without any secondary interaction, SEC should yield a characteristic Gaussian-shaped peak without the presence of any heterogeneity. As shown in the figure below when individual peaks F (ab) 2 and Fab + Fc from SEC separation (panel A) were applied to the reversed-phase chromatographic (RPC) column, a number of hydrophobic variants eluted (panel B) in the increasing order of their hydrophobicity (Figure 28). Mechanism of papain digestion is discussed in Section 2.6. Though papain digestion yields primarily the Fab fragments, F(ab’)2 fragment can be generated if the papain is first activated with 10 mM cysteine. Following the completion of the reaction, the excess needs to be removed by gel filtration. Size exclusion chromatography cannot differentiate these heterogenic impurities or hydrophobic variants, which are not sufficiently different in the size or hydrodynamic radii from each other. Similarly, a number of other chromatographic modes, other than RPC, can also be used as an orthogonal technique. The extent of the heterogeneity present in the SEC peak can only be confirmed by an orthogonal analysis.
\nAnalysis of papain digested IgG1 fragments. (A) Chromatogram of papain digested IgG1 fragments separated on a TSKgel SuperSW3000 column. (B) Individual peaks F (ab) 2 and Fab + Fc from the SEC separation (panel A) were applied to a reversed-phase chromatographic (RPC) TSKgel protein C4–300 column.
Similarly, a reversed-phase chromatography column can also be used as a complimentary chromatography column along with SEC as shown below (Figure 29). The elution order of elution of the peaks is simply reversed as expected.
\nSeparation of PEG (MW 5000)-lysozyme and PEG (MW 30,000)-lysozyme on a SEC TSKgel SuperSW3000 column (A) followed by chromatography of the SEC fractions 1–4 using a reverse phase TSKgel protein C4–300 column (B).
The PEG-conjugated species were more strongly retained by RPC, than the different forms of intact lysozyme. The order of elution in RPC is opposite to the order of size-based separation in SEC [35].
\nWith the advancement of the liquid chromatographic instruments from HPLC to UHPLC and with the advancement in column packing technology with smaller particle sizes, an easy method transfer from HPLC to UHPLC is becoming necessary. UHPLC has much lower extra column volume and can withstand more than 1000 bar besides other optimized instrumental features while conventional HPLC has maximum operable pressure of 400 bars. Differences between HPLC and UHPLC instruments give rise to many challenges when a method needs to be transferred from one system to another and if equivalent separation profiles are obtained. On the other hand, there are analysts who do not possess the UHPLC but still want to get advantage of smaller particle size SEC columns while using a traditional HPLC instrument. The effect of extra column volume in band broadening in conventional HPLC needs to be reduced and optimized by reducing the diffusion in the tubing between the injection valve and the column and between the column and the UV cell by using smaller ID tubing and micro flow cell. A column compatible with both UHPLC and conventional HPLC instruments may be helpful for easy method transfer in both cases.
\nAs an example, an SEC HPLC method for the separation of a mAb using 5 μm, 7.8 mm ID × 30 cm SEC column (TSKgel G3000SWXL) was transferred to a 2 μm, 4.6 mm ID × 30 cm SEC column (TSKgel UP-SW3000 SEC) on a UHPLC instrument (Figure 30). The mobile phase and other chromatographic parameters were not changed except the flow rate reduced for the 2-μm column.
\nComparison of the SEC HPLC method with the SEC UHPLC system. Separation efficiency of column TSKgel G3000SWXL (A) and column TSKgel UP-SW3000 SEC (B) was compared by loading a mixture of standard proteins (Thy = thyroglobulin bovine, γ-glo = γ-globulins from bovine blood, ova = albumin chicken egg grade VI, riboA = ribonuclease A type I-A from bovine pancreas, pAba = p-aminobenzoid acid) on both columns.
TSKgel UP-SW3000 columns feature the same pore size as the well-established TSKgel G3000SWxl columns. With the use of the 2-μm column, the resolution between peaks 2–3, 3–4, 4–5, 5–6, and 6–7 increased respectively by 15, 25, 33, 24, and 37% compared to the 5-μm column. Retention time consistency was maintained, and similar separation profile was obtained at lower flow rate. A smaller 2-μm particle size column yielded a twofold higher sensitivity. The 2-μm columns yielded a back pressure acceptable for use in both HPLC and UHPLC.
\nSimilarly, a method can be transferred directly from HPLC to UHPLC, without any change in conditions using 2 μm TSKgel UP-SW3000 columns as shown in Figure 31 [42].
\nComparison of the separation efficiency of a 2-μm TSKgel UP-SW3000 column applied in HPLC and UHPLC mode. Conditions were the same as in
Desalting is a process to remove or reduce salt from the liquid, such as protein sample solution. Desalting by gel filtration chromatography (GFC) is the preferred method in biochemical laboratories to reduce the salt concentration or to exchange the buffer of a biopolymer solution. The main advantage of desalting by GFC over dialysis is the faster analysis time. Desalting may be needed for various reasons. Proteins eluting at high or elevated salt concentrations may need to be desalted to lower salt concentration prior to its use for the next step. Protein samples may also contain denaturants such as sodium dodecyl sulfate (SDS), guanidine hydrochloride, and urea which need to be removed. Desalting and buffer exchange of proteins or polynucleotides can also be performed by dialysis, ultra-filtration, or by using spin columns. Desalting columns are characterized by a low exclusion limit and a large pore volume. Salts can fully access all pores, while proteins and other high MW species are excluded. Analytical columns packed with conventional packing materials such as dextran, cellulose, and polyacrylamide have limited physical stability and are not suitable when fast desalting is desired. Requirements for a fast desalting SEC column are [1] an inert matrix, [2] a large pore volume that is fully accessible to common salts and buffer components, [3] a pore size distribution that excludes the component(s) of interest from accessing the pores, and [4] sufficient mechanical strength to allow the use of the column in standard HPLC equipment. As an example, a 15-μm particle size TSKgel BioAssist DS column is composed of a stationary phase where the mechanical strength of the polyacrylamide gel is fourfold higher as compared to conventional gel by urea cross-linking. Conventionally, polyacrylamide beads have been prepared by reversed-phase suspension polymerization or by using a spray dry method. The uniform and more pressure-stable polyacrylamide beads packed in TSKgel BioAssist DS columns were prepared using a normal phase suspension method as shown in Figure 32 [43].
\nPrinciple of the generation of pressure-stable polyacrylamide beads which can be packed in TSKgel BioAssist DS columns.
Fast desalting with excellent reproducibility could be carried out within 5 min using conventional HPLC system and TSKgel BioAssist DS Columns (4.6 mm ID and 10 mm ID) (Figure 33). All the proteins (see table below) eluted with the same retention time closer to void volume irrespective of their size (see the figures below), while salt and other small impurities eluted at longer retention time as a function of their size. Refractive index was used as a detector in this study since salts do not have any chromophore.
\nDesalting of proteins using a TSKgel BioAssist DS column with 4.6 mm ID or 10 mm ID packed with pressure-stable polyacrylamide beads. The retention times of standard proteins and salt are shown.
SEC columns designed for desalting using a HPLC instrument can be useful for the desalting of proteins and polynucleotides at analytical and semi-preparative scale.
\nFor many years, SEC columns have been used to separate various nucleic acid species such as DNA, RNA, and tRNA as well as their constituent bases, adenine, guanine, thymine, cytosine, and uracil. In medicine, several primary nucleobases are the basis for the nucleoside analogues and other synthetic analogs which are used as anticancer and antiviral agents. Nucleobase modifications are the basis of oligonucleotide-based therapeutics, making their purification very important.
\nHydrophilic interaction chromatography (HILIC) is a variant of normal phase liquid chromatography which uses hydrophilic stationary phases with reversed-phase type eluents. It is applied for the separation of polar hydrophilic compounds.
\nChemically bonded diol-coated phases in size exclusion chromatography (SEC) columns demonstrate high polarity and hydrogen bonding properties. They do not contain ionizable groups compared to the unreacted free residual silanols, making them appropriate for the HILIC mode.
\nFigure 34 [44] shows the separation of four nucleobases using a TSKgel SuperSW mAb HTP Column under SEC condition and in HILIC mode at pH 7.4 (Figure 34).
\nSeparation of four nucleobases using a TSKgel SuperSW mAb HTP column under SEC condition (left panel) and in HILIC mode (right panel) at pH 7.4.
As expected, due to the similarities in molecular masses between the four compounds, significant interference is observed among the peaks of interest, particularly the three pyrimidine derivatives, when separated on the TSKgel SuperSW mAb HTP column under SEC conditions. The late elution of adenine (relative to the other three compounds) may be attributed to possible interactions between the stationary phase and the derivatized purine compound, leading to a shift toward a longer retention time. When the same SEC column is used in the HILIC mode, the order of elution of the analytes does not correlate with their molecular mass (as in SEC separations), but instead is based on their relative hydrophilicity. This note demonstrates the benefits of using a SEC column in HILIC mode for the superior resolution of four nucleobases, as opposed to using the column in the SEC mode or using another type of a HILIC column.
