Indication and properties of MSC products impact their manufacturing.
\r\n\tThe primary objective of this book is to provide the specialists involved in the clinical management and experimental research of acute and chronic leukemias updates on the theoretical aspects as well as state-of-the-art diagnostic and clinical management of acute and chronic leukemias.
\r\n\r\n\tThe book is intended to cover a broad spectrum of leukemia-related topics such as:
\r\n\t-novel and still evolving insights into the biology and diagnosis and how these result in new drug approvals and new therapeutic options with a focus on molecular and immunotherapeutic targeted therapeutics,
\r\n\t- the molecular and functional features of leukemic stem cells and their interaction with the microenvironment,
\r\n\t- preleukemic hematopoiesis and clonal diversity,
\r\n\t- new standard treatment algorithms,
\r\n\t- mechanisms of resistance and disease progression
\r\n\t- diagnosis and management of rare acute leukemia subtypes,
\r\n\t- Covid-19 aspects in specific leukemia categories,
\r\n\t- real-world data
\r\n\t- new drugs in development
Cell therapy is a growing clinical research and healthcare sector in which living cells are introduced into a patient in an attempt to ameliorate or cure a disease. Stem cell therapy is one of the most promising fields within this sector because the introduced cells have the capacity to differentiate, allowing the repopulation of diseased organs with healthy cells, or to allow even complete organ regeneration. This chapter will focus on one specific type of stem cell (MSCs), which are variously defined as mesenchymal stem cells, mesenchymal stromal cells, or (most recently) medicinal signaling cells [1]. These various definitions reflect the controversial origin and functionality of MSCs and uncertainty about their clinical potential [2, 3]. Following encouraging initial results, the unclear or disappointing outcomes of some MSC clinical trials have clouded the picture [4], but the pioneers of this approach still regard MSCs as a promising therapeutic option [5]. One of the key issues in the deployment of MSCs is ensuring they are safe and effective, which requires a well-characterized manufacturing process.
\nIn order to provide enough MSCs for cell therapy, donor cells must be isolated from tissue and then expanded
MSCs are classified as advanced therapeutic medicinal products (ATMPs) under regulations in Europe and the US. Many countries follow the regulations laid down by the US Food and Drug Administration (FDA), which defines MSCs as cell therapy products, whereas the European Medicines Agency (EMA) defines MSCs as cell-based medicinal products and distinguishes between somatic cell therapy medicinal products (SCTMPs) and tissue engineered products (TEPs) [7]. This means that clinical studies and drug approval are covered by a specific regulatory framework applied at the national or regional level. Manufacturing must therefore be compliant with good manufacturing practice (GMP) regulations that have been tailored for ATMPs, following strict criteria for product specification and release for clinical use. However, the regulatory framework for MSC manufacturing is confounded by ambiguous product definitions reflecting regional differences in the way the regulations are implemented. For example, the EMA requires GMP compliance and manufacturing authorization for phase I material, whereas the FDA does not apply this requirement until phases II and III, and in Canada, GMP compliance is not strictly required at any phase [8]. Even so, various MSC products have been manufactured under these different regulatory jurisdictions and have proceeded through clinical development, in some cases gaining market authorization from the local regulatory agency [9]. Most of these products are allogenic, which means that MSCs from one or more healthy donors are expanded, processed, and stored and then applied to patients as an off-the-shelf product (Table 1). In 2016, the allogenic MSC product TEMCELL (developed by Mesoblast) was licensed to JCR Pharmaceuticals, which received market authorization in Japan under a fast-track protocol for patients with steroid-refractory acute GvHD. Mesoblast also conducted a phase III trial with this product in the US, involving 60 patients of the same indication, achieving the primary endpoints (NCT02336230). In 2018, ALOFISEL (Takeda Pharma), an expanded allogenic adipose-derived MSC product, was approved by the EMA to treat complex perianal fistula in patients with Crohn’s disease. This was supported by a placebo-controlled trial involving 212 patients [10]. Stempeucel (Stempeutics), an expanded allogenic MSC product, received market authorization from the Drug Controller General of India to treat limb ischemia in patients with Buerger’s disease. However, it is limited to 200 patients on a cost-recovery basis, and a postmarket surveillance study is required. Ninety patients have already received an injection of this MSC product in a phase II trial, achieving a significantly better outcome than standard care [11]. CARTISTEM (Medipost) is an allogenic culture-expanded umbilical cord blood MSC product to treat knee articular cartilage defects in patients with osteoarthritis, grade IV, and following approval for the South Korean market in 2012, its clinical outcomes have remained stable over 7 years of follow-up studies [12]. Several autologous MSC products have also been approved in South Korea, meaning that the MSCs are isolated from the patient’s own tissue and then manipulated/expanded in a patented process and re-injected into the patient 4–6 weeks later. NEURONATA-R (Corestem) and Cellgram-AMI (Pharmicell) are autologous bone marrow-derived MSCs indicated for amyotrophic lateral sclerosis and acute myocardial infarction, respectively. Two other MSC products derived from adipose-tissue have been approved (Anterogen): a mixture of autologous adipose-derived MSCs with other cells for subcutaneous tissue defects (Queencell) and a pure adipose-derived MSC product for Crohn’s fistula treatment (Cupistem) [9]. NEURONATA-R has been designated as an orphan drug by the EMA and FDA.
\n\n | Product 1 | \nProduct 2 | \n
---|---|---|
Exemplary products | \nALOFISEL | \nQueencell | \n
Indication | \nCrohn’s disease, perianal fistula | \nRegeneration of subcutaneous tissue | \n
Patients per year | \n23,000 (in EU)* | \nn.d. | \n
Cell type | \nAllogenic MSCs | \nAutologous, patient-specific MSCs | \n
Cell source | \nAdipose tissue | \nAdipose tissue | \n
Cells per dose | \n1.2 × 108 MSCs | \n7 × 107** | \n
Therapeutic relevant cell properties*** | \nAnti-inflammation, immune modulation | \nRegeneration, anti-apoptosis | \n
Manufacturing type | \nBulk manufacturing | \nPatient-specific batch | \n
Batch size | \nLarge (min. 100–1000 doses per batch) | \nSmall (1 dose per batch) | \n
Scalability of production | \nScale up | \nScale out, several batches in parallel | \n
Product storage | \nFrozen, off-the-shelf | \nNo storage | \n
Stability under storage | \nStable >6 month, frozen | \nFresh, stable max. 24 hours | \n
Indication and properties of MSC products impact their manufacturing.
0.003% of all citizens (741 million) in Europe are putative patients.
Stromal vascular fraction contains MSCs and other cell types such as preadipocytes, endothelial progenitor cells, pericytes, mast cells, and fibroblast.
Following both products have different critical quality attributes (CQAs) and the manufacturing processes have different critical process parameters (CPPs).
n.d. not determined.
This brief survey of the market shows that the promise of MSC therapy is materializing, with positive efficacy data in controlled clinical trials followed by regulatory approval for a small number of products.
\nAlthough MSCs have been used in cell therapy applications for many years, the fundamental biology of these cells and their precise therapeutic properties are not fully understood. MSCs were initially isolated from bone marrow (bm-MSCs) based on their plastic adherence, but today they are usually isolated from adipose tissue (ad-MSCs) or umbilical cord blood (uc-MSCs), which are more accessible [13]. MSCs are also found in various other adult, fetal, and perinatal tissues [14]. Regardless of their origin, MSCs are heterogeneous and polyclonal cells, with at least three subpopulations defined based on morphology. Type I MSCs are spindle-shaped proliferating cells resembling fibroblasts. Type II MSCs are large, flat, epithelial-like cells, which are more senescent than type I cells and feature visible cytoskeletal structures and granules. Finally, type III MSCs are small round cells with a high capacity for self-renewal [15]. The heterogeneity of MSCs can be considered beneficial in that it ensures that some therapeutically active cells are present, but it reduces the maximum potential efficacy because some of the cells are inactive. However, even monoclonal MSCs become heterogeneous during expansion [16].
\nDespite the heterogeneity described above, the International Society of Cell Therapy has published a set of minimal criteria that must be met before cells can be defined as MSCs. Such cells must (i) show plastic adherence; (ii) be able to differentiate into cartilage, bone, and fat tissue
It is important to note that MSCs cannot be defined merely as a collection of surface markers because this says nothing about their therapeutic effect (Figure 1). Initially, the therapeutic potential of MSCs was believed to reflect their ability to migrate into damaged tissues, differentiate
Properties of MSCs and their mode of action. MSCs modulate the host immune systems, e.g., by secreting various trophic factors. Thereby, they reduce inflammation, promote neoangiogenesis, and prevent apoptosis and fibrosis. Further, they stimulate local stem cells to develop new tissue. TSG-6, tumor necrosis factor-inducible gene 6 protein also known as TNF-stimulated gene 6 protein; STC1, stanniocalcin 1; IL-4/6/10, interleukins 4, 6 and 10; CCL20, macrophage inflammatory protein-3; IDO, indoleamine 2,3-dioxygenase; PGE2, prostaglandin E2; VEGF, vascular endothelial growth factor; FGF-2, basic fibroblast growth factor; HGF, hepatocyte growth factor; IGF-1, insulin-like growth factor 1; CXCL12, stromal cell-derived factor 1; MMP1/2/9, matrix metalloproteinase-1/2/9.
The immunomodulatory properties of MSCs and their secretion of anti-inflammatory molecules and extracellular vesicles are an important therapeutic functionality [14]. MSCs are therefore logical candidates for the treatment of immune disorders, including GvHD, inflammatory bowel disease, multiple sclerosis, rheumatoid arthritis, and diabetes [21]. MSCs also secrete peptides and factors that promote the regeneration of damaged tissue by stimulating cell proliferation and migration, promoting angiogenesis, and suppressing apoptosis and fibrosis [14]. The regenerative capacity of MSCs has been used to treat Alzheimer’s disease, bone and cartilage diseases, diabetes, myocardial infarction, and osteoarthritis [22]. Another advantage of MSCs is that they do not form teratomas
The biological complexity and heterogeneity of MSCs hamper the translation of laboratory-scale experiments into industrial processes for cost-effective and reliable manufacturing. This can be addressed by developing MSC manufacturing processes that adhere to quality-by-design (QbD) principles [24]. QbD provides a rational framework and integrates scientific knowledge and risk analysis into process development. It is guided by a thorough understanding of the fundamental biology and engineering principles underlying an MSC product and its production process. QbD begins with a description of the desired product quality characteristics, known as the quality target product profile (QTPP). This is used to identify critical quality attributes (CQAs), which are physical, chemical, and biological attributes that define the quality of the product. The QTPP for MSCs describes properties such as identity, purity, and potency, which will be unique for each MSC product and dependent on the therapeutic indication.
\nFor MSCs, identity often means the cell phenotype, but as discussed above, there is no agreement on a single definition. Identity is often demonstrated by confirming a typical morphology and/or karyotype [25] and by detecting the presence or absence of surface markers. The minimal criteria for MSCs (see above) have led to a misconception that cells meeting these criteria are equivalent in identity and therapeutic functionality. In polyclonal MSC populations, the presence of multiple cell types can be a clinical benefit as stated above [26], and this should be reflected in the identity attributes.
\nThe functionality and potency of MSCs are closely linked to their therapeutic efficacy and thus the clinical outcome, but potency is used to demonstrate manufacturing consistency for batch release so a measurable property is required. Viability can fulfill the role of a potency indicator because only living cells can act as a therapeutic entity. Potency can also be measured using
If the therapeutic effect of MSCs is conferred by the secretome, then the differentiation potential may not be the primary determinant of potency. The profile of secreted factors would be a more appropriate measure, and this could be determined by multiplex enzyme-linked immunosorbent assays (ELISAs) or mass spectroscopy [31]. However, a clear link between the secretome profile and
Impurities are unwanted components from within the process, whereas contaminants come from outside the process. Impurities during MSC manufacturing include unwanted cell types, particles (e.g., residual microcarriers, or plastics and fibers from manufacturing equipment and materials), or components of culture medium. Contaminants include bacteria, fungi, viruses, endotoxins, and mycoplasma. The heterogeneity of MSCs makes it difficult to detect unwanted cell types. MSC preparations should ideally be pure, but fibroblasts are often present as impurities. Cell-specific sorting based on the marker CD166 (which is expressed at higher levels on MSCs) and CD9 (which is expressed at higher levels on fibroblasts) may help to achieve sufficient purity [33]. In other cases, it may be sufficient if most of the cells in the final product (>98%) fulfill the ISCT minimal criteria based on MSC surface markers. All other impurities and contaminants must be measured and the maximum residual levels must be defined to ensure safety and efficacy. A final sterilization step is not possible when the product is living cells, so the entire MSC production process must be carried out under aseptic conditions.
