Flexural and impact properties of PP and PP-bagasse biocomposites.
\r\n\tAnimal food additives are products used in animal nutrition for purposes of improving the quality of feed or to improve the animal’s performance and health. Other additives can be used to enhance digestibility or even flavour of feed materials. In addition, feed additives are known which improve the quality of compound feed production; consequently e.g. they improve the quality of the granulated mixed diet.
\r\n\r\n\tGenerally feed additives could be divided into five groups:
\r\n\t1.Technological additives which influence the technological aspects of the diet to improve its handling or hygiene characteristics.
\r\n\t2. Sensory additives which improve the palatability of a diet by stimulating appetite, usually through the effect these products have on the flavour or colour.
\r\n\t3. Nutritional additives, such additives are specific nutrient(s) required by the animal for optimal production.
\r\n\t4.Zootechnical additives which improve the nutrient status of the animal, not by providing specific nutrients, but by enabling more efficient use of the nutrients present in the diet, in other words, it increases the efficiency of production.
\r\n\t5. In poultry nutrition: Coccidiostats and Histomonostats which widely used to control intestinal health of poultry through direct effects on the parasitic organism concerned.
\r\n\tThe aim of the book is to present the impact of the most important feed additives on the animal production, to demonstrate their mode of action, to show their effect on intermediate metabolism and heath status of livestock and to suggest how to use the different feed additives in animal nutrition to produce high quality and safety animal origin foodstuffs for human consumer.
",isbn:"978-1-83969-404-2",printIsbn:"978-1-83969-403-5",pdfIsbn:"978-1-83969-405-9",doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!1,hash:"8ffe43a82ac48b309abc3632bbf3efd0",bookSignature:"Prof. László Babinszky",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/10496.jpg",keywords:"Technological Feed Additives, Feed Industry, Quality of Compound Feed, Non-Antibiotic Growth Promoter, Product Quality, Additive Enzymes, Digestibility of Nutrients, NSP Enzymes, Farm Animals, Livestock, Immunity, Microbiome",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:null,numberOfDimensionsCitations:null,numberOfTotalCitations:null,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"November 24th 2020",dateEndSecondStepPublish:"December 22nd 2020",dateEndThirdStepPublish:"February 20th 2021",dateEndFourthStepPublish:"May 11th 2021",dateEndFifthStepPublish:"July 10th 2021",remainingDaysToSecondStep:"2 months",secondStepPassed:!0,currentStepOfPublishingProcess:4,editedByType:null,kuFlag:!1,biosketch:"Professor Emeritus from the University of Debrecen, Hungary who authored 297 publications (papers, book chapters) and edited 3 books. Member of various committees and chairman of the World Conference of Innovative Animal Nutrition and Feeding (WIANF).",coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"53998",title:"Prof.",name:"László",middleName:null,surname:"Babinszky",slug:"laszlo-babinszky",fullName:"László Babinszky",profilePictureURL:"https://mts.intechopen.com/storage/users/53998/images/system/53998.jpg",biography:"László Babinszky is Professor Emeritus of animal nutrition at the University of Debrecen, Hungary. From 1984 to 1985 he worked at the Agricultural University in Wageningen and in the Institute for Livestock Feeding and Nutrition in Lelystad (the Netherlands). He also worked at the Agricultural University of Vienna in the Institute for Animal Breeding and Nutrition (Austria) and in the Oscar Kellner Research Institute in Rostock (Germany). From 1988 to 1992, he worked in the Department of Animal Nutrition (Agricultural University in Wageningen). In 1992 he obtained a PhD degree in animal nutrition from the University of Wageningen.He has authored 297 publications (papers, book chapters). He edited 3 books and 14 international conference proceedings. His total number of citation is 407. \r\nHe is member of various committees e.g.: American Society of Animal Science (ASAS, USA); the editorial board of the Acta Agriculturae Scandinavica, Section A- Animal Science (Norway); KRMIVA, Journal of Animal Nutrition (Croatia), Austin Food Sciences (NJ, USA), E-Cronicon Nutrition (UK), SciTz Nutrition and Food Science (DE, USA), Journal of Medical Chemistry and Toxicology (NJ, USA), Current Research in Food Technology and Nutritional Sciences (USA). From 2015 he has been appointed chairman of World Conference of Innovative Animal Nutrition and Feeding (WIANF).