\nIt is a recommended practice to protect the column from potential sources of contamination during the SEC separation of mAbs and other proteins. Standards and mobile phases should be filtered through a 0.45-μm syringe filter. A frit filter used between injector and column will also be an additional help. The use of guard columns is highly recommended. Guard columns being short, of similar ID and with the same stationary phase do not possess any separation power. The slight change in the retention time due to small increase in length remains constant in consecutive injections. Using guard columns can prolong the lifetime of the analytical column. The guard column needs to be changed before the dirty material spills over to the analytical column. Frits at different parts of the HPLC instruments need to be changed intermittently. Please refer to the picture below which clearly shows how much dirty materials are trapped by these frits (Figure 35). Frequent changes of the frits are necessary to avoid spillage of the dirty materials to the columns. Phosphate buffer pH 6.8, very commonly used for the protein analysis by SEC is prone to bacterial growth. The column will get clogged and dirty, eventually leading to failure of the analysis and breakdown of the system. Previously I mentioned that sodium azide (NaN3) was used as an antibacterial agent to prevent fouling of the phosphate buffer.
\nComparison of a dirty frit with a new frit.
During the SEC of mAbs and proteins, mobile phases containing salts are constantly used and can deposit on the different parts of the HPLC system (Figure 36). The pump head assembly may get affected from the deposition of salts, resulting into a rise of back pressure as well as causing damage to the sapphire piston. The pump head assembly can be cleaned with distilled water and the back pressure issue is resolved. Mobile phase containing high salt or a combination of high salt and organic solvent can create this problem more than any other low salt aqueous mobile phases. The system should not remain idle with these types of mobile phases. The deposition may happen anywhere within the HPLC system and also inside the column, or in the pump, the injector, the in-line filter or the tubing.
\nSalt depositions at screws of the column.
The use of a surfactant is necessary for certain SEC applications as discussed earlier, but surfactants may change the bonding phase, so it may be necessary to dedicate the column for that particular application.
\nOccasionally protein samples are adsorbed onto the packing material. When this occurs, it is time to clean the SEC column. At the beginning of the separation, when the SEC column is new and operating correctly and the mAb monomer is yielding a well-resolved peak, it is better to establish baseline data and acceptable running conditions criteria. Then, if one of the performance characteristics of the SEC column changes by 10% or more, it is prudent that cleaning is necessary. The acceptance criteria can be more stringent as needed. Similar cleaning of the whole system including the flow cell may be necessary.
\nIf cleaning is necessary, it is better to try cleaning the column in reversed direction at half the flow rate while the column is detached from the detector to prevent detector damage since the proteins have a tendency to adhere to the quartz material of the detector. Since the dirty materials may remain trapped over the frit outside the column, cleaning in reversed flow direction may be the easiest way to clean the column. Once the column is cleaned using the vendor suggested protocol, the analyst must qualify the column using the QC method and QC pass criteria under normal flow direction.
\nIt is always recommended to read the operational conditions and specification sheet (OCS) before the column is used. Occasionally, we may intend to use the column outside the specification for a short period of time, but it is the responsibility of the analyst to monitor if the column is functioning properly. For example, in Section 2.5, it has been shown that an analysis could be repeated to monitor the stability of the antibody over time using a TSKgel SuperSW mAb-HTP column at 0.75 mL/min. The used flow rate (0.75 mL/min) was higher than the recommended maximum flow rate of 0.5 mL/min, as mentioned in the operational conditions and specification (OCS) sheet for this column. Though the column could successfully be used at this higher flow rate without compromising the column health, the effect of the higher flow rate on the column lifetime for prolonged use was not investigated. It is always wise to operate within recommended maximum flow rate to remain in safe side in order to prolong the lifetime of the column. It is always necessary to keep an eye on the back pressure, peak parameters, and so on to monitor the column health.
\nHPLC system and analytical chromatography columns are costly. Method development is time-consuming and costly too. It is important to employ an HPLC system that is optimized with regard to extra-column band broadening to take full advantage of the high column efficiency that can be obtained on analytical columns. For any troubleshooting situation, the problem is one of the following: [1] the column, [2] the sample, [3] the mobile phases, [4] the instrumentation, and [5] personal errors. The same is true for size exclusion chromatography related troubleshooting issues too. Proper maintenance of the column and instrument is much more needed and important to avoid the troubleshooting in the beginning. Many troubleshooting issues can be avoided by taking proper handling the top three major components of any HPLC analysis. The importance of the use of a protein standard mixture and the standard mAb to monitor the column health from time to time is always a good idea as mentioned in brief in Section 2.5. Maintaining a SEC column is an art and good performance, which will result in a prolonged column lifetime.
\nSize-exclusion chromatography is a great tool for the purification of monoclonal antibodies. The secondary interaction of the stationary phase needs to be taken care of by controlling the stationary phase as well as by optimizing the mobile phase. The effect of particle size and pore size of the stationary phase on SEC separation needs to be clearly understood. A right column selection for the purification will depend on the separation range and the slope of the calibration curve. Separation of HMW and LMW species to purify the monomer is critical to get the pure mAb without heterogenic impurities, which might be immunogenic to human. Enzymatic digestion and forced degradations are necessary to characterize the monoclonal antibodies, and SEC columns need to be rightfully selected based on the separation goal. Mobile phase additives may be necessary to improve the separation, and few examples are discussed. Since online LC/MS is becoming popular, a mobile phase compatibility is needed as discussed in this chapter. Since size exclusion chromatography cannot separate the heterogenic species without much difference in hydrodynamic radii, an orthogonal technique is necessary. A similarly complimentary technique is also helpful for complete characterization of the monoclonal antibodies. SEC column developed, and optimization of surface and pore characteristics is critical for the separation of monoclonal antibodies and other proteins by HPLC and UHPLC instrument. With the advancement of UHPLC, an easy method transfer between HPLC and UHPLC is becoming necessary. Ease of method transfer using a column with dual functionality for its use in both HPLC and UHPLC is helpful. Desalting using a conventional SEC HPLC column can be very useful in removing the unwanted salts and additives. A robust separation with excellent reproducibility needs a size exclusion chromatography column with optimized packing as well as the knowledge about the tips and tricks to maintain the column lifetime
\nYoshimi Hashimoto, Toshiaki Nishi, Kazuaki Muranaka, Kosuke Araki, Yasutami Mitoma, Shigeru Nakatani, Hasegawa, Sachiyuki, Oscar Yamasaki, Toru Satoh – Tosoh Corporation, Separation Centre, Japan for their contributions in a number of references and overall support. Tosoh Bioscience LLC and Tosoh Corporation, Tosoh Bioscience LLC, Tosoh Bioscience Gmbh
\nMany 3D hydrodynamic and water quality models have been developed since the 1960s, and different numerical solution techniques have been used to solve the governing equations. The most popular numerical models and the basis that other models has been built based on are POM [1, 2], ECOM [1, 3], NCOM [4, 5], FVCOM [6, 7], EFDC [8], TRIM-3D [9], UnTRIM [10], GLLVHT [11], and DNS [12].
\nDuring the development stage of any numerical model, verification tests need to be performed to ensure that model foundations are valid. The 3D simulation models available in market have been tested either by comparing the predictions with the analytical solution, field data, or both. As a result, each verification approach has its advantages and disadvantages depending on the model complexity (governing equations used to develop the model and assumptions used to simplify the problem).
\nAll three-dimensional models available to simulate surface waterbodies do not have outputs related to the model of volume balance performance (see the user manuals of the above popular models). Therefore, the user does not know the model preserves volume or not during the simulation period even though the model gives results. In addition, most 3D users run the simulation for a very short time (even for seconds), thinking the model is stable, since the 3D numerical models require long computation time to run. Thus, the need to develop a new volume balance tool arises based on these issues related to 3D hydrodynamic numerical models used in practice for surface waterbodies.
\nIn this work, the volume balance approach was used as a tool to measure how a model preserves volume during the simulation time by calculating the accumulated error over time as a percent. Therefore, the modeler can monitor the model performance over time and decide whether the model is robust or not while running the model rather than waiting until the end of simulation.
\nTo implement the volume balance approach, the three-dimensional model W3 developed by [13] for modeling hydrodynamics, temperature, and water quality in surface waterbodies was employed. Using the finite differences, the model solves the governing equations of continuity, free surface, momentums, and mass transport. Comparisons with analytical solutions and field data were carried out for verifying and validating the W3 model [13, 14, 15, 16, 17].
\nThe model of volume balance was performed by comparing the water volume in the model domain during a time period with the water volume entering and leaving the same domain during the same period of time.