\nFrom the QTPP list, CQAs must be identified, which directly influence the safety and efficacy of the MSC product. This means that a risk assessment is carried out to reduce the QTPP list to the most influential attributes based on impact and certainty. According to ICHQ8, a CQA is “
Therapeutic applications of MSCs require at least 1 × 108 cells per dose, which is many more than can be isolated by tissue aspiration. All MSC production processes must therefore include an
The properties of MSCs are strongly influenced by the environment because MSCs in nature interact with surrounding cells and tissues, with the extracellular matrix and with various bioactive molecules. Even in an artificial environment like a bioreactor or T-flask, MSCs are very sensitive to their environment, and the most influential factors give rise to CPPs. By identifying CPPs that affect MSC quality, the process can be designed to favor the recovery of MSCs with specific phenotypes of interest, in this case those with the greatest therapeutic efficacy [34, 35]. The CPPs affecting MSC quality are discussed in more detail below.
\nDuring MSC isolation, the seeding density is important because all sources contain different quantities of MSCs. For example, only 1 in 100,000 bone marrow cells is an MSC, whereas in adipose tissue, the ratio is nearer to 1 in 100 [36]. If plastic adherence is selected as a strategy for MSC isolation, the number of adherent cells therefore differs according to the source if a similar number of tissue cells are seeded. Standardization during this step can be achieved by isolating MSCs using a strategy of surface marker sorting, allowing a defined number of cells to be seeded into the culture vessel. The seeding density selected for the
Several basal media have been shown to influence MSC expansion and potency, including Dulbecco’s modified Eagle’s medium (DMEM), Iscove’s modified Dulbecco’s medium (IMDM), and MEM alpha (αMEM) [37]. One of the key components of these media is glucose, which is the main carbon source for MSCs. Glucose may be provided at physiological concentrations (1 g/L) or higher (up to 4.5 g/L), the latter variously described as having a negative effect on MSC proliferation and growth factor secretion [38] or no effect at all [39]. Glutamine as a second carbon source is present at concentrations of 2–4 mM and appears essential for MSC growth [40], but its impact on MSC properties is complex, with contradictory results [41, 42, 43]. Glutamine is unstable at 37°C and spontaneously degrades to form ammonia. GlutaMAX (dipeptide Ala-Gln) is recommended instead of glutamine to promote MSC expansion [44]. Lactate and ammonia are the most abundant waste products formed by MSCs, and both therefore have the potential to inhibit growth. It therefore follows that glucose, glutamine, lactate, and ammonia levels should be considered as CPPs for the production of MSCs. Several other amino acids may also be relevant, given that the amino acid metabolism of MSCs differs from that of commercial cell lines such as Chinese hamster ovary (CHO) cells [42].
\nBasal media formulations must be supplemented to achieve MSC expansion. The most important supplement is fetal calf serum (FCS), which is added to a final concentration of 5–20%. FCS strongly influences MSC growth and phenotype, but the specific effectors are unknown because the composition of FCS is variable and lot-dependent [45]. The use of FCS for the manufacture of clinical MSC products is discouraged nowadays, in line with the drive to eliminate all raw materials of animal origin. The complex, uncertain, and variable composition of FCS also makes it difficult to validate for GMP-compliant processes. Finally, the manufacturing process must accommodate steps to eliminate FCS from the final product to avoid potential immunogenicity and allergenicity [46]. FCS can be replaced with human serum and its derivatives, such as human platelet lysate, which promotes MSC growth [47]. However, the same lot-dependent quality issues described above for FCS also apply to human serum [48]. The most acceptable alternative is serum-free or preferably chemically-defined medium, the latter not only serum-free but also lacking any hydrolysates or supplements of unknown composition. MSCs grow well in several commercial serum-free media, including BD Mosaic MSC Serum-free (BD Biosciences), RoosterNourish (Rooster Bio), Mesencult-XF (Stemcell Technologies), StemPro MSC SFM Xeno-Free (Invitrogen), TheraPEAK MSCGM-CD (Lonza), and PPRF-msc6, STK1 and STK2 (Abion) [49]. Growth in chemically-defined medium has also been demonstrated [50]. However, although MSCs showed excellent growth in these serum-free media, they reached senescence earlier, and there were changes in morphology, surface marker profiles, and potency [51]. This does not mean that serum-free and chemically-defined media should be avoided‑it is still better to use these media for MSC expansion in order to meet GMP requirements‑but further investigations are required to optimize the media composition. The development of serum-free media is mainly driven by companies, which tend not to disclose the precise composition, making it difficult for other researchers to build on the results. In serum-free and defined media, supplemental growth factors such as FGF2 and PDGF are needed to stimulate MSC proliferation, but they also influence MSC potency [18]. Accordingly, chemically-defined media would be preferable for the
MSCs are aerobic cells and any culture vessel must therefore ensure an adequate supply of oxygen. However, the oxygen saturation in standard T-flasks (21% O2) is far removed from nature (5–7% O2) [34]. MSCs therefore tend to be oversaturated with oxygen, which can increase the concentration of damaging reactive oxygen species (ROS). Several studies have confirmed that hypoxia enhances MSC proliferation, stabilizes their cell fate, and prevents apoptosis by reducing the levels of caspase-3 [52]. However, rather than imposing hypoxia by preconditioning the cells, it may be better to impose hypoxia during the entire expansion phase, because this mimics their natural niche [53].
\nIn addition to oxygen saturation, temperature and pH are CPPs in every process and can be monitored and controlled very easily. Typically,
Other CPPs include the parameters grouped under the term hydrodynamics, referring to the potential impact of aeration and agitation. Aeration is required to supply oxygen to the MSCs, but as well as affecting the oxygen saturation, it also generates forces that cause physical stress. In T-flasks, aeration is achieved by the diffusion of oxygen through the surface of the medium, whereas bioreactors must be actively aerated by, e.g., bubbling gas into the liquid. The bursting gas bubbles (cavitation) generate strong forces that can damage cells, although the stress can be reduced by controlling the bubble size [55]. Agitation in bioreactors is generally achieved with impellers, which help to disperse gas (and therefore contribute to aeration) but also maintain a homogenous suspension of cells and nutrients. The creation of a homogenous environment is advantageous because it avoids gradients of pH, nutrients, or waste products, whose effect on MSCs is unpredictable. Homogenization can also be achieved using pumps or is facilitated by air bubbles. Agitation always generates shear forces, so it is necessary to balance the homogeneity of the cultivation system and the impact of the hydrodynamic forces on the MSCs. Although excessive shear stress is detrimental, hydrodynamic forces can also stimulate MSC growth and increase potency [43]. For these reasons, the mode and rate of aeration and the method and intensity of agitation are CPPs that must be carefully optimized for each process.
\nMSCs are anchorage-dependent cells, so the properties of the growth surface also have a significant impact on the process and must be investigated and selected carefully. However, unlike the parameters discussed above, the growth surface does not have to be monitored or controlled during MSC production, so it falls outside the technical definition of a CPP. The expression of certain surface markers by MSCs reflects the stiffness of the growth surface, so it is clear that the surface affects the phenotype [56]. As stated above, the ability to adhere to plastic surfaces is one of the minimal criteria that define MSCs, and tissue-culture plastic is therefore the most commonly-used growth surface. Although all commercial tissue-culture plasticware has a polypropylene base, the surface is often treated differently, and this changes the behavior and properties of the adherent MSCs [37]. MSCs further grow on other surface materials, e.g., glass [57] or dextran [58]. When MSCs are cultivated in serum-free medium, cell growth often requires that the surface is coated with further adhesion-promoting factors, such as fibronectin, vitronectin, or the peptide RGD.
\nGiven that MSCs are anchorage-dependent cells, the harvesting of cells at the end of the
All the approved allogenic MSC products described earlier are cryopreserved, allowing them to be offered as off-the-shelf products that can be stored until quality control and batch release are completed. The use of cryopreserved allogeneic MSCs is the only feasible therapeutic strategy for acute tissue injury syndromes such as stroke, sepsis, or myocardial infarction, because the patient is likely to die before sufficient quantities of autologous MSCs could be prepared. However, cryopreservation and thawing have a massive impact on the potency of MSCs [66]. Indeed, even without optimization, fresh MSCs are much more potent than frozen ones [35]. A rule of thumb is to freeze the cells slowly (e.g., 1°C/min) but to thaw them quickly (e.g., direct transfer from storage to a 37°C water bath). The impact of multiple freeze-thaw cycles must be evaluated carefully [67]. The composition of the freezing medium is also important because it often contains dimethyl sulfoxide (DMSO) and FCS as cryoprotectants, the first being cytotoxic and the second undesirable for the reasons already discussed above. Nontoxic alternatives lacking DMSO and FCS have been tested and may be more compatible with MSCs intended for clinical applications [68, 69, 70].
\nIn summary, the expansion of MSCs in bioreactors involves multiple CPPs including (i) the source of the initial MSCs before expansion, (ii) the impact of cell density and age, (iii) the effects of the culture medium, (iv) the properties of the bioreactor and aeration/agitation systems, and (iv) the method used for cell harvest and storage. The impact of these CPPs on the quality of MSCs can only be determined by designing robust assays for (i)
For the 989 interventional clinical trials involving MSCs reported thus far (www.clinicaltrials.gov, search term: mesenchymal stem cell OR mesenchymal stromal cell, 2019/09/27), the MSCs were expanded
The manufacture of protein therapeutics is almost always carried out in bioreactors because they are scalable, controllable via integrated process analytical technology, and most process steps can be automated. This is not the case for MSC products, and a survey of GMP manufacturing at US academic centers has revealed major differences in the various process steps (cell isolation, expansion, and characterization). In the context of cell expansion, 80% of the centers surveyed above used T-flasks or cell factories, whereas only 20% mainly used bioreactors. A broad range of seeding densities was used for cultivation (50–2500 cells/cm2) and the cultivation time ranged from 1 to 28+ days. The cultivation medium was supplemented with FCS (lot-selected or not) or donor-pooled human platelet lysate (in-house product or commercial product) [71]. All of the centers expanded MSCs under GMP conditions, but with huge variations in the protocol. The production of MSCs in T-flasks is adequate for a small number of patients (30 T-flasks each with a growth surface of 175 cm2 would be required per patient, assuming each patient is dosed with 416 million cells and the harvesting efficiency is 8 × 104 cells/cm2 [72]). But for larger clinical trials with >100 patients, the resources required for cell culture would become unsupportable (assuming the conditions stated above, a trial with 140 patients would require 4200 T-flasks filling 32 standard 160-L incubators and 9 full-time personnel to handle the cells). Expansion in T-flasks might also be sufficient for autologous cell therapy, given that only a single patient is involved and it would not be necessary to produce more than 10 doses. However, even for small-scale manufacturing, an automated bioreactor system would offer several advantages over manual cultivation. Given that the entire manufacturing process must be aseptic, closed bioreactors provide much better insurance against contamination than an open culture system based on T-flaks. For allogenic MSC products, where up to 1 million doses are produced per batch, bioreactors are the only feasible manufacturing option (Figure 2).
\nManufacturing of autologous and allogenic MSC products. Autologous MSC products are isolated from the patient’s own tissue, whereas for allogenic MSC products a healthy donor from the same specie donates cells. In the isolation and expansion, there are few differences between the two types of MSC products. Most common sources are bone marrow, adipose tissue, and umbilical cord (blood), all three giving different amount of MSCs. The expansion for both MSC product types differs in scale. Storage is only needed for allogenic MSC products. If we expect that 0.4 × 106 MSC are isolated per donation and one dose to treat a single patient is about 120 × 106 MSCs, an expansion factor of at least 300-fold is needed. If more doses should be produced from one isolate, e.g., because the patient needs several treatments or in case of allogenic MSC products, the expansion factor dramatically increases.
When an MSC product advances from research to commercial manufacturing, the
Stirred tank reactors are the most widely-used devices for large-scale MSC expansion. They are often used with microcarriers, which are small beads that increase the surface area available for cell attachment, although MSCs can also be grown in bioreactors as aggregates or spheroids. The expansion of MSCs growing on microcarriers is typically a batch-mode manufacturing process because the cells are harvested at a predetermined density. However, fed-batch processes involve a smaller inoculum (100 cells/cm2, equivalent to five cells per microcarrier) and can thus achieve better economy and a higher expansion factor [77, 78]. There should be minimal (if any) agitation at the beginning of the expansion phase to allow for cell attachment to the microcarriers (if used) or otherwise for the formation of aggregates. However, agitation is required following attachment in order to homogenize the suspension and avoid the formation of large clumps. As discussed above, agitation is an important CPP and the parameters must be optimized based on the unique combination of system properties (e.g., impeller type/speed and microcarrier size/amount) to keep microcarriers or aggregates in suspension without causing shear damage, and these parameters must be optimized at different manufacturing scales [79].