\r\nHis main research areas are related to pig and poultry nutrition: elimination of harmful effects of heat stress by nutrition tools, energy- amino acid metabolism in livestock, relationship between animal nutrition and quality of animal food products (meat).",institutionString:"University of Debrecen",position:null,outsideEditionCount:0,totalCites:0,totalAuthoredChapters:"1",totalChapterViews:"0",totalEditedBooks:"0",institution:{name:"University of Debrecen",institutionURL:null,country:{name:"Hungary"}}}],coeditorOne:null,coeditorTwo:null,coeditorThree:null,coeditorFour:null,coeditorFive:null,topics:[{id:"25",title:"Veterinary Medicine and Science",slug:"veterinary-medicine-and-science"}],chapters:null,productType:{id:"1",title:"Edited Volume",chapterContentType:"chapter",authoredCaption:"Edited by"},personalPublishingAssistant:{id:"185543",firstName:"Maja",lastName:"Bozicevic",middleName:null,title:"Ms.",imageUrl:"https://mts.intechopen.com/storage/users/185543/images/4748_n.jpeg",email:"maja.b@intechopen.com",biography:"As an Author Service Manager my responsibilities include monitoring and facilitating all publishing activities for authors and editors. From chapter submission and review, to approval and revision, copyediting and design, until final publication, I work closely with authors and editors to ensure a simple and easy publishing process. I maintain constant and effective communication with authors, editors and reviewers, which allows for a level of personal support that enables contributors to fully commit and concentrate on the chapters they are writing, editing, or reviewing. I assist authors in the preparation of their full chapter submissions and track important deadlines and ensure they are met. I help to coordinate internal processes such as linguistic review, and monitor the technical aspects of the process. As an ASM I am also involved in the acquisition of editors. 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Venkateswarlu",coverURL:"https://cdn.intechopen.com/books/images_new/371.jpg",editedByType:"Edited by",editors:[{id:"58592",title:"Dr.",name:"Arun",surname:"Shanker",slug:"arun-shanker",fullName:"Arun Shanker"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"878",title:"Phytochemicals",subtitle:"A Global Perspective of Their Role in Nutrition and Health",isOpenForSubmission:!1,hash:"ec77671f63975ef2d16192897deb6835",slug:"phytochemicals-a-global-perspective-of-their-role-in-nutrition-and-health",bookSignature:"Venketeshwer Rao",coverURL:"https://cdn.intechopen.com/books/images_new/878.jpg",editedByType:"Edited by",editors:[{id:"82663",title:"Dr.",name:"Venketeshwer",surname:"Rao",slug:"venketeshwer-rao",fullName:"Venketeshwer Rao"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}},{type:"book",id:"4816",title:"Face Recognition",subtitle:null,isOpenForSubmission:!1,hash:"146063b5359146b7718ea86bad47c8eb",slug:"face_recognition",bookSignature:"Kresimir Delac and Mislav Grgic",coverURL:"https://cdn.intechopen.com/books/images_new/4816.jpg",editedByType:"Edited by",editors:[{id:"528",title:"Dr.",name:"Kresimir",surname:"Delac",slug:"kresimir-delac",fullName:"Kresimir Delac"}],productType:{id:"1",chapterContentType:"chapter",authoredCaption:"Edited by"}}]},chapter:{item:{type:"chapter",id:"63434",title:"Biocomposites from Colombian Sugarcane Bagasse with Polypropylene: Mechanical, Thermal and Viscoelastic Properties",doi:"10.5772/intechopen.80753",slug:"biocomposites-from-colombian-sugarcane-bagasse-with-polypropylene-mechanical-thermal-and-viscoelasti",body:'\nSince several decades ago, biocomposites have emerged as an option aimed to solve several issues within the composite materials science. In most of published cases in the literature, the use of natural fibers in combination with polymers is carried out to achieve some degree of reinforcement from the fibers to the polymer. Many studies report the use of natural fibers such as flax, hemp, jute, sisal, coconut fiber, banana, and fique, among many others [1], using an extensive variety of polymer matrices like polyethylene [2, 3], polypropylene [4], polystyrene [5], polyester resins [6], and natural rubber [7]. Clear effects have been seen in the improvement of mechanical performance. For example, usually Young’s modulus and tensile and flexural strength increase when natural fibers are compounded in percentages from 10 to 40%, which make the composites stiffer than its matrix counterpart [8]. Also, improvement in impact strength has been observed [9]. These findings give to the natural fibers a real opportunity to replace to some extent the use of fiberglass, aramid, and other synthetic fibers usually used for polymer reinforcement since, on top of their reinforcement ability, natural fibers are also cheap and have a much lower density than fiberglass, as previously stated in literatures [10, 11, 12]. However, the interest in utilization of natural fibers in biocomposites goes beyond their advantages for formulating new and mechanically improved materials. Interest is also driven by a global concern about the impact of plastics in the environment and a growing consciousness of the need for establishing a circular economy where residues like biomass and lignocellulosic can be valued and used as new raw material for industrial processes [13, 14]. In that regard it makes sense to use the agroforestry residues of extensive crops in a way that results in a neutral CO2 process, like the fabrication of composites, instead