\nLet Vol be the accumulated water volume in the model domain over time. Then,
\nwhere Volinitial = the initial water volume within the domain; Volin = the accumulated water volume entering the domain; and Volout = the accumulated water volume leaving the domain.
\nThus, the error over time can be calculated as follows:
\nwhere Volinternal is the water volume within the domain at any time during the simulation period.
\nA subroutine was added to the model to check the volume preservation by calculating % error at every time step. A lower % error represents more accurate model predictions. The error should reach a constant value with time and should not grow with time. If % error grows with time exponentially, this implies that the model goes unstable (blows up). Two tests implementing the volume balance check were performed. One of these tests examined the volume balance over a rectangular domain, and the other tests evaluated the volume balance over an irregular domain. Both tests were performed over a period of 100 days based on the same real meteorological data, calculated solar short radiation, and constant inflow and outflow. The meteorological data are shown in Figures 1
Wind speed input data.
Wind direction input data.
Air temperature input data.
Dew point input data.
Cloud cover input data.
The physical domain was divided into computational cells of 1000 × 500 × 1 (
Irregular physical domain and the input bathymetry.
Using a time step of 35 s and a degree of implicitness (θ) of 1, the code was run for the simulation period. Figure 7 presents the model predictions of the surface velocity field at Julian day 100. The model results showed good performance in following the bends at the boundaries. Furthermore, the volume balance error gave a good agreement in preserving volume in which the percent error reached a constant low value over time as shown in Figure 8, which is a semilog plot of the percent error with time. The corresponding water levels at three locations over time were shown in Figure 9, denoting a very small change (≅0.005 m) in the surface layer thickness resulting from the free water surface waves.
\nSurface velocity field for the irregular domain at Julian day 100.
Volume balance for the irregular domain using θ = 1.
Surface layer thickness over time for the irregular domain using θ = 1.
Since the W3 model uses the degree of implicitness to switch between the fully implicit numerical scheme and the fully explicit scheme, the effect of the degree of implicitness on the accumulated error was evaluated by running the code using θ = 0.5 with the same inputs that were used with θ = 1. The results showed that using the semi-implicit scheme of θ = 0.5 produces less percent error than using θ = 1. Figure 10 shows the percent error after running the code for day 100 using two degrees of implicitness (θ = 1 and θ = 0.5).
\nThe volume balance for the irregular domain using θ = 1 and θ = 0.5.
In addition and in order to make sure that the numerical answers do not depend on the grid resolution, a grid refinement was performed, and the associated volume error was assessed. The code was run using θ = 0.5 with three horizontal grid resolutions 1000 × 500, 500 × 500, and 500 × 125 (
The effect of grid refinement.
Model verification is the first step after building any new hydrodynamic numerical model for surface waterbody simulation. In this chapter, a new volume balance approach was introduced for verifying the three-dimensional hydrodynamic numerical models in surface waterbody simulation. This technique provides information about whether the code preserves fluid mass or not by calculating the volume balance percent error over time during a model simulation. The model results indicated that the model is considered numerically stable if the volume balance error reaches a constant value over time. In addition, even though the model degree of implicitness had a reasonable volume balance error (less than 0.1%), the semi-implicit numerical scheme had slightly better volume balance error than the fully implicit scheme.
\nThe authors thank the Department of Civil and Environmental Engineering, Portland State University, Portland, OR, USA, for their help in doing this research in association with the Iraqi Ministry of Higher Education and Scientific Research, University of Babylon.
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His studies in robotics lead him not only to a PhD degree but also inspired him to co-found and build the International Journal of Advanced Robotic Systems - world's first Open Access journal in the field of robotics.",institutionString:null,institution:{name:"TU Wien",country:{name:"Austria"}}},{id:"441",title:"Ph.D.",name:"Jaekyu",middleName:null,surname:"Park",slug:"jaekyu-park",fullName:"Jaekyu Park",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/441/images/1881_n.jpg",biography:null,institutionString:null,institution:{name:"LG Corporation (South Korea)",country:{name:"Korea, South"}}},{id:"465",title:"Dr",name:"Christian",middleName:null,surname:"Martens",slug:"christian-martens",fullName:"Christian Martens",position:null,profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",biography:null,institutionString:null,institution:null},{id:"479",title:"Dr.",name:"Valentina",middleName:null,surname:"Colla",slug:"valentina-colla",fullName:"Valentina Colla",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/479/images/358_n.jpg",biography:null,institutionString:null,institution:{name:"Sant'Anna School of Advanced Studies",country:{name:"Italy"}}},{id:"494",title:"PhD",name:"Loris",middleName:null,surname:"Nanni",slug:"loris-nanni",fullName:"Loris Nanni",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/494/images/system/494.jpg",biography:"Loris Nanni received his Master Degree cum laude on June-2002 from the University of Bologna, and the April 26th 2006 he received his Ph.D. in Computer Engineering at DEIS, University of Bologna. On September, 29th 2006 he has won a post PhD fellowship from the university of Bologna (from October 2006 to October 2008), at the competitive examination he was ranked first in the industrial engineering area. He extensively served as referee for several international journals. He is author/coauthor of more than 100 research papers. He has been involved in some projects supported by MURST and European Community. 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Delac received his B.Sc.E.E. degree in 2003 and is currentlypursuing a Ph.D. degree at the University of Zagreb, Faculty of Electrical Engineering andComputing. His current research interests are digital image analysis, pattern recognition andbiometrics.",institutionString:null,institution:{name:"University of Zagreb",country:{name:"Croatia"}}},{id:"557",title:"Dr.",name:"Andon",middleName:"Venelinov",surname:"Topalov",slug:"andon-topalov",fullName:"Andon Topalov",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/557/images/1927_n.jpg",biography:"Dr. Andon V. Topalov received the MSc degree in Control Engineering from the Faculty of Information Systems, Technologies, and Automation at Moscow State University of Civil Engineering (MGGU) in 1979. He then received his PhD degree in Control Engineering from the Department of Automation and Remote Control at Moscow State Mining University (MGSU), Moscow, in 1984. From 1985 to 1986, he was a Research Fellow in the Research Institute for Electronic Equipment, ZZU AD, Plovdiv, Bulgaria. In 1986, he joined the Department of Control Systems, Technical University of Sofia at the Plovdiv campus, where he is presently a Full Professor. He has held long-term visiting Professor/Scholar positions at various institutions in South Korea, Turkey, Mexico, Greece, Belgium, UK, and Germany. And he has coauthored one book and authored or coauthored more than 80 research papers in conference proceedings and journals. 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Aalborg University has Two Satellite Campuses, one in Copenhagen (Aalborg University Copenhagen) and the other in Esbjerg (Aalborg University Esbjerg).\n· He is a member of prestigious IEEE (Institute of Electrical and Electronics Engineers), and IAENG (International Association of Engineers) organizations. \n· He is the chief Editor of the Journal of Software Engineering.\n· He is the member of the Editorial Board of International Journal of Computer Science and Software Technology (IJCSST) and International Journal of Computer Engineering and Information Technology. \n· He is also the Editor of Communication in Computer and Information Science CCIS-20 by Springer.\n· Reviewer For Many Conferences\nHe is the lead person in making collaboration agreements between Aalborg University and many universities of Pakistan, for which the MOU’s (Memorandum of Understanding) have been signed.\nProfessor Akbar is working in Academia since 1990, he started his career as a Lab demonstrator/TA at the University of Sussex. After finishing his P. hD degree in 1992, he served in the Industry as a Scientific Officer and continued his academic career as a visiting scholar for a number of educational institutions. In 1996 he joined National University of Science & Technology Pakistan (NUST) as an Associate Professor; NUST is one of the top few universities in Pakistan. In 1999 he joined an International Company Lineo Inc, Canada as Manager Compiler Group, where he headed the group for developing Compiler Tool Chain and Porting of Operating Systems for the BLACKfin processor. The processor development was a joint venture by Intel and Analog Devices. In 2002 Lineo Inc., was taken over by another company, so he joined Aalborg University Denmark as an Assistant Professor.