\nFixed bed reactors are also widely used for MSC expansion, and in this case, the cells are grown either on macrocarriers or as capsules (500 μm diameter), both of which form a stable bed at the reactor base. The production of homogeneous conditions in the bed can be frustrated by the development of channels and gradients in the bed, particularly in large-scale systems [80]. The shear forces in fixed bed reactors are low (~0.5 × 10−5 N/cm2) and consistent throughout the reactor with no peaks near the impeller; the shear forces also remain constant at all scales [81]. The
The earlier sections highlighted several challenges that must be overcome to develop robust processes for the expansion of MSCs in bioreactors, which are summarized briefly below. Furthermore, our current understanding of the CPPs affecting MSC production is rudimentary at best, and more work is required to determine the impact of hydrodynamic factors on the CQAs. Precise online monitoring tools are needed to control CPPs effectively and to measure their influence on cell viability, potency, and secretory profiles. An increase in process understanding will facilitate process modeling, to fulfill the requirements of process analytic technology as a prerequisite for GMP manufacturing.
\nThe major challenge for MSC therapy is the development of an
Polyclonal MSCs often show the most potent therapeutic effects, but clonal impoverishment occurs during lengthy expansion phases and this must be avoided if potency is compromised. However, even monoclonal MSCs become heterogeneous over time, generating subpopulations with different morphologies and surface marker profiles. The therapeutic outcome can only be predicted if the MSC pool does not change during expansion, and the well-controlled conditions in bioreactors can therefore help to ensure that the cell products remain homogeneous.
\nUltimately, even bioreactor-based processes for MSC expansion are constrained by the inbuilt replication limit of MSCs, which leads to senescence after a certain number of generations. Stem cells by definition have an unlimited capacity for self-renewal, but this property is lost
The production of MSCs with standardized properties would be facilitated by the development of standardized validated potency assays so that results obtained in different laboratories are truly comparable. The ISCT has taken steps in this direction by publishing standards for the harmonization of potency assays. In a matrix assay approach, they propose to use quantitative RNA analysis for selected gene products, flow cytometry to detect functionally-relevant surface markers, and protein-based assays to map the secretome and determine the immunomodulatory potency of MSCs [88].
\nMSCs are typically the sole product of any MSC cultivation process, but in some applications, the MSCs are used as helper cells to deliver a different product or they are used as a vehicle to produce a specific cellular component. In each case, the CQAs differ significantly from the standard MSC manufacturing process and other CPPs must therefore be considered. We discuss two examples below.
\nMSCs are potent therapeutics, but researchers are seeking new ways to achieve the same therapeutic effect without the drawbacks associated with MSC manufacturing, such as the limited availability of potent cells, the complex transfusion process, and the entrapment of MSCs in nontarget organs [89]. As discussed earlier, the therapeutic effect of MSCs reflects the secretion of cytokines, growth factors, and other paracrine signaling molecules, particularly via the release of extracellular vesicles that interact directly with target cells and deliver their contents into the cytosol. The advantage of these vesicles over whole MSCs is their much greater stability, which means they can be manufactured, stored, and shipped without losing therapeutic efficacy [90, 91].
\nThe large-scale manufacturing of extracellular vesicles requires the cultivation of MSCs, which secrete these vesicles directly in the culture medium. Scalable production methods are not yet available, and vesicles are currently produced in T-flasks or cell factories without process monitoring. Bioreactors could be used to scale up production, and given there is no need to harvest the MSCs, it would be possible to consider a wider range of bioreactor systems than the relatively narrow selection favored for MSC manufacturing. A fixed bed bioreactor has been used for the continuous production and harvesting of extracellular vesicles, which increased the yield 10-fold compared to T-flasks [92]. Stirred tank reactors with microcarriers might also be suitable, but they have not yet been used for vesicle production [57]. The cells would be exposed to shear forces caused by the impellers and air bubble cavitation, and this may influence vesicle production and potency [93].
\nThe effect of different process parameters on the production of MSC-derived extracellular vesicles has been investigated at the laboratory scale. For example, hMSCs and their vesicles are primed by hypoxic conditions or changes in medium composition, such as the removal of FCS or the addition of priming factors like IFNγ and TNFα [34, 35]. The yield of extracellular vesicles can also be increased by preparing spheroids that mimic
There is currently no standardized large-scale production platform for primed hMSC-derived vesicles, but even if such a platform existed, a corresponding purification process would be required. The laboratory-scale purification of vesicles captured from the culture medium is currently based on a combination of ultracentrifugation, dead-end filtration, precipitation, and size exclusion chromatography, which are difficult to scale up [94, 95]. However, tangential-flow filtration can also be used for large-scale purification, washing, and buffer exchange, and this method should be investigated in more detail for vesicle purification [94, 96]. Extracellular vesicles are even more sensitive to process changes than MSCs, so the influence of multiple cell-dependent, culture, and process parameters on the potency of these vesicles must be determined.
\nThe ability of MSCs to restore the activity of dysfunctional cells
The major challenge of cocultivation is to balance the demands of two completely different cell types. In large scales, the distribution of cells becomes heterogeneous, which can lead to instability within the bioreactor and lower cell viability. A well-balanced and tightly controlled culture environment is needed to stabilize large-scale cocultures. Because secreted factors are important for the cocultivation of MSCs and beta cells, the hydrodynamic forces in bioreactors, which influence the distribution of secreted molecules, must be considered at an early stage [100]. Furthermore, the optimal cocultivation ratio of the cells must be determined. Established processes can be modified to achieve a new process setup for cocultivation, but it is often beneficial to separate cell expansion from cocultivation (i.e., first expand the pure cultures to generate the cells needed for the coculture and then combine them to improve the function of beta cells in a second process step). For the expansion step, it can be sufficient to improve the growth of beta cells using conditioned medium from the cultivation of MSCs. Alternatively, the expansion and functionalization of beta cells can be combined in one process step [101]. The CPPs for such a complex process can be difficult to identify, but the CQAs of the beta cells are most relevant if the aim of the process is to produce functionalized beta cells for drug screening or cell therapy. Even so, the potency of the MSCs must not be neglected because they are required to stimulate the beta cells. Accordingly, the MSCs must be expanded under controlled and standardized conditions that maximize their beneficial impact on beta cells. In the future, cocultivation bioreactor concepts for MSCs and beta cells must be tested to allow the completely aseptic expansion and cocultivation of both cell types.
\nMSCs are potent therapeutic agents, but their complexity and environmental sensitivity make the GMP-compliant manufacturing of MSC products extremely challenging. Given the range of tissue sources, isolation procedures, and expansion protocols, it is unclear whether MSC products are similar enough across manufacturing sites and whether results can be considered comparable even within the same study. Moreover, the incomplete definition of MSCs makes it difficult to develop objective release criteria. These issues strongly argue for the harmonization and standardization of MSC manufacturing processes, release criteria, and potency assays. The regulatory standards for MSCs are still evolving, and different standards apply in different jurisdictions. MSCs are living cells and cannot be held to the same standards as chemical entities or biopharmaceuticals, both of which can be tested against rigorous and objective quality criteria. The regulations for MSCs should be more flexible, acknowledging that each MSC product is developed for a specific indication, and unique platform technologies, CQAs, and CPPs may therefore be necessary for each manufacturing process. One of the most important platform technologies is the use of bioreactors for cell expansion, because this is the only current strategy that can bring MSC therapy into routine practice. MSCs can also be used as production aids for other products, including beta cells for drug screening or diabetes therapy, and novel biological agents such as extracellular vesicles. In the future, they could even be used for commodity products such as artificial meat. But in all these applications, a robust and scalable manufacturing process will be necessary.
\nWe would like to thank the Hessen State Ministry of Higher Education, Research and the Arts for the financial support within the Hessen initiative for scientific and economic excellence (LOEWE-Program, LOEWE Center DRUID (Novel Drug Targets against Poverty-Related and Neglected Tropical Infectious Diseases)). We also received financial support from the Strategic Research Fund of the THM (University of Applied Sciences Mittelhessen). The authors acknowledge Dr. Richard M Twyman for revising the paper.
\nThe authors declare no conflict of interest.
Water covers about 70 per cent of the Earth’s surface, makes up about 75 per cent of human body mass, and is the basic material that all living things need to live. The fact that water covers more than two-thirds of the Earth’s surface makes it hard to believe that it is a scarce resource and that less than 1% of the total water on this planet is readily accessible for drinking or other uses. Approximately 97% of the earth’s water is salt water contained in lakes or seas; just 3% is fresh water. However, 68 per cent of freshwater on Earth is enclosed in the Antarctic and Greenland ice caps (30%) while just 0.3 per cent is enclosed in surface waters, including lakes, rivers, reservoirs, springs and streams. Water quality can be defined by its physical, chemical, biological and esthetic characteristics (appearance and smell) as well as by its fitness for the beneficial uses it has in the past provided for human and animal drinking, for the promotion of a healthy aquatic life, for irrigation of the land and for recreation. A safe water ecosystem is when it meets the standard in term of water maintains a rich, diverse population of species and is conducive for the consumption of public health. Water is of course, the basic liquid medium for living matter; thus, it is uniquely vulnerable to contamination by living creatures, including those that cause disease to humans. Aquatic contamination occurs as a result of the introduction by humans of either direct discharges into the water body or indirect substances or/energy that may result in the degradation of the water quality of any water body that poses a danger to human health, harms living organisms and hinders aquatic activities such as fishing and polluted water quality with respect to its use. Contamination mechanisms including suspension, solution and biochemical alteration is not inherently separate and distinct from each other and all of these complex processes may only occur in water. However, growing anthropogenic activities, such as urbanization, Industries, agricultural waste, etc. and natural processes, reduce water quality and pose a danger to all modes of life. Most people live in underdeveloped countries still depend on unprotected/contaminated water sources as their primary sources of drinking water and at the same time, as their means of waste disposal, which can cause outbreaks of waterborne diseases. The discharge of industrial waste into water bodies constitutes approximately 62 per cent of the overall source of heavy metals such as lead (Pb), zinc (Zn), copper (Cu), nickel (Ni), cadmium (Cd) and chromium (Cr) [1]. It is important to write about contamination caused by heavy trace elements, since untreated waste materials discharged by industry or agriculture worldwide are very concerned about the current disposal of waste materials containing heavy metals such as mercury, cadmium, lead, copper and arsenic due to growing concentrations in many waters.