of using them for energy production through combustion. An example of the potential of lignocellulosic materials is the region of Valle del Cauca in the South West of Colombia, which has a large influence of the sugarcane industry. It produces 80% of all Colombian sugar, and also it counts for 50% of all local agricultural production. This industry produces a lot of agroforestry residues, approximately 6 million tons of sugarcane bagasse a year [15]. The availability and low cost of this residue are thought as competitive advantages for the development of sustainable biocomposites in this region. That is the main reason behind of our resent research: the valorization of sugarcane residues by their use in natural fiber reinforced polymer composites (NFPC). In regard to the use of sugarcane bagasse, some studies have reported its use as reinforcement for polypropylene, polyester, recycled PET, PVC, HIPS, and HDPE and as agents and/or compatibilizing treatments such as aluminates and mercerization (NaOH treatment) and the use of ethylene and methyl acrylate as copolymers and benzyl chloride [16, 17, 18, 19]. However, to the best of our knowledge, there have been no reports of the use of silanes as compatibilizing agents in sugarcane bagasse microfibers, in order to improve their adhesion to polymer matrices. In this chapter, polypropylene bagasse (PP bagasse) biocomposites prepared through extrusion, injection, and thermocompression molding will be evaluated. The morphology as well as the mechanical, thermal, and thermomechanical properties of the biocomposites will be investigated with the aim to understand the effect of the chemical treatments of the bagasse fibers on the polypropylene (PP) matrix properties.
\nSugarcane bagasse was obtained from a local sugar mill and kindly provided by Sucromiles S.A. Hexadecyltrimethoxysilane and sodium hydroxide were analytical-grade reagents from Aldrich (Wisconsin, USA). Absolute ethanol was a product from Merck (PA, USA). Polypropylene homopolymer 01H41 was obtained from Essentia (Cartagena, Colombia).
\nSugarcane as received was cleaned with distilled water in order to take off soil and residues from the lignocellulosic material. Clean bagasse was later dried at 60°C for 6 h until constant weight. Around 500 g of bagasse was then grounded using a Kinematica™ Polymix™ PX-MFC 90 D Lab mill drive and a sieve size of 200 μm. The sample was divided in three groups: one used as it was obtained after milling with no further treatment. A second group was treated with an aqueous solution of 8% NaOH using a 1:1 bagasse/solution ratio during 2 h, in order to remove lignin from the bagasse’s surface. A third group of fibers were silanized after lignin removal. For silanization, a solution of 2 × 10−3 M of octadecyltrimethoxysilane in an 8:2 ethanol/water ratio as solvent was prepared. The pH of solution was kept at 3 using acetic acid. A time of 10 minutes was allowed for hydrolysis after addition of silane and before the solution was sprayed over bagasse fibers using a plastic spray bottle. Wet fibers were allowed to dry at 90°C for 24 h in a forced air oven. After drying, fibers were kept in plastic bags until used in the composition process with polypropylene.
\nEach group of fibers was characterized by thermal gravimetric analysis (TGA) using a nonreactive atmosphere (N2 at 50 mL/min) from 25 to 650°C at a heating rate of 10°C/min using a TGA/DSC 2 STAR system, from Mettler Toledo. Surface structure of fibers was characterized by scanning electron microscopy (SEM), and chemical maps are also obtained by electron dispersed spectroscopy (EDS) using an ultra-high-resolution analytical FE-SEM SU-70 from Hitachi. All samples were sputtered with gold before analysis. Silicon content on fibers was quantified by flame atomic absorption spectrometry (FAAS). Around 0.5 g of fibers was calcinated at 550°C for 4 h in porcelain crucibles. After calcination each sample was treated with 2 mL of HF (48–50%) and 98 mL of H2SO4 0.08 M. Samples were kept for 24 h in polypropylene containers for 24 h and then filtered. Quantification using Analist 800 from Perkin Elmer was performed using a nitrous oxide/acetylene flame.
\nBiocomposites were compounded in a counter-rotating twin screw extruder Thermo Scientific HAAKE™ Polylab. In all cases fibers were physically premixed with polypropylene pellets in a plastic bag using 20% of fiber in weight. About 500 g of fiber-polypropylene mix was introduced in the extruder at 70 rpm. A temperature gradient from 140 to 170°C from the feeder to the melting zone was used. The outcoming cord of composite from the extruder was pelletized using a rotating cutter which produced pellets of about 5 mm long.
\nAfter the pelletization process, the PP and PP-bagasse biocomposite samples were dried in an oven at 85°C followed by injection molding process at 180°C. A BOY XS (BOY Machines, Inc., USA) microinjection molding machine was used to prepare samples (3*12.7*60 mm) for flexural and impact tests. An injection pressure of 68 bar and a back pressure of 18 bar were used.