\nProfessor Akbar has truly a multi-disciplined career and he continued his legacy and making progress in many areas of his interests both in teaching and research. He has contributed in stochastic estimation of control area especially, in the Multiple Target Tracking and Interactive Multiple Model (IMM) research, Ball & Beam Control Problem, Robotics, Levitation Control. He has contributed in developing Algorithms for Fingerprint Matching, Computer Vision and Face Recognition. He has been supervising Pattern Recognition, Formal Languages and Distributed Processing projects for several years. He has reviewed many books on Management, Computer Science. Currently, he is an active and permanent reviewer for many international conferences and symposia and the program committee member for many international conferences.\nIn teaching he has taught the core computer science subjects like, Digital Design, Real Time Embedded System Programming, Operating Systems, Software Engineering, Data Structures, Databases, Compiler Construction. 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These reactions occur through a regular radical chain causing growth of polymer by three steps, namely, initiation, propagation, and termination. To understand ionizing radiation-induced polymerization, the water radiolysis must be taken into consideration. This chapter explores the mechanism of water molecules radiolysis paying especial attention to the basic regularities of solvent radicals’ interaction with the polymer molecules for forming the crosslinked polymer. Water radiolysis is the main engine of the polymerization processes, especially the “free-radical polymerization.” The mechanisms of the free-radical polymerization and crosslinking will be discussed in detail later. Since different polymers respond differently to radiation, it is useful to quantify the response, namely in terms of crosslinking and chain scission. A parameter called the G-value is frequently used for this purpose. It represents the chemical yield of crosslinks, scissions and double bonds, etc. For the crosslinked polymer, the crosslinking density increases with increasing the radiation dose, this is reflected by the swelling degree of the polymer while being immersed in a compatible solvent. If crosslinking predominates, the crosslinking density increases and the extent of swelling decreases. If chain scission predominates, the opposite occurs. A further detailed discussion of these aspects is presented throughout this chapter.",book:{id:"6149",slug:"ionizing-radiation-effects-and-applications",title:"Ionizing Radiation Effects and Applications",fullTitle:"Ionizing Radiation Effects and Applications"},signatures:"Mohamed Mohamady Ghobashy",authors:[{id:"212371",title:"Dr.",name:"Mohamed",middleName:null,surname:"Mohamady Ghobashy",slug:"mohamed-mohamady-ghobashy",fullName:"Mohamed Mohamady Ghobashy"}]},{id:"53504",doi:"10.5772/66925",title:"Applications of Ionizing Radiation in Mutation Breeding",slug:"applications-of-ionizing-radiation-in-mutation-breeding",totalDownloads:3478,totalCrossrefCites:9,totalDimensionsCites:13,abstract:"As a predicted result of increasing population worldwide, improvements in the breeding strategies in agriculture are valued as mandatory. The natural resources are limited, and due to the natural disasters like sudden and severe abiotic stress factors, excessive floods, etc., the production capacities are changed per year. In contrast, the yield potential should be significantly increased to cope with this problem. Despite rich genetic diversity, manipulation of the cultivars through alternative techniques such as mutation breeding becomes important. Radiation is proven as an effective method as a unique method to increase the genetic variability of the species. Gamma radiation is the most preferred physical mutagen by plant breeders. Several mutant varieties have been successfully introduced into commercial production by this method. Combinational use of in vitro tissue culture and mutation breeding methods makes a significant contribution to improve new crops. Large populations and the target mutations can be easily screened and identified by new methods. Marker assisted selection and advanced techniques such as microarray, next generation sequencing methods to detect a specific mutant in a large population will help to the plant breeders to use ionizing radiation efficiently in breeding programs.",book:{id:"5451",slug:"new-insights-on-gamma-rays",title:"New Insights on Gamma Rays",fullTitle:"New Insights on Gamma Rays"},signatures:"Özge Çelik and Çimen Atak",authors:[{id:"147362",title:"Dr.",name:"Özge",middleName:null,surname:"Çelik",slug:"ozge-celik",fullName:"Özge Çelik"},{id:"147364",title:"Prof.",name:"Çimen",middleName:null,surname:"Atak",slug:"cimen-atak",fullName:"Çimen Atak"}]},{id:"32846",doi:"10.5772/36950",title:"Current Importance and Potential Use of Low Doses of Gamma Radiation in Forest Species",slug:"current-importance-and-potential-use-of-low-doses-of-gamma-radiation-in-forest-species",totalDownloads:5277,totalCrossrefCites:2,totalDimensionsCites:13,abstract:null,book:{id:"1590",slug:"gamma-radiation",title:"Gamma Radiation",fullTitle:"Gamma Radiation"},signatures:"L. G. Iglesias-Andreu, P. Octavio-Aguilar and J. Bello-Bello",authors:[{id:"110581",title:"Dr.",name:"Lourdes",middleName:null,surname:"Iglesias-Andreu",slug:"lourdes-iglesias-andreu",fullName:"Lourdes Iglesias-Andreu"}]},{id:"58410",doi:"10.5772/intechopen.72074",title:"Radiation-Induced Degradation of Organic Compounds and Radiation Technologies for Purification of Aqueous Systems",slug:"radiation-induced-degradation-of-organic-compounds-and-radiation-technologies-for-purification-of-aq",totalDownloads:1415,totalCrossrefCites:8,totalDimensionsCites:12,abstract:"Environmental application of radiation technologies is an important part of radiation processing. Radiation treatment of aqueous systems contaminated with organic compounds is a promising method of water and wastewater purification and corresponding technologies are being developed. In this chapter, the following aspects of radiation treatment process are considered: sources of contamination and major contaminants of water and wastewater; primary processes in aqueous systems initiated by ionizing radiation; principal ways of contaminant conversion as consequences of primary processes (complete mineralization of organic compounds, partial decomposition of organic molecules resulted in detoxification, decolorization, disinfection of polluted water, and improvement in biological degradation of contaminant, polymerization of monomers’ contaminants, oxidation-reduction processes, and coagulation of colloids); sources of ionizing radiation; and main equipment applied in radiation technologies of aqueous system purification.",book:{id:"6149",slug:"ionizing-radiation-effects-and-applications",title:"Ionizing Radiation Effects and Applications",fullTitle:"Ionizing Radiation Effects and Applications"},signatures:"Igor E. Makarov and Alexander V. Ponomarev",authors:[{id:"213652",title:"Dr.",name:"Igor",middleName:null,surname:"Makarov",slug:"igor-makarov",fullName:"Igor Makarov"},{id:"213657",title:"Dr.",name:"Alexander",middleName:null,surname:"Ponomarev",slug:"alexander-ponomarev",fullName:"Alexander Ponomarev"}]}],mostDownloadedChaptersLast30Days:[{id:"32842",title:"Sterilization by Gamma Irradiation",slug:"sterilization-by-gamma-irradiation",totalDownloads:74766,totalCrossrefCites:37,totalDimensionsCites:85,abstract:null,book:{id:"1590",slug:"gamma-radiation",title:"Gamma Radiation",fullTitle:"Gamma Radiation"},signatures:"Kátia Aparecida da Silva Aquino",authors:[{id:"102109",title:"Dr.",name:"Katia",middleName:"Aparecida Da S.",surname:"Aquino",slug:"katia-aquino",fullName:"Katia Aquino"}]},{id:"32837",title:"Environmental Gamma-Ray Observation in Deep Sea",slug:"environmental-gamma-ray-observation-in-deep-sea-",totalDownloads:2917,totalCrossrefCites:4,totalDimensionsCites:6,abstract:null,book:{id:"1590",slug:"gamma-radiation",title:"Gamma Radiation",fullTitle:"Gamma Radiation"},signatures:"Hidenori Kumagai, Ryoichi Iwase, Masataka Kinoshita, Hideaki Machiyama, Mutsuo Hattori and Masaharu Okano",authors:[{id:"108174",title:"Dr.",name:"Hidenori",middleName:null,surname:"Kumagai",slug:"hidenori-kumagai",fullName:"Hidenori Kumagai"},{id:"108237",title:"Dr.",name:"Masa",middleName:null,surname:"Kinoshita",slug:"masa-kinoshita",fullName:"Masa Kinoshita"},{id:"137650",title:"Dr.",name:"Ryoichi",middleName:null,surname:"Iwase",slug:"ryoichi-iwase",fullName:"Ryoichi Iwase"},{id:"137656",title:"Dr.",name:"Hideaki",middleName:null,surname:"Machiyama",slug:"hideaki-machiyama",fullName:"Hideaki Machiyama"},{id:"146918",title:"Dr.",name:"Mutsuo",middleName:null,surname:"Hattori",slug:"mutsuo-hattori",fullName:"Mutsuo Hattori"},{id:"146919",title:"Dr.",name:"Masaharu",middleName:null,surname:"Okano",slug:"masaharu-okano",fullName:"Masaharu Okano"}]},{id:"58998",title:"Ionizing Radiation-Induced Polymerization",slug:"ionizing-radiation-induced-polymerization",totalDownloads:1784,totalCrossrefCites:8,totalDimensionsCites:17,abstract:"Ionizing radiation can induce some kinds of reactions, other than polymerization, such as dimerization, oligomerization, curing, and grafting. These reactions occur through a regular radical chain causing growth of polymer by three steps, namely, initiation, propagation, and termination. To understand ionizing radiation-induced polymerization, the water radiolysis must be taken into