Poultry farms are one of the world’s leading sources of high-grade and palatable protein-rich food (eggs and meat) but domestic, industrial and agricultural poultry waste is regularly disposed of without treatment into water bodies, especially in most developing countries. Poultry farming is a lucrative global trade in animal husbandry that raises domesticated birds such as chickens, ducks, quails, pigeons, guinea fowl, turkeys and geese to produce meat or eggs for food originating in the agricultural period. According to the World Watch Institute, 74% of meat consumed worldwide is from poultry meat, and 68% of eggs are derived intensively from poultry, while more than 60 billion chickens are killed annually for consumption [2]. There is little doubt that the demand and therefore the production of poultry will continue to increase relative to the world population, the economy and also the increase in the production of poultry wastes. Poultry waste is used as manure in many fields, but when disposed of in a water body without treatment, it may cause significant problems for aquatic life due to the presence of heavy metals in it. Poultry waste as a mixture of different media involving feces, bedding materials, wasted feeds and feathers, represent favorable media for wide range of chemical and biological hazards include many food-borne pathogens like
Due to increased demand for livestock meats and eggs, there is also a need for increased use of trace elements (some of which are also heavy metals’) as nutritional supplements in poultry diets to boost feed quality, promote weight gain and prevent disease, resulting in increased concentration of trace elements added to poultry diets. However, poultry feeds, whether natural or locally sourced or improved by special manufacturing processes, have been reported to be affected by the content of heavy metals in the feed [4]. Many heavy metals are also added to poultry feed as supplements, including copper (Cu), manganese (Mn), iron (Fe), selenium (Se), zinc (Zn) which are important nutrients needed for various biochemical and physiological functions in species, and a lack of supply of these micronutrients results in a number of deficiency diseases or syndromes [5]. Iron and Cu are added to prevent anemia, selenium is added to prevent oxidative cell damage, and Zn and Mn are added to ensure proper egg shell deposition and feather growth [5]. Calcium (Ca2+) is added for bone formation, while in mature laying fowl the majority of dietary calcium is used for egg formation and plays a role in blood clotting and intracellular communication. Antioxidants are added to delay the deterioration of vitamins in poultry feed and tranquilizers may be used to keep flocks quiet in the house and during transport to another pen. A wide variety of antimicrobial drugs are commonly administered to poultry feed as prophylaxis and/or growth promoter and most of the oral applied antibiotics are poorly absorbed in the poultry gut, and then consequently those large amounts of antibiotics were excreted in feces and urine to the environment. Approximately 90% of the applied antibiotics might be excreted as the parent compound [6]. The most common antibiotics such as bactracin, chlortetracycline, monesin, tylosin, penicillin, chloramphenicol and virginiamycin can be applied to poultry feed to fight diseases, pests and increase the supply of certain nutrients that transferred through the food chain to humans that induce antimicrobial resistance in humans. Topical pesticides are used as a repellent against flies, lice, bugs, mice and reptiles that can harm or destroy them. WHO/FAO [7], NRC [8], EU [9, 10, 11, 12] and SON [13] set acceptable levels of metals in animals, but excessive or deficient use of these metals may lead to deformity in the body or to health problems, some of which may cause serious toxicity, which may lead to the death of the animal (Tables 1 and 2). However, pollutants from poultry waste can have detrimental environmental consequences (air, soil and water) if their waste is poorly handled or untreated prior to disposal in the aquatic setting. The disposal of waste produced by the poultry industry is a long-standing concern due to the contribution of nutrients or as a source of heavy metal contamination to our water bodies. Livestock manure may be used as fertilizer in the agricultural sector, it may also degrade the quality of the environment, especially surface and ground water, if it is not properly managed [14]. Untreated poultry waste can degrade water quality when discharged directly to surface water by runoff. The key environmental and health threats associated with animal waste are the introduction of toxins into water sources, such as nutrient limitation (nitrogen and phosphorus), organic matter, sediments, bacteria and heavy metals, which have harmful effects on the living organism and change the nature of the water. However, all mineral elements, whether considered to be necessary or potentially harmful, can have an adverse food impact on humans and animals if they are included in the diet at an overly high concentration [15]. Trace mineral bioavailability is characterized as the proportion of the component consumed that is used for biochemical or physiological purposes [16]. In order to have a high bioavailability, the mineral component must be readily absorbed and rapidly integrate by the body. Bioavailability is mainly influenced by the chemical form of the mineral or the amount in the diet, the amount in the body of the animal, the concentration of other minerals in the diet, its age and the physiological state of the animal to which it is fed. The risk lies in the accumulation of manure-borne metals, as they are not biodegradable and ultimately become phytotoxic, and the long-term use of poultry waste on the soil could lead to the accumulation of heavy/trace elements that increase the potential bioavailability and toxicity of metals in the environment. Such accumulation has the potential to limit soil function, contaminate water and cause toxicity to plants, animals and humans via the food chain. Their bioavailability is determined by physical, chemical and biological factors such as temperature, adsorption, sequestration, lipid solubility and water partition coefficients, whereas biological factors such as species characteristics, trophic interactions and biochemical/physiological adaptation also play an important role [17]. Poultry waste is more toxic than other animal waste due to the high concentration of heavy metals in poultry feed which is not directly absorbed by the body of the animal and egestion as a waste product, while the land application of poultry manure may result in the absorptions of toxicants by plants, animals and humans through absorption, ingestion, bioaccumulation or other processes.
Trace/heavy metals | FAO/WHO [7] and EU [9, 10, 11, 12] | National Research council [8] | |
---|---|---|---|
Metals requirement in total diet dry (mg/kg) | Metals requirement in normal diet (ppm) | Toxic level in total diet (ppm) | |
Cadmium | 1 mg/kg | ||
Chromium | 0.01 mg/kg | ||
Cobalt | 1 mg/kg | ||
Copper | 100 mg/kg | 6–8 ppm | 250–800 ppm |
Iron | 45–80 mg/kg | 50–80 ppm | 4,500 ppm |
Iodine | 0.3–0.4 ppm | 625 ppm | |
Lead | 1–5 mg/kg | ||
Manganese | 20–60 mg/kg | ||
Molybdenum | 3–5 ppm | 20–10 ppm | |
Mercury | 0.5 mg/kg | ||
Nickel | 0.05 mg/kg | ||
Selenium | 5–20 pm | ||
Zinc | 600 mg/kg | 40–75 ppm | 800–4,000 ppm |
Permissible limits of trace/heavy metals requirements as an additive in poultry feed.
Trace mineral | Egg Layer | Brolier | Brolier Breeder | Cockerel | |||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Chick Mash (0–8 Weeks) | Grower Mash (9–17 Weeks) | Layer 1 (18–45 Weeks) | Layer 2 (46–72 Weeks) | Pre-Starter (0–8 Days) | Starter (9–21 Days) | Finisher (22–42 Days) | Broiler Breeder Starter (0–8 Weeks) | Broiler Breeder Grower (9–17 Weeks) | Broiler Breeder Female 1 (18–45 Weeks) | Broiler Breeder Female 2 (46–72 Weeks) | Cockerel Starter (0–8 Weeks) | Cockerel Finisher (9 Weeks –Market) | |
Manganese (mg) | 60 | 60 | 60 | 60 | 60 | 30 | 30 | 60 | 60 | 90 | 90 | 60 | 60 |
Iron (mg) | 30 | 30 | 30 | 30 | 80 | 60 | 60 | 30 | 30 | 30 | 30 | 30 | 30 |
Copper (mg) | 6 | 6 | 6 | 6 | 5 | 4 | 4 | 6 | 6 | 12 | 12 | 6 | 6 |
Zinc (mg) | 60 | 60 | 60 | 60 | 40 | 35 | 35 | 60 | 60 | 100 | 100 | 60 | 60 |
Iodine (mg) | 0.5 | 0.5 | 0.5 | 0.5 | 0.4 | 0.4 | 0.4 | 0.5 | 0.5 | 0.5 | 0.5 | 0.5 | 0.5 |
Selenium (mg) | 0.3 | 0.3 | 0.3 | 0.3 | 0.2 | 0.1 | 0.1 | 0.3 | 0.3 | 0.3 | 0.3 | 0.3 | 0.3 |
The word heavy/or trace metals is sometimes used loosely, as they contain a number of metals, some of which are not heavy and some of which are not metals. Heavy metals are a wide class of inorganic chemicals that are harmful to both human and environmental health. Heavy metals are commonly referred to as metals with a minimum density of more than 5 gm/cm3 and adversely impacting the environment and living organisms. Heavy metals include all metals and metalloids except alkali and alkaline earth elements. Some heavy metals are necessary for enzymatic activity and can inhibit enzyme activity when natural concentrations are exceeded. Although some heavy metals are needed as micronutrients, they may be toxic at higher levels than their requirements. In addition, elements such as C, H, O, N, P, S, K, Ca and Mg are often required by majority of species in very small amounts. These elements are called trace elements, such as Fe, Mn, Cu, Co and Mo, and are usually considered to be necessary for most organisms, although V, B and Zn are confirmed to be essential in at least some cases. Most of these trace elements function in an enzyme or in an active group in an enzyme. Since heavy metals cannot be degraded, they are deposited, assimilated or incorporated into water, soil and marine organisms, causing heavy metal contamination in water bodies. Essentials include iron, copper, zinc, cobalt, manganese, chromium, molybdenum, selenium, tin, nickel and vanadium. The deficiency or elevation of these elements can affect the body’s normal physiological activities and biochemical processes, resulting in abnormal cell metabolism, development, reproductive disorder and severe oxidative. Non-essential metals are lead, cadmium and mercury. Cobalt, copper, chromium, iron, manganese, nickel, molybdenum, selenium, tin and zinc, sometimes known as trace metals. As a result, the majority of heavy metals, whether necessary or not, are potentially harmful to all living organisms, depends on many factors, such as dosage intake, species chemical composition, age of organisms, gender, genetic make-up and nutritional status of exposed individuals [17]. They have various effects on species depending on dosage exposure and durations of consumption: acute poisoning occurs when exposed to high doses over a short period of time, and chronic poisoning or bioaccumulation occurs when exposed to low doses over a long period of time. ‘Toxic metals, including ‘heavy metals, ‘are individual metals and metal products that have harmful human health effects either by direct or indirect exposure. Trace minerals or heavy metals used in animal feed are often expressed either as parts per million (ppm) or as milligrams per kilogram (mg/kg) of dietary dry matter. In very small quantities, many of these metals are required to sustain life and become toxic in large quantities. They can build ups in biological systems and become a major health hazard” [18]. The term heavy metal refers to any metallic chemical elements that have a comparatively high densities compared to water and are found in traces in different matrices. Their heaviness and toxicity are interrelated as heavy metals are capable of causing toxic or toxic at low concentrations and, if present in animal feed, pose significant health hazards to poultry meat consumers due to biomagnification effects in the body of the animal [19, 20, 21, 22]. Heavy metals are normal components of the earth’s crust that are not depleted or damaged in the atmosphere and are harmful to human health because they appear to be bioaccumulate for a long period of time, e.g. mercury (Hg), cadmium (Cd), arsenic (As), chromium (Cr), thallium (Tl) and lead (Pb). Bioaccumulation refers to the rise in the concentration of the chemical in the body of the organism over time as opposed to the chemical concentration in the atmospheres. Accumulation of compounds in the organism at any time taken up is processed faster than broken down (metabolized) or excreted. Toxicity could result from any heavy metal, but ten (10) of them are among the top twenty hazardous substances considered to be toxic by several agencies due to their health implications, including arsenic, cadmium, chromium, cobalt, copper, iron, lead, manganese, mercury and platinum [17, 23]. In recent decades, the levels of these metals have risen in our environments as a result of human inputs and activities [24, 25, 26]. There are 35 different metals that are of considerable concern to human health due to residential or industrial exposure. They are widely present in the environment and animal diet as a food supplement and are needed in small quantities to maintain good health, but in larger amounts they become harmful or unsafe due to their accumulation in the animal’s body over time and may cause serious illness or death. Considering the great variety of heavy metals in the environment, their concentration in various feed chains, it is difficult to achieve a lower level of toxicity than the detection limit for all elements in all products [27]. The European Union, the United States, Asia and other countries are aware of all these problems and as a result numerous laws have been implemented to regulate all heavy metal contamination, reduce the risk of human exposure in the food chain and develop detection methods to control these pollutants in the food chain [28].
Arsenic is used in poultry production for growth promotion and for controlling intestinal parasites in which they are fed with arsenic compound called roxarsone (3-nitro-4-hydroxyphenylarsonic acid) while three-quarters of arsenic in feed will be excreted out as poultry waste into environment [29]. Arsenic is a natural soil constituents with concentrations of up to 500 mg/kg. In its essential form, arsenic is insoluble in water, but many of the arsenates are highly soluble. Much if not all-natural water contains compounds of arsenic. Arsenic is the most common cause of acute heavy metal poisoning in adults and is number 1 in the Top 20 List of ATSDR. Arsenic can also be present in water sources worldwide, contributing to contamination of shellfish, cod and haddock. The target organs are the blood, kidney, central nervous, digestive and skin systems [30]. Arsenic is noted for its human toxicity when ingestion of as little as 100 mg typically results in serious poisoning and 130 mg has been shown to be fatal [31]. Several incidents have shown that arsenic in water can be carcinogenic, that skin and probably liver cancers are due to arsenic in drinking water [32, 33].
Lead is number 2 on the “Top 20 List.” for the ATSDR. Lead accounts for most cases of pediatric heavy metal poisoning [30]. Goal organs are bone, brain, blood, kidney, and thyroid gland [23, 34]. Some natural water contains as much as 0.8 mg/l of lead in solution [35]. These concentrations are also found in mountain streams that flow through limestone and galena. It causes acute and chronic toxicity and causes a wide variety of physiological, biochemical and behavioral dysfunctions in humans, animals and aquatic species. Addition of lead to the diet results in a dose-related rise in the concentration of Pb in different organs in the body of animals such as the kidney, blood stream, liver and tibia. It induces oxidative stress that suppresses growth efficiency and decreases feed intake and body weight loss.
The number 3 of ATSDR’s “Top 20 List” is mercury and naturally generated in the environment by degassing the earth’s crust, by volcanic emissions [36]. It is available in three forms: elemental mercury, organic and inorganic mercury. Atmospheric mercury is spread across the globe by winds and returns to the planet in runoff, collecting in marine food chains and fish in lakes [37]. Many researchers believe that dental amalgam could be due to a source of mercury toxicity. Mercurochrome and merthiolate are still in use in drugs, while algaecides are the main possible sources of mercury by inhalation. The organic form is readily absorbed in the gastrointestinal tract (90–100%); Less but nevertheless large amounts of inorganic mercury are absorbed in gastrointestinal tract (7–15%) and the target organs are majorly brain and kidneys [30].
Cadmium is a derivative from the smelting or mining activities of lead and zinc in environment and it occupied 7 position on ATSDR’s “Top 20 list.” It also used in nickel cadmium batteries production, PVC plastics, and paint pigments industries. It can also find in Cigarettes, as well as in soil as a result of insecticides, fungicides, sludge, and other commercial fertilizers that contain cadmium compound in agriculture or in reservoirs that contain shellfish. Other sources of cadmium contamination are from dental alloys, electroplating, engine oil and automobile exhaust. Inhalation of cadmium accounts for 15–50 per cent of assimilate into the respiratory tracts; 2–7 per cent of the ingested cadmium is absorbed into the gastrointestinal system while main target organs are the liver, placenta, kidneys, lungs, brain and bones [30]. Cadmium is moderately harmful to all species and is a cumulative toxin in mammals. In low concentrations, the use of trivalent chromium as an additive in animal diets may induce rapid growth for the animal in order to improve the quality of the meat produced, but often poultry owners may add trivalent chromium in excesses for rapid growth of their animals in order to obtain further value, which may have adverse effects on animals such as those injured and poisonous to the animal. It appears to be concentrated in the kidneys, liver, pancreas and thyroid of humans and other mammals. Humans can be exposed to this metal mainly through inhalation and ingestion, and can suffer from acute and chronic intoxication. Kar and Patra [38] reported that the Cd concentration sometimes increases in feeds, fodders, water bodies, and tissues of livestock which causes metabolic, structural, and functional changes of different organs of all animals. In poultry birds, bioaccumulation of Cd occurs in several organs mainly in the liver, kidney, lung, and reproductive organs due to its continuous exposure. Intake of Cd reduces growth and egg laying performance and feed conversion efficiency in poultry. Chronic exposure of Cd at low doses can also alter the microscopic structures of tissues, particularly in the liver, kidney, brain, pancreas, intestine, and reproductive organs due to increased contents of Cd in these tissues. Continuous Cd exposure causes increased oxidative stresses at cellular levels due to over-production of reactive oxygen species, exhausting antioxidant defense mechanisms. This leads to disruption of biologically relevant molecules, particularly nucleic acid, protein and lipid, and subsequently apoptosis, cell damage, and necrotic cell death. The histopatholocal changes in the liver, kidneys, and other organs are adversely reflected in hemogram and serum biochemical and enzyme activities.
Iron does not appear on the ATSDR’s “Top 20 List, “ but it is a heavy metal of concerns, particularly because ingesting dietary iron supplements may acutely poison young children. Uses of Fe as additives in feed formation have many disadvantages such as low bioavailability, high hydroscopicity and oxidative, high excretion and so on [39]. Iron deficiency is still a major problem in several segments of the livestock production causes microcytic, hypochromic anemia in chickens. Iron also plays a role in other enzymes involved in oxygen transport and the oxidative process, including catalase, peroxidases, flavoprotein enzymes and cytochromes. Approximately two-thirds of body iron is found in hemoglobin (red blood cells and myoglobin in the muscles), while 20% is present in labile forms in the liver, spleen and other tissues, with the remainder not available in tissues such as myosin and actmysin and in metalloenzymes. The iron in hemoglobin is essential for the proper function of every organ and tissue of the body. The iron requirement of chicks fed casein, dextrose, and isolated soybean protein concentrate-based diet was studied by Aoyagi and Baker [40]. Ingestion accounts for most of the toxic effects of iron because iron is absorbed rapidly in the gastrointestinal tract and other target organs are the liver, cardiovascular system, and kidneys [30].
Zinc plays an important role in biological process in animal including immune function, growth, development and reproduction. It is component of many enzymes contributing in the energy metabolism, protein synthesis and degradation biosynthesis of nuclei acids, carbon dioxide, transport and many more. Its performance major role as an antioxidant in diet, growth and development, production, immunity and stress related issues. It is important in animal diets formation because it influences economic profitability of egg modifying. Zinc has a beneficial impacts on the growth and reproduction of livestock. Due to the low zinc and copper contents of some home-grown feeds compared to guidelines and varying bioavailability, supplementation of these metals is essential for most livestock species and is usually added as mineral supplements to dairy rations [7, 9, 10]. Zinc deficiency causes growth retardation and irregular production of feathers in poultry animals. Feather spattering occurs towards the end of the feather while severity of the spattering ranges from no feathers on the wings and tail to minor defects in the growth of some of the barbels and the hog joint may be widened. Zinc deficiency can causes the long bones of the legs and wings to be shortened and thickened. Other signs include loss of appetite, decreased feed use quality, and death in extreme cases. Zinc deficiencies in the breeding diet decreases egg production and hatchability. Embryos developed in zinc-deficient eggs display a wide range of skeletal anomalies in the head, limbs and vertebrae. The hatched chicks will also not stand, eat or drink [41]. Proper zinc supplementation has been shown to be effective in reducing the early mortality of poultry animals and zinc supplementation is typically applied to animal diets in the form of zinc oxide or zinc sulfate. Latest comparisons of bioavailability in chicks suggest that feed grade zinc oxide has just 44–78 per cent of zinc sulfate availability when added to refined or functional diets [42, 43, 44]. Zinc toxicities can cause health problems, and prolonged consumption can also lead to negative side effects such as nausea and vomiting, loss of appetite, diarrhea, abdominal cramping and immunity. The risk associated with zinc deficiency could cause gastrointestinal diseases such as Crohn’s disease, decreased immunity, thinning of hair, decreased appetite, weight loss, skeletal malformations, poor bone mineralization, immunological dysfunction, mood disorders, dry skin, fertility problems and impaired wound healing, inadequate dietary intake, poor absorption, genetic mutations. Symptoms of extreme zinc deficiency include impaired growth and development, delayed sexual maturity, chronic diarrhea, impaired wound healing and behavioral problems [45, 46].
Nickel is an essential element required in low amount for animal growth and it is required for activities of vitamin B12 and biotin during metabolism of odd-chain fatty acids in animals [47]. Depending on the dose and length of exposure, as an immunotoxic and carcinogen agent, nickel can cause several health problems such as contact dermatitis, cardiovascular disease, asthma, lung fibrosis, and respiratory tract cancer [48, 49]. However, the exposure of human beings mainly concerns oral ingestion through water and food as nickel may be a contaminant in drinking water and/or food [50]. Although the molecular mechanisms of nickel-induced neurotoxicity are not yet clear, oxidative stress and mitochondrial dysfunction have a significant role to play. Mitochondrial nickel-induced damage can occur due to impaired mitochondrial membrane potential, decreased mitochondrial ATP concentration and degradation of mitochondrial DNA [51]. Nickel, high concentrations of which can affect human health badly, can accumulate on plants, animals, and soil.
Based on the study by Eloma et al. [56], which analyzed six potentially toxic elements (PTEs) from poultry feeds such as Cd, Cr, Cu, Pb, Mn, Ni and Zn, four feed forms (starter, grower, finisher and layer) from four producers coded A, B, C and D were sold in Ebony State, Nigeria. The mean concentrations of metals recorded from poultry feeds were as following: Chromium (11.9–7.90 mg/kg); Copper (5.10–7.91 mg/kg); Cadmium (0.49–0.76 mg/kg); Lead (7.17–9.47 mg/kg); Manganese (26.9–34.9 mg/kg); Nickel (3.80–6.50 mg/kg) and Zinc (27.8–38.4 mg/kg). The result of these findings was compared with European Union standard of PTEs maximum acceptable concentration in feed while Pb and Ni concentrations were above the maximum acceptable limits that is risk to human health. Thus, there is a need for continuous monitoring of feed compositions. Lead and Ni exceeded permissible limits by European Union in feed as stipulated, but the perilous elements such as Cr, Cu and Zn were also high in feed. There is however a need for continuous monitoring of feed compositions and also for the introduction of practices that will not introduce PTEs into the system. It also recommended that a proximate study be carried-out on poultry feeds to determine its moisture content, ash content, crude fiber, lipid, crude protein, carbohydrate and metabolizable energy [57, 58, 59].
Kabir and Bhuyan [60] were conducted to determine the heavy metal content of hens (
Korish and Attia [61] conducted research on heavy metal content in feed, litter, meat, meat products, liver and table eggs of chickens. Concentrations of heavy metals were examined in chicken meat, meat products, feed, litter, as well as laying hen eggs to track the regularity of this metals in the market products and their protection for human consumption as recommended daily allowance (RDA). Samples were collected from most popular poultry products in Saudi Arabia. A total of 45 samples from frozen broiler meat, fresh beef, liver, frankfurter and burger were collected from the same brand. However, 60 table eggs were collected from four different commercial brands while the edible parts of egg were analyzed to determine the levels of mineral elements present in it. In addition, 30 samples from different feed and litter were collected from the starter feed, grower feed, diets of layer broilers and laying hens. The findings showed that there were extensive amounts of most trace or heavy metals in the various meat sources while liver had the highest concentration of all elements examined, except for Co, Cr and Ni. The highest amount of Chromium concentration was recorded in fresh meat, followed by frozen meat. Trace or heavy metals such as Mn, Co, Ni and Pb were not detected in frozen or fresh meat. The chicken burger and the frankfurter samples have similar concentrations of trace/heavy metal except for Zn and Mn which had higher concentrations was observed in frankfurter compared to burger sample. There were significant differences between zinc concentration of the different sources of eggs. Fe was significantly higher in beef meat compared to poultry meat but the opposite trend for Zn was observed. All heavy metals concentration in were higher in liver than the eggs, except for Chromium while the burger had higher concentrations of Cu and Co. finally, it concluded that Cd, Pb, As and Se are not detected in chicken meat and eggs produced which indicate that no human hazards from these toxic elements. However, the liver had the highest concentration of all heavy metals examined, except for Cr, and the intake of Pb and Cd from the broiler liver was higher than the RDA for adults. Burgers and frankfurters, showed higher concentrations of Pb, Cd and Ni than chicken meat and table eggs, implying a potential human health danger. Therefore, in order to enhance the quality of poultry products for human consumption, adequate legislation is required to regulate the quality of poultry products, as well as feed/food and chicken litter. In addition, critical measurements should be used for the detoxification of heavy metals from waste. The relationship between the minerals in poultry production and the diet of poultry and poultry litter remains fertile for further study.
Study of Dahri et al. [62] on the investigation of concentrations of heavy metals; lead and chromium in chicken feed collected from commercial poultry feed markets and local poultry farms in Hyderabad Sindh. A total of eight samples of poultry feeds, four of which were commercial feed samples and four of which were local feed samples collected in polyethylene bags. The samples were analyzed using the Aurora Al1200 Atomic Absorption Spectrophotometer (AAS) for heavy metals; lead (Pb) and chromium (Cr). Relatively higher concentrations of lead (Pb) have been found in commercial feed samples. Data obtained from the present study for lead and chromium beyond the allowable limit, i.e., 0.05 and 0.1 ppm as recommended by WHO/FAO. Lead (Pb) and chromium (Cr) metals are important for the growth of poultry, but they may become toxic if the concentrations exceed the allowable limits. Excessive quantities of metals taken by animals make their way to the human body, which is extremely dangerous to human health. Heavy metal contamination is prevalent in the Hyderabad district and thus in the present report, the amount of poultry feed is alarming. The nutritional values of the feed are therefore calculated from the concentrations of lead and chromium above the allowable level in the feed content.
Aquatic ecosystems are highly complex, diverse and subject to a variety of internal and external relationships that are subject to change over time. Public health issues are among the pollutants that the concentration of heavy metals in marine environments enters humans through food chains. Heavy metal contamination may occur from many causes, but most generally results from metal purification, e.g., copper smelting and nuclear fuel preparation. Following the introduction of heavy metal pollutants into the flow, whether from natural or anthropogenic sources, they divide between aqueous (pore water and overlying water) and solid phases (sediment, suspended particulate matter and biota). Anthropogenic metals can persistently persist inside water bodies, or these elements are absorbed by silt, likely absorbed by animals, and accumulate in the food chain, beginning easily with plankton, such as filtering zooplankton, benthos, or fish, and eventually transferred to humans. Unlike organic contaminants that lose biodegradation toxicity, heavy metals cannot be degraded/decayed and thus pose a different form of remediation challenge. Heavy metals such as lead, mercury, iron, cadmium, aluminum and magnesium are found in water supplies. If these metals are found in the sediment, they enter the food chain through plants and aquatic animals.