consideration. This chapter explores the mechanism of water molecules radiolysis paying especial attention to the basic regularities of solvent radicals’ interaction with the polymer molecules for forming the crosslinked polymer. Water radiolysis is the main engine of the polymerization processes, especially the “free-radical polymerization.” The mechanisms of the free-radical polymerization and crosslinking will be discussed in detail later. Since different polymers respond differently to radiation, it is useful to quantify the response, namely in terms of crosslinking and chain scission. A parameter called the G-value is frequently used for this purpose. It represents the chemical yield of crosslinks, scissions and double bonds, etc. For the crosslinked polymer, the crosslinking density increases with increasing the radiation dose, this is reflected by the swelling degree of the polymer while being immersed in a compatible solvent. If crosslinking predominates, the crosslinking density increases and the extent of swelling decreases. If chain scission predominates, the opposite occurs. A further detailed discussion of these aspects is presented throughout this chapter.",book:{id:"6149",slug:"ionizing-radiation-effects-and-applications",title:"Ionizing Radiation Effects and Applications",fullTitle:"Ionizing Radiation Effects and Applications"},signatures:"Mohamed Mohamady Ghobashy",authors:[{id:"212371",title:"Dr.",name:"Mohamed",middleName:null,surname:"Mohamady Ghobashy",slug:"mohamed-mohamady-ghobashy",fullName:"Mohamed Mohamady Ghobashy"}]},{id:"53780",title:"Gamma-Ray Spectrometry and the Investigation of Environmental and Food Samples",slug:"gamma-ray-spectrometry-and-the-investigation-of-environmental-and-food-samples",totalDownloads:2501,totalCrossrefCites:1,totalDimensionsCites:1,abstract:"Gamma radiation consists of high‐energy photons and penetrates matter. This is an advantage for the detection of gamma rays, as gamma spectrometry does not need the elimination of the matrix. The disadvantage is the need of shielding to protect against this radiation. Gamma rays are everywhere: in the atmosphere; gamma nuclides are produced by radiation of the sun; in the Earth, the primordial radioactive nuclides thorium and uranium are sources for gamma and other radiation. The technical enrichment and use of radioisotopes led to the unscrupulously use of radioactive material and to the Cold War, with over 900 bomb tests from 1945 to 1990, combined with global fallout over the northern hemisphere. The friendly use of radiation in medicine and for the production of energy at nuclear power plants (NPPs) has caused further expositions with ionising radiation. This chapter describes in a practical manner the instrumentation for the detection of gamma radiation and some results of the use of these techniques in environmental and food investigations.",book:{id:"5451",slug:"new-insights-on-gamma-rays",title:"New Insights on Gamma Rays",fullTitle:"New Insights on Gamma Rays"},signatures:"Markus R. 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He is also a faculty member in the Molecular Oncology Program. He obtained his MSc and Ph.D. at Oregon State University and Texas Tech University, respectively. He pursued his postdoctoral studies at Rutgers University Medical School and the National Institutes of Health (NIH/NIDDK), USA. His research focuses on biochemistry, biophysics, genetics, molecular biology, and molecular medicine with specialization in the fields of drug design, protein structure-function, protein folding, prions, microRNA, pseudogenes, molecular cancer, epigenetics, metabolites, proteomics, genomics, protein expression, and characterization by spectroscopic and calorimetric methods.",institutionString:"University of Health Sciences",institution:null},{id:"180528",title:"Dr.",name:"Hiroyuki",middleName:null,surname:"Kagechika",slug:"hiroyuki-kagechika",fullName:"Hiroyuki Kagechika",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/180528/images/system/180528.jpg",biography:"Hiroyuki Kagechika received his bachelor’s degree and Ph.D. in Pharmaceutical Sciences from the University of Tokyo, Japan, where he served as an associate professor until 2004. He is currently a professor at the Institute of Biomaterials and Bioengineering (IBB), Tokyo Medical and Dental University (TMDU). From 2010 to 2012, he was the dean of the Graduate School of Biomedical Science. Since 2012, he has served as the vice dean of the Graduate School of Medical and Dental Sciences. He has been the director of the IBB since 2020. Dr. Kagechika’s major research interests are the medicinal chemistry of retinoids, vitamins D/K, and nuclear receptors. He has developed various compounds including a drug for acute promyelocytic leukemia.",institutionString:"Tokyo Medical and Dental University",institution:{name:"Tokyo Medical and Dental University",country:{name:"Japan"}}},{id:"268659",title:"Ms.",name:"Xianquan",middleName:null,surname:"Zhan",slug:"xianquan-zhan",fullName:"Xianquan Zhan",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/268659/images/8143_n.jpg",biography:"Dr. Zhan received his undergraduate and graduate training in the fields of preventive medicine and epidemiology and statistics at the West China University of Medical Sciences in China during 1989 to 1999. He received his post-doctoral training in oncology and cancer proteomics for two years at the Cancer Research Institute of Human Medical University in China. In 2001, he went to the University of Tennessee Health Science Center (UTHSC) in USA, where he was a post-doctoral researcher and focused on mass spectrometry and cancer proteomics. Then, he was appointed as an Assistant Professor of Neurology, UTHSC in 2005. He moved to the Cleveland Clinic in USA as a Project Scientist/Staff in 2006 where he focused on the studies of eye disease proteomics and biomarkers. He returned to UTHSC as an Assistant Professor of Neurology in the end of 2007, engaging in proteomics and biomarker studies of lung diseases and brain tumors, and initiating the studies of predictive, preventive, and personalized medicine (PPPM) in cancer. In 2010, he was promoted to Associate Professor of Neurology, UTHSC. Currently, he is a Professor at Xiangya Hospital of Central South University in China, Fellow of Royal Society of Medicine (FRSM), the European EPMA National Representative in China, Regular Member of American Association for the Advancement of Science (AAAS), European Cooperation of Science and Technology (e-COST) grant evaluator, Associate Editors of BMC Genomics, BMC Medical Genomics, EPMA Journal, and Frontiers in Endocrinology, Executive Editor-in-Chief of Med One. He has\npublished 116 peer-reviewed research articles, 16 book chapters, 2 books, and 2 US patents. His current main research interest focuses on the studies of cancer proteomics and biomarkers, and the use of modern omics techniques and systems biology for PPPM in cancer, and on the development and use of 2DE-LC/MS for the large-scale study of human proteoforms.",institutionString:null,institution:{name:"Xiangya Hospital Central South University",country:{name:"China"}}},{id:"40482",title:null,name:"Rizwan",middleName:null,surname:"Ahmad",slug:"rizwan-ahmad",fullName:"Rizwan Ahmad",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/40482/images/system/40482.jpeg",biography:"Dr. Rizwan Ahmad is a University Professor and Coordinator, Quality and Development, College of Medicine, Imam Abdulrahman bin Faisal University, Saudi Arabia. Previously, he was Associate Professor of Human Function, Oman Medical College, Oman, and SBS University, Dehradun. Dr. Ahmad completed his education at Aligarh Muslim University, Aligarh. He has published several articles in peer-reviewed journals, chapters, and edited books. His area of specialization is free radical biochemistry and autoimmune diseases.",institutionString:"Imam Abdulrahman Bin Faisal University",institution:{name:"Imam Abdulrahman Bin Faisal University",country:{name:"Saudi Arabia"}}},{id:"41865",title:"Prof.",name:"Farid A.",middleName:null,surname:"Badria",slug:"farid-a.-badria",fullName:"Farid A. Badria",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/41865/images/system/41865.jpg",biography:"Farid A. Badria, Ph.D., is the recipient of several awards, including The World Academy of Sciences (TWAS) Prize for Public Understanding of Science; the World Intellectual Property Organization (WIPO) Gold Medal for best invention; Outstanding Arab Scholar, Kuwait; and the Khwarizmi International Award, Iran. He has 250 publications, 12 books, 20 patents, and several marketed pharmaceutical products to his credit. He continues to lead research projects on developing new therapies for liver, skin disorders, and cancer. Dr. Badria was listed among the world’s top 2% of scientists in medicinal and biomolecular chemistry in 2019 and 2020. He is a member of the Arab Development Fund, Kuwait; International Cell Research Organization–United Nations Educational, Scientific and Cultural Organization (ICRO–UNESCO), Chile; and UNESCO Biotechnology France",institutionString:"Mansoura University",institution:{name:"Mansoura University",country:{name:"Egypt"}}},{id:"329385",title:"Dr.",name:"Rajesh K.",middleName:"Kumar",surname:"Singh",slug:"rajesh-k.