The effects of heavy metals on marine species vary from a small drop in the rate of growth to death. Pollutants entering inshore waters and estuaries cause severe problems, causing significant harm to the life and activities of living aquatic species and also to the mass mortality of organisms. The gradual and irreversible accumulation of these metals in the various organs of the creatures of life contributes to long-term metal-related diseases due to their toxicity, endangering aquatic biota and other organisms [36]. Heavy metal pollution may have detrimental effects on the ecological balance of the recipient ecosystem and the diversity of marine species [63, 64, 65]. Among the animal species, fish are those that cannot avoid the adverse effects of these contaminants [66, 67, 68]. These metals are responsible not only for the deterioration of the water quality of the body, but also for the death of a variety of aquatic species [69]. The disposal of these wastes adversely affects water bodies, changes their chemical composition and causes harm to both humans and aquatic organisms [70, 71, 72]. These heavy metals (arsenic, cadmium, chromium, cobalt, copper, iron, lead, manganese, mercury and platinum) become toxic by accumulations of flora and fauna in the body tissues and then move through the food chain from fish to humans [73, 74]. Cadmium causes certain problems similar to those caused by mercury, which are much more harmful than mercury; Daphina, Scenedesmus and
Mercury emission experiments in aquatic environments indicate that recovery from pollution will take place within a limited period of time following the cessation of pollution input [81]. The embryonic and larval stages of marine organisms are typically the most susceptible periods of the life cycle for heavy metals and other toxicants. Copper accelerated the mortality of
The great concern lies in the excessive accumulation of macro-minerals (Ca, Si, Fe), trace elements (Cu, Mn, Zn, Se), heavy metals (Pb, Hg, Cd), medicinal drugs (antibiotics, coccidiostatics, sulfa drugs, etc.), anti-metabolites, insecticides, herbicides, wood preservatives, mycotoxins and hormones, harmful organisms transmittable other non-nutritional excretory via wastes to man. Poultry waste contains considerable amounts of nutrients (nitrogen and phosphorus) and other excreted substances such as hormones, antibiotics, harmful pathogens and heavy metals. Leaching and runoff of these substances has the potential to contamination both the surface water and/or nearby groundwater (Steinfeld et al., in [82]). Thus, increased outputs of phosphorus and nitrate to fresh water which can caused severe water quality problems like accelerate eutrophication in surface waters due to high inputs of organic substances and nutrients through runoffs which can result into accumulation pollution nutrient-sensitive ecosystems resulting in biodiversity losses such as fish kills due to hypoxia/anoxia and high levels of ammonia, harmful toxic algal blooms, decreases in water clarity, widespread anoxia, declines in submerged aquatic vegetation, shifts in pH, and depletion of oxygen. A drop in the level of dissolved oxygen in surface water has deleterious effects on fish populations [83, 84]. Furthermore, eutrophication can spur the growth of toxic microorganisms, such as
Poultry waste contains toxic metals that are bioaccumulate in the body of aquatic organisms and become biomagnification through food chain to next trophic level which can cause health hazard to human such as arsenic which is carcinogen and may also lead to heart disease, diabetes, and a decline in mental functioning. These harmful bacteria and chemicals present in poultry waste threaten the human health and aquatic organisms globally. Also of concern is the issue of air quality affected by dust particles, releases significant emissions of gases (methane, hydrogen sulfides, sulfur dioxide and ammonia) offensive odors and other pollutants such as volatile organic compounds (VOCs), particulate matter (PM), nitrogenous compounds during the decomposition of poultry waste that contributed to climate change which is a global concern and deleterious health effects (both chronic and acute) including respiratory conditions (i.e., bronchitis, asthma in children), heart disease, and lung cancer [85, 86]. Poultry waste as an important source of nutrients for many edible crops, may also contain some biological hazards that can threaten human health [87]. Poultry waste could be a source of human pathogens such as Salmonella, Campylobacter and Listeria that can potentially contaminate both edible crops and environment, which consequently leads to food-borne diseases [88, 89]. Poultry litter contains wide and diverse counts of microorganism including both of gram positive and negative bacteria. Among the bacterial and fungal species that are biological hazard to human health was recorded from poultry litter and waste such as
Heavy metal contamination is known to cause numerous diseases worldwide, such as minamata disease (organic mercury poisoning), iItai-itai disease (cadmium poisoning), arsenic acid poisoning, and asthma induced air pollution (Matsuo, 2003). The worst examples of accumulation are recorded in Japan, where hundreds of fishermen were killed by consuming fish containing too much mercury (Minamata Disease, [37]) or cadmium (itai-itai disease Kobayashi, 1971). These tragedies resulted from the tragic coincidence of a predominantly fish-eating and fish-eating population, which while having high concentrations of mercuries, did not show any symptoms. A special feature makes the mercury issue more important because under anaerobic conditions after sedimentation, it can be transformed to yet more toxic methylmercury compounds, which like metals, may accumulate in organisms that are possibly adsorbed to –SH groups in enzymes and even in food chains. Conversion to methylmercury is a bacteriological conversion involving methane bacteria. As a part of this reaction, mercury adsorbed to sediments may be mobilized after they are settled (which would also contribute to anaerobic conditions) and accumulate in fish via food chains. Copper is another aspect that creates concern in contaminated water. Approximately 1 g of copper causes acute illness in humans. Some freshwater algae tend to require either cobalt or vitamin B12. Copper is also less harmful than mercury, and incidents like Minamata are not likely to occur, nor do drinking water appear to be poisoning. Copper disposal is so common and widespread, however that amounts of copper in aquatic environments can typically be high enough to cause harm to species. If humans eat food tainted with heavy metals and their concentrations are amplified because they cannot be excreted. If the concentration exceeds a lethal level, it can results in brain injury or death. Lead affects the central and peripheral nervous systems, organs, bones and kidneys. Lead does not have an advantageous biological role and is believed to store in the body. Lead acquaintance can cause antagonistic effects on human health, particularly in young children and pregnant women, as Pb is a neurotoxin that always disrupts normal brain development. It accumulates in the skeleton, induces bone mobilization during pregnancy, lactation exposure to fetuses and breastfed babies. Cellular and molecular lead can increase the incidence of carcinogenic events associated with DNA damage and suppress DNA repair and tumor controls. Lead is a toxic metal that is particularly harmful to children. Health issues caused by low levels of chronic exposure to heavy metals may take years for humans to develop and may be related to heavy metals [36]. Most of the exposure to heavy metal contamination has been clinically shown to be associated with causing free radical harm leading to: heart attacks, strokes, cancer and several circulatory disorders other than cardiovascular diseases that do not cause death, but may affect the quality of life. Any of these include: Impotence, Asthma, Diabetes, Exhaustion, Alzheimer’s Disease, Memory Loss. Lead poisoning is a severe, very common form of heavy metal poisoning and the symptoms of lead poisoning in children are close to those of attention deficit hyperactivity disorder (ADHD). Lead poisoning also 07 triggers behavioral and cognitive disabilities, nervousness, headaches, and many other associated symptoms.
Metals may be extracted from aqueous sources such as chemical precipitation, lime coagulation, ion exchange, reverse osmosis and solvent extraction [90]. Other methods include electrodialysis, ultrafiltration and biosorption; [36]. At present, phytoremediation (plants or microorganisms) is being used at an early stage to extract heavy metals from water, sediment and soil by concentrating them in their organic matter.
Increased regular releases of anthropogenic activities such as untreated poultry waste containing heavy metals into the aquatic environment globally, particularly in underdeveloped countries, could pose a risk to aquatic species as well as affect the ecological balance that can be transmitted via the food chain to humans and could pose serious human health problems. Regulation of the use of heavy metals as an additive or complete elimination of heavy metals in animal feed should be carried out in order to mitigate the human health risks associated with the use of animal products and the contamination of the atmosphere by manure. Preventive steps should be taken to minimize the level of heavy metal contamination in the marine ecosystems by destroying animals when the concentration is too high or consumed and transmitted to humans.
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Every year, the number of palm oil mills increases rapidly, thus increasing the capacity of fresh fruit bunch waste or effluent discharge. Based on the data from the Malaysian Palm Oil Board in 2012, Malaysia produced 99.85 million tons of fresh fruit bunch (FFB) per year. However, about 5–5.7 tons of water was required in order to sterilize the palm fruit bunches and clarify the extracted oil to produce 1 ton of crude palm oil resulting in 50% of the water turning into palm oil mill effluent (POME). POME is one of the major environmental pollutants in Malaysia. The characteristics of POME and its behavior, if discharged directly, in water are described in this chapter. The suspended solid and nutrient content in POME could be able to support the growth of algae. This chapter aims to demonstrate that POME could be used as a main source for algae production, and this effluent is one of the main environmental problems in the tropical region especially in Malaysia.",book:{id:"6730",slug:"palm-oil",title:"Palm Oil",fullTitle:"Palm Oil"},signatures:"Hesam Kamyab, Shreeshivadasan Chelliapan, Mohd Fadhil Md Din,\nShahabaldin Rezania, Tayebeh Khademi and Ashok Kumar",authors:[{id:"225957",title:"Dr.",name:"Hesam",middleName:null,surname:"Kamyab",slug:"hesam-kamyab",fullName:"Hesam Kamyab"},{id:"237449",title:"Dr.",name:"Shreeshivadasan",middleName:null,surname:"Chelliapan",slug:"shreeshivadasan-chelliapan",fullName:"Shreeshivadasan Chelliapan"},{id:"241504",title:"Dr.",name:"Mohd Fadhil",middleName:null,surname:"Md Din",slug:"mohd-fadhil-md-din",fullName:"Mohd Fadhil Md Din"},{id:"241505",title:"Dr.",name:"Shahabaldin",middleName:null,surname:"Rezania",slug:"shahabaldin-rezania",fullName:"Shahabaldin Rezania"},{id:"241506",title:"Dr.",name:"Tayebeh",middleName:null,surname:"Khademi",slug:"tayebeh-khademi",fullName:"Tayebeh Khademi"},{id:"241508",title:"Dr.",name:"Ashok",middleName:null,surname:"Kumar",slug:"ashok-kumar",fullName:"Ashok Kumar"}]},{id:"38711",doi:"10.5772/51238",title:"Hydrogen Storage for Energy Application",slug:"hydrogen-storage-for-energy-application",totalDownloads:12184,totalCrossrefCites:17,totalDimensionsCites:40,abstract:null,book:{id:"2873",slug:"hydrogen-storage",title:"Hydrogen Storage",fullTitle:"Hydrogen Storage"},signatures:"Rahul Krishna, Elby Titus, Maryam Salimian, Olena Okhay, Sivakumar Rajendran, Ananth Rajkumar, J. M. G. Sousa, A. L. C. Ferreira, João Campos Gil and Jose Gracio",authors:[{id:"25491",title:"Dr.",name:"Elby",middleName:null,surname:"Titus",slug:"elby-titus",fullName:"Elby Titus"}]},{id:"29876",doi:"10.5772/35875",title:"Petroleum Asphaltenes",slug:"petroleum-asphaltenes",totalDownloads:14174,totalCrossrefCites:25,totalDimensionsCites:40,abstract:null,book:{id:"2288",slug:"crude-oil-emulsions-composition-stability-and-characterization",title:"Crude Oil Emulsions",fullTitle:"Crude Oil Emulsions - Composition Stability and Characterization"},signatures:"Lamia Goual",authors:[{id:"106226",title:"Dr.",name:"Lamia",middleName:null,surname:"Goual",slug:"lamia-goual",fullName:"Lamia Goual"}]},{id:"60752",doi:"10.5772/intechopen.76412",title:"Biomaterial from Oil Palm Waste: Properties, Characterization and Applications",slug:"biomaterial-from-oil-palm-waste-properties-characterization-and-applications",totalDownloads:2816,totalCrossrefCites:22,totalDimensionsCites:38,abstract:"Oil palm are among the best known and most extensively cultivated plant families, especially Indonesia and Malaysia. Many common products and foods are derived from oil palm, its making them one of the most economically important plants. On the other hand, declining supply of raw materials from natural resources has motivated researchers to find alternatives to produce new materials from sustainable resources like oil palm. Oil palm waste is possibly an ideal source for cellulose-based natural fibers and particles. Generally, oil palm waste such as oil palm empty fruit bunches, oil palm trunk, oil palm shell and oil palm ash are good source of biomaterials. Lack of sufficient documentation of existing scientific information about the utilization of oil palm waste raw materials for biomaterial production is the driving force behind the this chapter. Incorporation of various types of biomaterial derived from oil palm waste resources as reinforcement in polymer matrices lead to the development of biocomposites products and this can be used in wide range of potential applications. Properties and characterization of biomaterial from oil palm waste will not only help to promote further study on nanomaterials derived from non-wood materials but also emphasize the importance of commercially exploit oil palm waste for sustainable products.",book:{id:"6730",slug:"palm-oil",title:"Palm Oil",fullTitle:"Palm Oil"},signatures:"Rudi Dungani, Pingkan Aditiawati, Sri Aprilia, Karnita Yuniarti, Tati\nKarliati, Ichsan Suwandhi and Ihak Sumardi",authors:[{id:"220081",title:"Dr.",name:"Pingkan",middleName:null,surname:"Aditiawati",slug:"pingkan-aditiawati",fullName:"Pingkan Aditiawati"},{id:"234728",title:"Dr.",name:"Rudi",middleName:null,surname:"Dungani",slug:"rudi-dungani",fullName:"Rudi Dungani"},{id:"249537",title:"Dr.",name:"Sri",middleName:null,surname:"Aprilia",slug:"sri-aprilia",fullName:"Sri Aprilia"},{id:"249539",title:"Dr.",name:"Karnita",middleName:null,surname:"Yuniarti",slug:"karnita-yuniarti",fullName:"Karnita Yuniarti"},{id:"249541",title:"Dr.",name:"Tati",middleName:null,surname:"Karliati",slug:"tati-karliati",fullName:"Tati Karliati"},{id:"249542",title:"Dr.",name:"Ichsan",middleName:null,surname:"Suwandi",slug:"ichsan-suwandi",fullName:"Ichsan Suwandi"},{id:"249543",title:"Dr.",name:"Ihak",middleName:null,surname:"Sumardi",slug:"ihak-sumardi",fullName:"Ihak Sumardi"},{id:"256251",title:"Dr.",name:"Sri",middleName:null,surname:"Hartati",slug:"sri-hartati",fullName:"Sri Hartati"}]},{id:"52155",doi:"10.5772/64828",title:"EOR Processes, Opportunities and Technological Advancements",slug:"eor-processes-opportunities-and-technological-advancements",totalDownloads:5447,totalCrossrefCites:16,totalDimensionsCites:33,abstract:"Enhanced oil recovery (EOR) processes are well known for their efficiency in incrementing oil production; however, the selection of the most suitable method to adopt for specific field applications is challenging. Hence, this chapter presents an overview of different EOR techniques currently applied in oil fields, the opportunities associated with these techniques, key technological advancements to guide the decision‐making process for optimum applicability and productivity and a brief review of field applications.",book:{id:"5143",slug:"chemical-enhanced-oil-recovery-ceor-a-practical-overview",title:"Chemical Enhanced Oil Recovery (cEOR)",fullTitle:"Chemical Enhanced Oil Recovery (cEOR) - a Practical Overview"},signatures:"Lezorgia Nekabari Nwidee, Stephen Theophilus, Ahmed Barifcani,\nMohammad Sarmadivaleh and Stefan Iglauer",authors:[{id:"37799",title:"Dr.",name:"Stefan",middleName:null,surname:"Iglauer",slug:"stefan-iglauer",fullName:"Stefan Iglauer"},{id:"179076",title:"Dr.",name:"Lezorgia",middleName:"Nekabari",surname:"Nwidee",slug:"lezorgia-nwidee",fullName:"Lezorgia Nwidee"},{id:"179077",title:"Prof.",name:"Ahmed",middleName:null,surname:"Barifcani",slug:"ahmed-barifcani",fullName:"Ahmed Barifcani"},{id:"179078",title:"Prof.",name:"Stephen",middleName:null,surname:"Theophilus",slug:"stephen-theophilus",fullName:"Stephen Theophilus"},{id:"189371",title:"Dr.",name:"Mohammad",middleName:null,surname:"Sarmadivaleh",slug:"mohammad-sarmadivaleh",fullName:"Mohammad Sarmadivaleh"}]}],mostDownloadedChaptersLast30Days:[{id:"52155",title:"EOR Processes, Opportunities and Technological Advancements",slug:"eor-processes-opportunities-and-technological-advancements",totalDownloads:5449,totalCrossrefCites:16,totalDimensionsCites:33,abstract:"Enhanced oil recovery (EOR) processes are well known for their efficiency in incrementing oil production; however, the selection of the most suitable method to adopt for specific field applications is challenging. Hence, this chapter presents an overview of different EOR techniques currently applied in oil fields, the opportunities associated with these techniques, key technological advancements to guide the decision‐making process for optimum applicability and productivity and a brief review of field applications.",book:{id:"5143",slug:"chemical-enhanced-oil-recovery-ceor-a-practical-overview",title:"Chemical Enhanced Oil Recovery (cEOR)",fullTitle:"Chemical Enhanced Oil Recovery (cEOR) - a Practical Overview"},signatures:"Lezorgia Nekabari Nwidee, Stephen Theophilus, Ahmed Barifcani,\nMohammad Sarmadivaleh and Stefan Iglauer",authors:[{id:"37799",title:"Dr.",name:"Stefan",middleName:null,surname:"Iglauer",slug:"stefan-iglauer",fullName:"Stefan Iglauer"},{id:"179076",title:"Dr.",name:"Lezorgia",middleName:"Nekabari",surname:"Nwidee",slug:"lezorgia-nwidee",fullName:"Lezorgia Nwidee"},{id:"179077",title:"Prof.",name:"Ahmed",middleName:null,surname:"Barifcani",slug:"ahmed-barifcani",fullName:"Ahmed Barifcani"},{id:"179078",title:"Prof.",name:"Stephen",middleName:null,surname:"Theophilus",slug:"stephen-theophilus",fullName:"Stephen Theophilus"},{id:"189371",title:"Dr.",name:"Mohammad",middleName:null,surname:"Sarmadivaleh",slug:"mohammad-sarmadivaleh",fullName:"Mohammad Sarmadivaleh"}]},{id:"60752",title:"Biomaterial from Oil Palm Waste: Properties, Characterization and Applications",slug:"biomaterial-from-oil-palm-waste-properties-characterization-and-applications",totalDownloads:2823,totalCrossrefCites:22,totalDimensionsCites:38,abstract:"Oil palm are among the best known and most extensively cultivated plant families, especially Indonesia and Malaysia. Many common products and foods are derived from oil palm, its making them one of the most economically important plants. On the other hand, declining supply of raw materials from natural resources has motivated researchers to find alternatives to produce new materials from sustainable resources like oil palm. Oil palm waste is possibly an ideal source for cellulose-based natural fibers and particles. Generally, oil palm waste such as oil palm empty fruit bunches, oil palm trunk, oil palm shell and oil palm ash are good source of biomaterials. Lack of sufficient documentation of existing scientific information about the utilization of oil palm waste raw materials for biomaterial production is the driving force behind the this chapter. Incorporation of various types of biomaterial derived from oil palm waste resources as reinforcement in polymer matrices lead to the development of biocomposites products and this can be used in wide range of potential applications. Properties and characterization of biomaterial from oil palm waste will not only help to promote further study on nanomaterials derived from non-wood materials but also emphasize the importance of commercially exploit oil palm waste for sustainable products.",book:{id:"6730",slug:"palm-oil",title:"Palm Oil",fullTitle:"Palm Oil"},signatures:"Rudi Dungani, Pingkan Aditiawati, Sri Aprilia, Karnita Yuniarti, Tati\nKarliati, Ichsan Suwandhi and Ihak Sumardi",authors:[{id:"220081",title:"Dr.",name:"Pingkan",middleName:null,surname:"Aditiawati",slug:"pingkan-aditiawati",fullName:"Pingkan Aditiawati"},{id:"234728",title:"Dr.",name:"Rudi",middleName:null,surname:"Dungani",slug:"rudi-dungani",fullName:"Rudi Dungani"},{id:"249537",title:"Dr.",name:"Sri",middleName:null,surname:"Aprilia",slug:"sri-aprilia",fullName:"Sri Aprilia"},{id:"249539",title:"Dr.",name:"Karnita",middleName:null,surname:"Yuniarti",slug:"karnita-yuniarti",fullName:"Karnita Yuniarti"},{id:"249541",title:"Dr.",name:"Tati",middleName:null,surname:"Karliati",slug:"tati-karliati",fullName:"Tati Karliati"},{id:"249542",title:"Dr.",name:"Ichsan",middleName:null,surname:"Suwandi",slug:"ichsan-suwandi",fullName:"Ichsan Suwandi"},{id:"249543",title:"Dr.",name:"Ihak",middleName:null,surname:"Sumardi",slug:"ihak-sumardi",fullName:"Ihak Sumardi"},{id:"256251",title:"Dr.",name:"Sri",middleName:null,surname:"Hartati",slug:"sri-hartati",fullName:"Sri Hartati"}]},{id:"66623",title:"Catalytic Dehydration of Glycerine to Acrolein",slug:"catalytic-dehydration-of-glycerine-to-acrolein",totalDownloads:1418,totalCrossrefCites:1,totalDimensionsCites:3,abstract:"The biodiesel production yields glycerine as a by-product in quantities around 10 vol% of produced biodiesel. Acrolein can be obtained from glycerine by a dehydration reaction. Catalytic processes in gas phase have been developed to obtain acrolein from a renewable feedstock using heterogeneous catalysts. The main process variables are the reaction temperature, the concentration of glycerol in water, and the space velocity in fixed-bed reactors. A thermodynamic study of the equilibrium has been made to estimate the conversion to equilibrium as a function of temperature. The reactors have been heated usually between 523 and 603 K. Generally, an aqueous glycerol solution is preheated in a preheating zone at a temperature enough to vaporize the feedstock, between 473 and 533 K, depending on the concentration of reactant required in the feed. Some of the most active catalysts in the gas-phase reaction (yield >70%) were NH4-La-β zeolite, Pd/LaY zeolite, hierarchical ZSM-5, WO3/ZrO2, WO3/TiO2, ZrOx-NbOx, WOx-NbOx, WO3-SiO2/ZrO2, NbOx-WOx/Al2O3, H3PO4-MCM-41, SAPO-40, NbPSi, Pd-H3PW12O40/Zr-MCM-41, H3PW12O40/Cs-SBA-15, H3PW12O40/Nb2O5, Cs-doped H4SiW12O40/Al2O3, H4SiW12O40/TiO2, and H4SiW12O40/SiO2.",book:{id:"8448",slug:"glycerine-production-and-transformation-an-innovative-platform-for-sustainable-biorefinery-and-energy",title:"Glycerine Production and Transformation",fullTitle:"Glycerine Production and Transformation - An Innovative Platform for Sustainable Biorefinery and Energy"},signatures:"Israel Pala Rosas, Jose Luis Contreras Larios , Beatriz Zeifert and José Salmones Blásquez",authors:[{id:"94936",title:"Dr.",name:"José Luis",middleName:null,surname:"Contreras",slug:"jose-luis-contreras",fullName:"José Luis Contreras"},{id:"284261",title:"Ph.D.",name:"Israel",middleName:null,surname:"Pala-Rosas",slug:"israel-pala-rosas",fullName:"Israel Pala-Rosas"},{id:"284262",title:"Dr.",name:"Jose",middleName:null,surname:"Salmones",slug:"jose-salmones",fullName:"Jose Salmones"},{id:"284263",title:"Dr.",name:"Beatriz",middleName:null,surname:"Zeifert",slug:"beatriz-zeifert",fullName:"Beatriz Zeifert"},{id:"295779",title:"Prof.",name:"Jose Luis",middleName:null,surname:"Contreras",slug:"jose-luis-contreras",fullName:"Jose Luis Contreras"}]},{id:"64816",title:"PVT Properties of Black Crude Oil",slug:"pvt-properties-of-black-crude-oil",totalDownloads:1561,totalCrossrefCites:0,totalDimensionsCites:1,abstract:"Precise PVT studies and behavior of phase-equilibrium of petroleum reservoir fluids are essential for describing these fluids and appraising their volumetric behavior at several pressure stages. There are numerous laboratory studies that can be performed on a reservoir sample. The amount of data desired determines the number of tests to be performed in the laboratory. Generally, there are three laboratory tests which characterize hydrocarbon fluids, namely primary study, constant mass depletion, and differential vaporization test. Generally, PVT properties are determined either experimentally or calculated theoretically through published correlations. This chapter presents different PVT laboratory tests that are required to understand the phase behavior of black oils.",book:{id:"7323",slug:"processing-of-heavy-crude-oils-challenges-and-opportunities",title:"Processing of Heavy Crude Oils",fullTitle:"Processing of Heavy Crude Oils - Challenges and Opportunities"},signatures:"Abdelaziz El-Hoshoudy and Saad Desouky",authors:[{id:"201556",title:"Dr.",name:"Abdelaziz",middleName:"Nasr",surname:"El-Hoshoudy",slug:"abdelaziz-el-hoshoudy",fullName:"Abdelaziz El-Hoshoudy"},{id:"210639",title:"Dr.",name:"Saad M.",middleName:null,surname:"Desouky",slug:"saad-m.