-singh",fullName:"Rajesh K. Singh",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/329385/images/system/329385.png",biography:"Dr. Singh received a BPharm (2003) and MPharm (2005) from Panjab University, Chandigarh, India, and a Ph.D. (2013) from Punjab Technical University (PTU), Jalandhar, India. He has more than sixteen years of teaching experience and has supervised numerous postgraduate and Ph.D. students. He has to his credit more than seventy papers in SCI- and SCOPUS-indexed journals, fifty-five conference proceedings, four books, six Best Paper Awards, and five projects from different government agencies. He is currently an editorial board member of eight international journals and a reviewer for more than fifty scientific journals. He received Top Reviewer and Excellent Peer Reviewer Awards from Publons in 2016 and 2017, respectively. He is also on the panel of The International Reviewer for reviewing research proposals for grants from the Royal Society. He also serves as a Publons Academy mentor and Bentham brand ambassador.",institutionString:"Punjab Technical University",institution:{name:"Punjab Technical University",country:{name:"India"}}},{id:"142388",title:"Dr.",name:"Thiago",middleName:"Gomes",surname:"Gomes Heck",slug:"thiago-gomes-heck",fullName:"Thiago Gomes Heck",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/142388/images/7259_n.jpg",biography:null,institutionString:null,institution:{name:"Universidade Regional do Noroeste do Estado do Rio Grande do Sul",country:{name:"Brazil"}}},{id:"336273",title:"Assistant Prof.",name:"Janja",middleName:null,surname:"Zupan",slug:"janja-zupan",fullName:"Janja Zupan",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/336273/images/14853_n.jpeg",biography:"Janja Zupan graduated in 2005 at the Department of Clinical Biochemistry (superviser prof. dr. Janja Marc) in the field of genetics of osteoporosis. Since November 2009 she is working as a Teaching Assistant at the Faculty of Pharmacy, Department of Clinical Biochemistry. In 2011 she completed part of her research and PhD work at Institute of Genetics and Molecular Medicine, University of Edinburgh. She finished her PhD entitled The influence of the proinflammatory cytokines on the RANK/RANKL/OPG in bone tissue of osteoporotic and osteoarthritic patients in 2012. From 2014-2016 she worked at the Institute of Biomedical Sciences, University of Aberdeen as a postdoctoral research fellow on UK Arthritis research project where she gained knowledge in mesenchymal stem cells and regenerative medicine. She returned back to University of Ljubljana, Faculty of Pharmacy in 2016. She is currently leading project entitled Mesenchymal stem cells-the keepers of tissue endogenous regenerative capacity facing up to aging of the musculoskeletal system funded by Slovenian Research Agency.",institutionString:null,institution:{name:"University of Ljubljana",country:{name:"Slovenia"}}},{id:"357453",title:"Dr.",name:"Radheshyam",middleName:null,surname:"Maurya",slug:"radheshyam-maurya",fullName:"Radheshyam Maurya",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/357453/images/16535_n.jpg",biography:null,institutionString:null,institution:{name:"University of Hyderabad",country:{name:"India"}}},{id:"418340",title:"Dr.",name:"Jyotirmoi",middleName:null,surname:"Aich",slug:"jyotirmoi-aich",fullName:"Jyotirmoi Aich",position:null,profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0033Y000038Ugi5QAC/Profile_Picture_2022-04-15T07:48:28.png",biography:"Biotechnologist with 15 years of research including 6 years of teaching experience. Demonstrated record of scientific achievements through consistent publication record (H index = 13, with 874 citations) in high impact journals such as Nature Communications, Oncotarget, Annals of Oncology, PNAS, and AJRCCM, etc. Strong research professional with a post-doctorate from ACTREC where I gained experimental oncology experience in clinical settings and a doctorate from IGIB where I gained expertise in asthma pathophysiology. A well-trained biotechnologist with diverse experience on the bench across different research themes ranging from asthma to cancer and other infectious diseases. An individual with a strong commitment and innovative mindset. Have the ability to work on diverse projects such as regenerative and molecular medicine with an overall mindset of improving healthcare.",institutionString:"DY Patil Deemed to Be University",institution:null},{id:"349288",title:"Prof.",name:"Soumya",middleName:null,surname:"Basu",slug:"soumya-basu",fullName:"Soumya Basu",position:null,profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0033Y000035QxIDQA0/Profile_Picture_2022-04-15T07:47:01.jpg",biography:"Soumya Basu, Ph.D., is currently working as an Associate Professor at Dr. D. Y. Patil Biotechnology and Bioinformatics Institute, Dr. D. Y. Patil Vidyapeeth, Pune, Maharashtra, India. With 16+ years of trans-disciplinary research experience in Drug Design, development, and pre-clinical validation; 20+ research article publications in journals of repute, 9+ years of teaching experience, trained with cross-disciplinary education, Dr. Basu is a life-long learner and always thrives for new challenges.\r\nHer research area is the design and synthesis of small molecule partial agonists of PPAR-γ in lung cancer. She is also using artificial intelligence and deep learning methods to understand the exosomal miRNA’s role in cancer metastasis. Dr. Basu is the recipient of many awards including the Early Career Research Award from the Department of Science and Technology, Govt. of India. She is a reviewer of many journals like Molecular Biology Reports, Frontiers in Oncology, RSC Advances, PLOS ONE, Journal of Biomolecular Structure & Dynamics, Journal of Molecular Graphics and Modelling, etc. She has edited and authored/co-authored 21 journal papers, 3 book chapters, and 15 abstracts. She is a Board of Studies member at her university. She is a life member of 'The Cytometry Society”-in India and 'All India Cell Biology Society”- in India.",institutionString:"Dr. D.Y. Patil Vidyapeeth, Pune",institution:{name:"Dr. D.Y. Patil Vidyapeeth, Pune",country:{name:"India"}}},{id:"354817",title:"Dr.",name:"Anubhab",middleName:null,surname:"Mukherjee",slug:"anubhab-mukherjee",fullName:"Anubhab Mukherjee",position:null,profilePictureURL:"https://intech-files.s3.amazonaws.com/0033Y0000365PbRQAU/ProfilePicture%202022-04-15%2005%3A11%3A18.480",biography:"A former member of Laboratory of Nanomedicine, Brigham and Women’s Hospital, Harvard University, Boston, USA, Dr. Anubhab Mukherjee is an ardent votary of science who strives to make an impact in the lives of those afflicted with cancer and other chronic/acute ailments. He completed his Ph.D. from CSIR-Indian Institute of Chemical Technology, Hyderabad, India, having been skilled with RNAi, liposomal drug delivery, preclinical cell and animal studies. He pursued post-doctoral research at College of Pharmacy, Health Science Center, Texas A & M University and was involved in another postdoctoral research at Department of Translational Neurosciences and Neurotherapeutics, John Wayne Cancer Institute, Santa Monica, California. In 2015, he worked in Harvard-MIT Health Sciences & Technology as a visiting scientist. He has substantial experience in nanotechnology-based formulation development and successfully served various Indian organizations to develop pharmaceuticals and nutraceutical products. He is an inventor in many US patents and an author in many peer-reviewed articles, book chapters and books published in various media of international repute. Dr. Mukherjee is currently serving as Principal Scientist, R&D at Esperer Onco Nutrition (EON) Pvt. Ltd. and heads the Hyderabad R&D center of the organization.",institutionString:"Esperer Onco Nutrition Pvt Ltd.",institution:null},{id:"319365",title:"Assistant Prof.",name:"Manash K.",middleName:null,surname:"Paul",slug:"manash-k.-paul",fullName:"Manash K. Paul",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/319365/images/system/319365.png",biography:"Manash K. Paul is a Principal Investigator and Scientist at the University of California Los Angeles. He has contributed significantly to the fields of stem cell biology, regenerative medicine, and lung cancer. His research focuses on various signaling processes involved in maintaining stem cell homeostasis during the injury-repair process, deciphering lung stem cell niche, pulmonary disease modeling, immuno-oncology, and drug discovery. He is currently investigating the role of extracellular vesicles in premalignant lung cell migration and detecting the metastatic phenotype of lung cancer via machine-learning-based analyses of exosomal signatures. Dr. Paul has published in more than fifty peer-reviewed international journals and is highly cited. He is the recipient of many awards, including the UCLA Vice Chancellor’s award, a senior member of the Institute of Electrical and Electronics Engineers (IEEE), and an editorial board member for several international journals.",institutionString:"University of California Los Angeles",institution:{name:"University of California Los Angeles",country:{name:"United States of America"}}},{id:"311457",title:"Dr.",name:"Júlia",middleName:null,surname:"Scherer Santos",slug:"julia-scherer-santos",fullName:"Júlia Scherer Santos",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/311457/images/system/311457.jpg",biography:"Dr. Júlia Scherer Santos works in the areas of cosmetology, nanotechnology, pharmaceutical technology, beauty, and aesthetics. Dr. Santos also has experience as a professor of graduate courses. Graduated in Pharmacy, specialization in Cosmetology and Cosmeceuticals applied to aesthetics, specialization in Aesthetic and Cosmetic Health, and a doctorate in Pharmaceutical Nanotechnology. Teaching experience in Pharmacy and Aesthetics and Cosmetics courses. She works mainly on the following