-desouky",fullName:"Saad M. Desouky"}]},{id:"64885",title:"Environmental Challenges Associated with Processing of Heavy Crude Oils",slug:"environmental-challenges-associated-with-processing-of-heavy-crude-oils",totalDownloads:876,totalCrossrefCites:2,totalDimensionsCites:6,abstract:"The petroleum industry is one of the largest industries in the world and plays a pivotal part in driving a nation’s economy. However, the exploration and exploitation of heavy crude oil have raised series of environmental challenges and caused increased concern for the communities where the oil refineries are cited. Activities such as gas flaring and oil spillage have led to the release of toxic organic and inorganic pollutants, which has resulted in acid rain, climate change, and contamination of soil, water, and air. These environmental hazards have caused adverse effects directly or indirectly to the ecosystem. This chapter offers a general overview of the processes involved in the processing and some of the potential environmental challenges associated with heavy crude oil processing.",book:{id:"7323",slug:"processing-of-heavy-crude-oils-challenges-and-opportunities",title:"Processing of Heavy Crude Oils",fullTitle:"Processing of Heavy Crude Oils - Challenges and Opportunities"},signatures:"Samuel O. 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Biochemistry examines macromolecules - proteins, nucleic acids, carbohydrates, and lipids – and their building blocks, structures, functions, and interactions. Much of biochemistry is devoted to enzymes, proteins that catalyze chemical reactions, enzyme structures, mechanisms of action and their roles within cells. Biochemistry also studies small signaling molecules, coenzymes, inhibitors, vitamins, and hormones, which play roles in life processes. Biochemical experimentation, besides coopting classical chemistry methods, e.g., chromatography, adopted new techniques, e.g., X-ray diffraction, electron microscopy, NMR, radioisotopes, and developed sophisticated microbial genetic tools, e.g., auxotroph mutants and their revertants, fermentation, etc. More recently, biochemistry embraced the ‘big data’ omics systems. Initial biochemical studies have been exclusively analytic: dissecting, purifying, and examining individual components of a biological system; in the apt words of Efraim Racker (1913 –1991), “Don’t waste clean thinking on dirty enzymes.” Today, however, biochemistry is becoming more agglomerative and comprehensive, setting out to integrate and describe entirely particular biological systems. The ‘big data’ metabolomics can define the complement of small molecules, e.g., in a soil or biofilm sample; proteomics can distinguish all the comprising proteins, e.g., serum; metagenomics can identify all the genes in a complex environment, e.g., the bovine rumen. 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Dr. Blumenberg’s research is focused on the epidermis, expression of keratin genes, transcription profiling, keratinocyte differentiation, inflammatory diseases and cancers, and most recently the effects of the microbiome on the skin. He has published more than 100 peer-reviewed research articles and graduated numerous Ph.D. and postdoctoral students.",institutionString:null,institution:{name:"New York University Langone Medical Center",institutionURL:null,country:{name:"United States of America"}}},subseries:[{id:"14",title:"Cell and Molecular Biology",keywords:"Omics (Transcriptomics; Proteomics; Metabolomics), Molecular Biology, Cell Biology, Signal Transduction and Regulation, Cell Growth and Differentiation, Apoptosis, Necroptosis, Ferroptosis, Autophagy, Cell Cycle, Macromolecules and Complexes, Gene Expression",scope:"The Cell and Molecular Biology topic within the IntechOpen Biochemistry Series aims to rapidly publish contributions on all aspects of cell and molecular biology, including aspects related to biochemical and genetic research (not only in humans but all living beings). We encourage the submission of manuscripts that provide novel and mechanistic insights that report significant advances in the fields. Topics include, but are not limited to: Advanced techniques of cellular and molecular biology (Molecular methodologies, imaging techniques, and bioinformatics); Biological activities at the molecular level; Biological processes of cell functions, cell division, senescence, maintenance, and cell death; Biomolecules interactions; Cancer; Cell biology; Chemical biology; Computational biology; Cytochemistry; Developmental biology; Disease mechanisms and therapeutics; DNA, and RNA metabolism; Gene functions, genetics, and genomics; Genetics; Immunology; Medical microbiology; Molecular biology; Molecular genetics; Molecular processes of cell and organelle dynamics; Neuroscience; Protein biosynthesis, degradation, and functions; Regulation of molecular interactions in a cell; Signalling networks and system biology; Structural biology; Virology and microbiology.",annualVolume:11410,isOpenForSubmission:!0,coverUrl:"https://cdn.intechopen.com/series_topics/covers/14.jpg",editor:{id:"165627",title:"Dr.",name:"Rosa María",middleName:null,surname:"Martínez-Espinosa",fullName:"Rosa María Martínez-Espinosa",profilePictureURL:"https://mts.intechopen.com/storage/users/165627/images/system/165627.jpeg",institutionString:null,institution:{name:"University of Alicante",institutionURL:null,country:{name:"Spain"}}},editorTwo:null,editorThree:null,editorialBoard:[{id:"79367",title:"Dr.",name:"Ana Isabel",middleName:null,surname:"Flores",fullName:"Ana Isabel Flores",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bRpIOQA0/Profile_Picture_1632418099564",institutionString:null,institution:{name:"Hospital Universitario 12 De Octubre",institutionURL:null,country:{name:"Spain"}}},{id:"328234",title:"Ph.D.",name:"Christian",middleName:null,surname:"Palavecino",fullName:"Christian Palavecino",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0033Y000030DhEhQAK/Profile_Picture_1628835318625",institutionString:null,institution:{name:"Central University of Chile",institutionURL:null,country:{name:"Chile"}}},{id:"186585",title:"Dr.",name:"Francisco Javier",middleName:null,surname:"Martin-Romero",fullName:"Francisco Javier Martin-Romero",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bSB3HQAW/Profile_Picture_1631258137641",institutionString:null,institution:{name:"University of Extremadura",institutionURL:null,country:{name:"Spain"}}}]},{id:"15",title:"Chemical Biology",keywords:"Phenolic Compounds, Essential Oils, Modification of Biomolecules, Glycobiology, Combinatorial Chemistry, Therapeutic peptides, Enzyme Inhibitors",scope:"Chemical biology spans the fields of chemistry and biology involving the application of biological and chemical molecules and techniques. In recent years, the application of chemistry to biological molecules has gained significant interest in medicinal and pharmacological studies. This topic will be devoted to understanding the interplay between biomolecules and chemical compounds, their structure and function, and their potential applications in related fields. Being a part of the biochemistry discipline, the ideas and concepts that have emerged from Chemical Biology have affected other related areas. 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Behind these definitions are hidden all the aspects of normal and pathological functioning of all processes that the topic ‘Metabolism’ will cover within the Biochemistry Series. Thus all studies on metabolism will be considered for publication.",annualVolume:11413,isOpenForSubmission:!0,coverUrl:"https://cdn.intechopen.com/series_topics/covers/17.jpg",editor:{id:"138626",title:"Dr.",name:"Yannis",middleName:null,surname:"Karamanos",fullName:"Yannis Karamanos",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002g6Jv2QAE/Profile_Picture_1629356660984",institutionString:null,institution:{name:"Artois University",institutionURL:null,country:{name:"France"}}},editorTwo:null,editorThree:null,editorialBoard:[{id:"243049",title:"Dr.",name:"Anca",middleName:null,surname:"Pantea Stoian",fullName:"Anca Pantea Stoian",profilePictureURL:"https://mts.intechopen.com/storage/users/243049/images/system/243049.jpg",institutionString:null,institution:{name:"Carol Davila University of Medicine and Pharmacy",institutionURL:null,country:{name:"Romania"}}},{id:"203824",title:"Dr.",name:"Attilio",middleName:null,surname:"Rigotti",fullName:"Attilio Rigotti",profilePictureURL:"//cdnintech.com/web/frontend/www/assets/author.svg",institutionString:null,institution:{name:"Pontifical Catholic University of Chile",institutionURL:null,country:{name:"Chile"}}},{id:"300470",title:"Dr.",name:"Yanfei (Jacob)",middleName:null,surname:"Qi",fullName:"Yanfei (Jacob) Qi",profilePictureURL:"https://mts.intechopen.com/storage/users/300470/images/system/300470.jpg",institutionString:null,institution:{name:"Centenary Institute of Cancer Medicine and Cell Biology",institutionURL:null,country:{name:"Australia"}}}]},{id:"18",title:"Proteomics",keywords:"Mono- and Two-Dimensional Gel Electrophoresis (1-and 2-DE), Liquid Chromatography (LC), Mass Spectrometry/Tandem Mass Spectrometry (MS; MS/MS), Proteins",scope:"With the recognition that the human genome cannot provide answers to the etiology of a disorder, changes in the proteins expressed by a genome became a focus in research. Thus proteomics, an area of research that detects all protein forms expressed in an organism, including splice isoforms and post-translational modifications, is more suitable than genomics for a comprehensive understanding of the biochemical processes that govern life. The most common proteomics applications are currently in the clinical field for the identification, in a variety of biological matrices, of biomarkers for diagnosis and therapeutic intervention of disorders. From the comparison of proteomic profiles of control and disease or different physiological states, which may emerge, changes in protein expression can provide new insights into the roles played by some proteins in human pathologies. Understanding how proteins function and interact with each other is another goal of proteomics that makes this approach even more intriguing. Specialized technology and expertise are required to assess the proteome of any biological sample. Currently, proteomics relies mainly on mass spectrometry (MS) combined with electrophoretic (1 or 2-DE-MS) and/or chromatographic techniques (LC-MS/MS). MS is an excellent tool that has gained popularity in proteomics because of its ability to gather a complex body of information such as cataloging protein expression, identifying protein modification sites, and defining protein interactions. The Proteomics topic aims to attract contributions on all aspects of MS-based proteomics that, by pushing the boundaries of MS capabilities, may address biological problems that have not been resolved yet.",annualVolume:11414,isOpenForSubmission:!0,coverUrl:"https://cdn.intechopen.com/series_topics/covers/18.jpg",editor:{id:"200689",title:"Prof.",name:"Paolo",middleName:null,surname:"Iadarola",fullName:"Paolo Iadarola",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bSCl8QAG/Profile_Picture_1623568118342",institutionString:null,institution:{name:"University of Pavia",institutionURL:null,country:{name:"Italy"}}},editorTwo:{id:"201414",title:"Dr.",name:"Simona",middleName:null,surname:"Viglio",fullName:"Simona Viglio",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bRKDHQA4/Profile_Picture_1630402531487",institutionString:null,institution:{name:"University of Pavia",institutionURL:null,country:{name:"Italy"}}},editorThree:null,editorialBoard:[{id:"72288",title:"Dr.",name:"Arli Aditya",middleName:null,surname:"Parikesit",fullName:"Arli Aditya Parikesit",profilePictureURL:"https://mts.intechopen.com/storage/users/72288/images/system/72288.jpg",institutionString:null,institution:{name:"Indonesia International Institute for Life Sciences",institutionURL:null,country:{name:"Indonesia"}}},{id:"40928",title:"Dr.",name:"Cesar",middleName:null,surname:"Lopez-Camarillo",fullName:"Cesar Lopez-Camarillo",profilePictureURL:"https://mts.intechopen.com/storage/users/40928/images/3884_n.png",institutionString:null,institution:{name:"Universidad Autónoma de la Ciudad de México",institutionURL:null,country:{name:"Mexico"}}},{id:"81926",title:"Dr.",name:"Shymaa",middleName:null,surname:"Enany",fullName:"Shymaa Enany",profilePictureURL:"https://s3.us-east-1.amazonaws.com/intech-files/0030O00002bRqB9QAK/Profile_Picture_1626163237970",institutionString:null,institution:{name:"Suez Canal University",institutionURL:null,country:{name:"Egypt"}}}]}]}},libraryRecommendation:{success:null,errors:{},institutions:[]},route:{name:"profile.detail",path:"/profiles/334585",hash:"",query:{},params:{id:"334585"},fullPath:"/profiles/334585",meta:{},from:{name:null,path:"/",hash:"",query:{},params:{},fullPath:"/",meta:{}}}},function(){var e;(e=document.currentScript||document.scripts[document.scripts.length-1]).parentNode.removeChild(e)}()