subjects: nanotechnology, cosmetology, pharmaceutical technology, aesthetics.",institutionString:"Universidade Federal de Juiz de Fora",institution:{name:"Universidade Federal de Juiz de Fora",country:{name:"Brazil"}}},{id:"219081",title:"Dr.",name:"Abdulsamed",middleName:null,surname:"Kükürt",slug:"abdulsamed-kukurt",fullName:"Abdulsamed Kükürt",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/219081/images/system/219081.png",biography:"Dr. Kükürt graduated from Uludağ University in Turkey. He started his academic career as a Research Assistant in the Department of Biochemistry at Kafkas University. In 2019, he completed his Ph.D. program in the Department of Biochemistry at the Institute of Health Sciences. He is currently working at the Department of Biochemistry, Kafkas University. He has 27 published research articles in academic journals, 11 book chapters, and 37 papers. He took part in 10 academic projects. He served as a reviewer for many articles. He still serves as a member of the review board in many academic journals.",institutionString:"Kafkas University",institution:{name:"Kafkas University",country:{name:"Turkey"}}},{id:"178366",title:"Associate Prof.",name:"Volkan",middleName:null,surname:"Gelen",slug:"volkan-gelen",fullName:"Volkan Gelen",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/178366/images/system/178366.jpg",biography:"Volkan Gelen is a Physiology specialist who received his veterinary degree from Kafkas University in 2011. Between 2011-2015, he worked as an assistant at Atatürk University, Faculty of Veterinary Medicine, Department of Physiology. In 2016, he joined Kafkas University, Faculty of Veterinary Medicine, Department of Physiology as an assistant professor. Dr. Gelen has been engaged in various academic activities at Kafkas University since 2016. There he completed 5 projects and has 3 ongoing projects. He has 60 articles published in scientific journals and 20 poster presentations in scientific congresses. His research interests include physiology, endocrine system, cancer, diabetes, cardiovascular system diseases, and isolated organ bath system studies.",institutionString:"Kafkas University",institution:{name:"Kafkas University",country:{name:"Turkey"}}},{id:"418963",title:"Dr.",name:"Augustine Ododo",middleName:"Augustine",surname:"Osagie",slug:"augustine-ododo-osagie",fullName:"Augustine Ododo Osagie",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/418963/images/16900_n.jpg",biography:"Born into the family of Osagie, a prince of the Benin Kingdom. I am currently an academic in the Department of Medical Biochemistry, University of Benin. 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She is a fellow member of the Royal Society of Chemistry UK and the American Chemical Society of the United States.",institutionString:"King Saud University",institution:{name:"King Saud University",country:{name:"Saudi Arabia"}}},{id:"49848",title:"Dr.",name:"Wen-Long",middleName:null,surname:"Hu",slug:"wen-long-hu",fullName:"Wen-Long Hu",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/49848/images/system/49848.jpg",biography:"Wen-Long Hu is Chief of the Division of Acupuncture, Department of Chinese Medicine at Kaohsiung Chang Gung Memorial Hospital, as well as an adjunct associate professor at Fooyin University and Kaohsiung Medical University. Wen-Long is President of Taiwan Traditional Chinese Medicine Medical Association. He has 28 years of experience in clinical practice in laser acupuncture therapy and 34 years in acupuncture. He is an invited speaker for lectures and workshops in laser acupuncture at many symposiums held by medical associations. He owns the patent for herbal preparation and producing, and for the supercritical fluid-treated needle. Dr. Hu has published three books, 12 book chapters, and more than 30 papers in reputed journals, besides serving as an editorial board member of repute.",institutionString:"Kaohsiung Chang Gung Memorial Hospital",institution:{name:"Kaohsiung Chang Gung Memorial Hospital",country:{name:"Taiwan"}}},{id:"298472",title:"Prof.",name:"Andrey V.",middleName:null,surname:"Grechko",slug:"andrey-v.-grechko",fullName:"Andrey V. Grechko",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/298472/images/system/298472.png",biography:"Andrey Vyacheslavovich Grechko, Ph.D., Professor, is a Corresponding Member of the Russian Academy of Sciences. He graduated from the Semashko Moscow Medical Institute (Semashko National Research Institute of Public Health) with a degree in Medicine (1998), the Clinical Department of Dermatovenerology (2000), and received a second higher education in Psychology (2009). Professor A.V. Grechko held the position of Сhief Physician of the Central Clinical Hospital in Moscow. He worked as a professor at the faculty and was engaged in scientific research at the Medical University. Starting in 2013, he has been the initiator of the creation of the Federal Scientific and Clinical Center for Intensive Care and Rehabilitology, Moscow, Russian Federation, where he also serves as Director since 2015. He has many years of experience in research and teaching in various fields of medicine, is an author/co-author of more than 200 scientific publications, 13 patents, 15 medical books/chapters, including Chapter in Book «Metabolomics», IntechOpen, 2020 «Metabolomic Discovery of Microbiota Dysfunction as the Cause of Pathology».",institutionString:"Federal Research and Clinical Center of Intensive Care Medicine and Rehabilitology",institution:null},{id:"199461",title:"Prof.",name:"Natalia V.",middleName:null,surname:"Beloborodova",slug:"natalia-v.-beloborodova",fullName:"Natalia V. Beloborodova",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/199461/images/system/199461.jpg",biography:'Natalia Vladimirovna Beloborodova was educated at the Pirogov Russian National Research Medical University, with a degree in pediatrics in 1980, a Ph.D. in 1987, and a specialization in Clinical Microbiology from First Moscow State Medical University in 2004. She has been a Professor since 1996. Currently, she is the Head of the Laboratory of Metabolism, a division of the Federal Research and Clinical Center of Intensive Care Medicine and Rehabilitology, Moscow, Russian Federation. N.V. Beloborodova has many years of clinical experience in the field of intensive care and surgery. She studies infectious complications and sepsis. She initiated a series of interdisciplinary clinical and experimental studies based on the concept of integrating human metabolism and its microbiota. Her scientific achievements are widely known: she is the recipient of the Marie E. Coates Award \\"Best lecturer-scientist\\" Gustafsson Fund, Karolinska Institutes, Stockholm, Sweden, and the International Sepsis Forum Award, Pasteur Institute, Paris, France (2014), etc. Professor N.V. Beloborodova wrote 210 papers, five books, 10 chapters and has edited four books.',institutionString:"Federal Research and Clinical Center of Intensive Care Medicine and Rehabilitology",institution:null},{id:"354260",title:"Ph.D.",name:"Tércio Elyan",middleName:"Azevedo",surname:"Azevedo Martins",slug:"tercio-elyan-azevedo-martins",fullName:"Tércio Elyan Azevedo Martins",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/354260/images/16241_n.jpg",biography:"Graduated in Pharmacy from the Federal University of Ceará with the modality in Industrial Pharmacy, Specialist in Production and Control of Medicines from the University of São Paulo (USP), Master in Pharmaceuticals and Medicines from the University of São Paulo (USP) and Doctor of Science in the program of Pharmaceuticals and Medicines by the University of São Paulo. Professor at Universidade Paulista (UNIP) in the areas of chemistry, cosmetology and trichology. Assistant Coordinator of the Higher Course in Aesthetic and Cosmetic Technology at Universidade Paulista Campus Chácara Santo Antônio. Experience in the Pharmacy area, with emphasis on Pharmacotechnics, Pharmaceutical Technology, Research and Development of Cosmetics, acting mainly on topics such as cosmetology, antioxidant activity, aesthetics, photoprotection, cyclodextrin and thermal analysis.",institutionString:null,institution:{name:"University of Sao Paulo",country:{name:"Brazil"}}},{id:"334285",title:"Ph.D. Student",name:"Sameer",middleName:"Kumar",surname:"Jagirdar",slug:"sameer-jagirdar",fullName:"Sameer Jagirdar",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/334285/images/14691_n.jpg",biography:"I\\'m a graduate student at the center for biosystems science and engineering at the Indian Institute of Science, Bangalore, India. I am interested in studying host-pathogen interactions at the biomaterial interface.",institutionString:null,institution:{name:"Indian Institute of Science Bangalore",country:{name:"India"}}},{id:"329248",title:"Dr.",name:"Md. Faheem",middleName:null,surname:"Haider",slug:"md.-faheem-haider",fullName:"Md. Faheem Haider",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/329248/images/system/329248.jpg",biography:"Dr. Md. Faheem Haider completed his BPharm in 2012 at Integral University, Lucknow, India. In 2014, he completed his MPharm with specialization in Pharmaceutics at Babasaheb Bhimrao Ambedkar University, Lucknow, India. He received his Ph.D. degree from Jamia Hamdard University, New Delhi, India, in 2018. He was selected for the GPAT six times and his best All India Rank was 34. Currently, he is an assistant professor at Integral University. Previously he was an assistant professor at IIMT University, Meerut, India. He has experience teaching DPharm, Pharm.D, BPharm, and MPharm students. He has more than five publications in reputed journals to his credit. Dr. Faheem’s research area is the development and characterization of nanoformulation for the delivery of drugs to various organs.",institutionString:"Integral University",institution:{name:"Integral University",country:{name:"India"}}},{id:"329795",title:"Dr.",name:"Mohd Aftab",middleName:"Aftab",surname:"Siddiqui",slug:"mohd-aftab-siddiqui",fullName:"Mohd Aftab Siddiqui",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/329795/images/15648_n.jpg",biography:"Dr. Mohd Aftab Siddiqui is currently working as Assistant Professor in the Faculty of Pharmacy, Integral University, Lucknow for the last 6 years. He has completed his Doctor in Philosophy (Pharmacology) in 2020 from Integral University, Lucknow. He completed his Bachelor in Pharmacy in 2013 and Master in Pharmacy (Pharmacology) in 2015 from Integral University, Lucknow. He is the gold medalist in Bachelor and Master degree. He qualified GPAT -2013, GPAT -2014, and GPAT 2015. His area of research is Pharmacological screening of herbal drugs/ natural products in liver and cardiac diseases. He has guided many M. Pharm. research projects. He has many national and international publications.",institutionString:"Integral University",institution:null},{id:"333824",title:"Dr.",name:"Ahmad Farouk",middleName:null,surname:"Musa",slug:"ahmad-farouk-musa",fullName:"Ahmad Farouk Musa",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/333824/images/22684_n.jpg",biography:"Dato’ Dr Ahmad Farouk Musa\nMD, MMED (Surgery) (Mal), Fellowship in Cardiothoracic Surgery (Monash Health, Aust), Graduate Certificate in Higher Education (Aust), Academy of Medicine (Mal)\n\n\n\nDato’ Dr Ahmad Farouk Musa obtained his Doctor of Medicine from USM in 1992. He then obtained his Master of Medicine in Surgery from the same university in the year 2000 before subspecialising in Cardiothoracic Surgery at Institut Jantung Negara (IJN), Kuala Lumpur from 2002 until 2005. He then completed his Fellowship in Cardiothoracic Surgery at Monash Health, Melbourne, Australia in 2008. He has served in the Malaysian army as a Medical Officer with the rank of Captain upon completing his Internship before joining USM as a trainee lecturer. He is now serving as an academic and researcher at Monash University Malaysia. He is a life-member of the Malaysian Association of Thoracic & Cardiovascular Surgery (MATCVS) and a committee member of the MATCVS Database. He is also a life-member of the College of Surgeons, Academy of Medicine of Malaysia; a life-member of Malaysian Medical Association (MMA), and a life-member of Islamic Medical Association of Malaysia (IMAM). Recently he was appointed as an Interim Chairperson of Examination & Assessment Subcommittee of the UiTM-IJN Cardiothoracic Surgery Postgraduate Program. As an academic, he has published numerous research papers and book chapters. He has also been appointed to review many scientific manuscripts by established journals such as the British Medical Journal (BMJ). He has presented his research works at numerous local and international conferences such as the European Association for Cardiothoracic Surgery (EACTS) and the European Society of Cardiovascular Surgery (ESCVS), to name a few. He has also won many awards for his research presentations at meetings and conferences like the prestigious International Invention, Innovation & Technology Exhibition (ITEX); Design, Research and Innovation Exhibition, the National Conference on Medical Sciences and the Annual Scientific Meetings of the Malaysian Association for Thoracic and Cardiovascular Surgery. He was awarded the Darjah Setia Pangkuan Negeri (DSPN) by the Governor of Penang in July, 2015.",institutionString:null,institution:{name:"Monash University Malaysia",country:{name:"Malaysia"}}},{id:"30568",title:"Prof.",name:"Madhu",middleName:null,surname:"Khullar",slug:"madhu-khullar",fullName:"Madhu Khullar",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/30568/images/system/30568.jpg",biography:"Dr. Madhu Khullar is a Professor of Experimental Medicine and Biotechnology at the Post Graduate Institute of Medical Education and Research, Chandigarh, India. She completed her Post Doctorate in hypertension research at the Henry Ford Hospital, Detroit, USA in 1985. She is an editor and reviewer of several international journals, and a fellow and member of several cardiovascular research societies. Dr. Khullar has a keen research interest in genetics of hypertension, and is currently studying pharmacogenetics of hypertension.",institutionString:"Post Graduate Institute of Medical Education and Research",institution:{name:"Post Graduate Institute of Medical Education and Research",country:{name:"India"}}},{id:"223233",title:"Prof.",name:"Xianquan",middleName:null,surname:"Zhan",slug:"xianquan-zhan",fullName:"Xianquan Zhan",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/223233/images/system/223233.png",biography:"Xianquan Zhan received his MD and Ph.D. in Preventive Medicine at West China University of Medical Sciences. He received his post-doctoral training in oncology and cancer proteomics at the Central South University, China, and the University of Tennessee Health Science Center (UTHSC), USA. He worked at UTHSC and the Cleveland Clinic in 2001–2012 and achieved the rank of associate professor at UTHSC. Currently, he is a full professor at Central South University and Shandong First Medical University, and an advisor to MS/PhD students and postdoctoral fellows. He is also a fellow of the Royal Society of Medicine and European Association for Predictive Preventive Personalized Medicine (EPMA), a national representative of EPMA, and a member of the American Society of Clinical Oncology (ASCO) and the American Association for the Advancement of Sciences (AAAS). He is also the editor in chief of International Journal of Chronic Diseases & Therapy, an associate editor of EPMA Journal, Frontiers in Endocrinology, and BMC Medical Genomics, and a guest editor of Mass Spectrometry Reviews, Frontiers in Endocrinology, EPMA Journal, and Oxidative Medicine and Cellular Longevity. He has published more than 148 articles, 28 book chapters, 6 books, and 2 US patents in the field of clinical proteomics and biomarkers.",institutionString:"Shandong First Medical University",institution:{name:"Affiliated Hospital of Shandong Academy of Medical Sciences",country:{name:"China"}}},{id:"297507",title:"Dr.",name:"Charles",middleName:"Elias",surname:"Assmann",slug:"charles-assmann",fullName:"Charles Assmann",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/297507/images/system/297507.jpg",biography:"Charles Elias Assmann is a biologist from Federal University of Santa Maria (UFSM, Brazil), who spent some time abroad at the Ludwig-Maximilians-Universität München (LMU, Germany). He has Masters Degree in Biochemistry (UFSM), and is currently a PhD student at Biochemistry at the Department of Biochemistry and Molecular Biology of the UFSM. His areas of expertise include: Biochemistry, Molecular Biology, Enzymology, Genetics and Toxicology. He is currently working on the following subjects: Aluminium toxicity, Neuroinflammation, Oxidative stress and Purinergic system. Since 2011 he has presented more than 80 abstracts in scientific proceedings of national and international meetings. Since 2014, he has published more than 20 peer reviewed papers (including 4 reviews, 3 in Portuguese) and 2 book chapters. He has also been a reviewer of international journals and ad hoc reviewer of scientific committees from Brazilian Universities.",institutionString:"Universidade Federal de Santa Maria",institution:{name:"Universidade Federal de Santa Maria",country:{name:"Brazil"}}},{id:"217850",title:"Dr.",name:"Margarete Dulce",middleName:null,surname:"Bagatini",slug:"margarete-dulce-bagatini",fullName:"Margarete Dulce Bagatini",position:null,profilePictureURL:"https://mts.intechopen.com/storage/users/217850/images/system/217850.jpeg",biography:"Dr. Margarete Dulce Bagatini is an associate professor at the Federal University of Fronteira Sul/Brazil. She has a degree in Pharmacy and a PhD in Biological Sciences: Toxicological Biochemistry. She is a member of the UFFS Research Advisory Committee\nand a member of the Biovitta Research Institute. She is currently:\nthe leader of the research group: Biological and Clinical Studies\nin Human Pathologies, professor of postgraduate program in\nBiochemistry at UFSC and postgraduate program in Science and Food Technology at\nUFFS. She has experience in the area of pharmacy and clinical analysis, acting mainly\non the following topics: oxidative stress, the purinergic system and human pathologies, being a reviewer of several international journals and books.",institutionString:"Universidade Federal da Fronteira Sul",institution:{name:"Universidade Federal da Fronteira Sul",country:{name:"Brazil"}}}]}},subseries:{item:{id:"23",type:"subseries",title:"Computational Neuroscience",keywords:"Single-Neuron Modeling, Sensory Processing, Motor Control, Memory and Synaptic Pasticity, Attention, Identification, Categorization, Discrimination, Learning, Development, Axonal Patterning and Guidance, Neural Architecture, Behaviours and Dynamics of Networks, Cognition and the Neuroscientific Basis of Consciousness",scope:"Computational neuroscience focuses on biologically realistic abstractions and models validated and solved through computational simulations to understand principles for the development, structure, physiology, and ability of the nervous system. 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