\nAlso, sheets of the different samples were obtained using a hot-plate press and a forced water circulation cooling system (LabPro 400, Fontijne Presses). To shape the specimens, stainless steel molds were used. The molding was conducted at a temperature of 185°C, with a pressure of 50 kN and a 15 minute cycle. Finally, the sheets were demolded and adjusted to the required dimensions for DMA tests (1.7*12.7*60 mm) using a computer numerical control router. Figure 1 shows the injected specimens of PP and a PP-bagasse biocomposite.
\nInjected specimens of PP and a PP-bagasse biocomposite.
Three-point bending flexural tests were performed with an INSTRON universal testing machine model 3366 according to the ASTM D 790–17. The tests were carried out on bars of rectangular cross section at 23°C and at a rate of crosshead motion in 1.3 mm/min. This rate was determined based on the dimensions of the specimen. Also, the distance between the supports was 50 mm, and the tests were conducted up to 5% strain. All the results were taken as the average value of five samples.
\nThe impact strength of PP and biocomposites were determined with an Izod Tinius Olsen impact pendulum equipped with a 4.53 N pendulum. Prior to the test, the materials were subjected to conditioning for 48 h at 50% relative humidity and a temperature of 25°C. The specimens were made following the standard ASTM D256, and the starting angle of the test was 55.80°. All the results were taken as the average value of five samples.
\nDifferential scanning calorimeter (DSC) and TGA test of the neat PP and biocomposites were carried out using a TGA/DSC 2 STAR system, from Mettler Toledo. DSC tests were carried out under nitrogen atmosphere (N2 at 50 mL/min) from 20 to 200°C at a scanning rate of 10°C/min, with a sample of 10 mg in aluminum pans. Melting temperatures (Tm) were determined from the first heating scans. TGA was carried out on 10 mg samples using a TA Q500 thermogravimeter at 10°C/min from 23 to 600°C under nitrogen flow. The thermal degradation temperatures considered were the onset of inflection (T0) and the temperature of maximum weight loss rate (Tmax).
\nPolymers and composites have a different response to mechanical loads in comparison with other materials. They can be studied as materials that in some cases behave as elastic solids and, in others, as viscous fluids. As such, its mechanical properties also depend on time, stress, and temperature. The present study of the viscoelastic performance was carried out in a DMA RSA-G2 with ACS-3 Air Chiller System. In order to identify the viscoelastic behavior of biocomposites, the following test modes were used:
\nTo begin the study of the viscoelastic response of biocomposites, the linear viscoelastic region for the PP matrix was identified. To find this region, strain sweeps were carried out at a defined temperature. The geometry used to perform these tests was three-point bending. A strain sweep test takes successive measurements with an increase in the strain. For these experiments, the RSA-G2 was programmed with a strain between 0.001 and 1%; the frequency was constant at 1 Hz. Measurements were made at 0, 30, and 60°C.
\nAfter finding the linear viscoelastic zone, temperature ramp tests were performed to observe the behavior of the PP matrix at different temperatures. These tests were performed between −60°C and 170°C, at 1 Hz, 3°C/min, and 0.01% of strain.
\nScanning electronic microscopy (SEM) of biocomposites was carried out on the cryogenic fracture surfaces of the specimens using a Quanta FEG 250 microscope operating at a voltage of 10 kV.
\nThe samples were previously sputter-coated with gold to increase their electric conductivity. The cross-sectional diameters of the dispersed phase were measured using ImageJ 1.8v (Wayne Rasband, National institutes of health, USA). Determinations were performed in different areas of the SEM images.
\nFlexural and impact properties of the materials were subjected to analysis of variance (ANOVA), and the Tukey’s test was applied at the 0.05 level of significance. All statistical analyses were performed using Minitab Statistical Software Release 12 (Pennsylvania, USA).
\nIn order to produce and tune a lignocellulosic material to improve the mechanical performance of natural fiber reinforced polymer composites (NFPC), it is very important to conceptualize adhesion as one of the most important factors to achieve such challenge [20, 21]. Adhesion on the polymer-fiber interface is said to follow one of the four common mechanisms: mechanical interlocking, electrostatic interactions, molecular entanglement, or chemical bonding [22]. Many commercial polymer-coupling additives like maleic and acrylate grafted thermoplastics work as adhesion enhancers in polyolefins by generating chemical bonds with the free alcohol functionality on the fiber’s cellulose [23]. In those aforementioned cases, the adhesion increases by conditioning the polymer to the fiber’s surface. Instead, when silanes are used to increase adhesion, in the fiber’s surface which is conditioned to interact with the olefin polymer matrix by promoting electrostatic interactions or chain molecular entanglement [24, 25].
\nWhen nonpolar silanes, like dodecyl, hexadecyl, or octadecyl triethoxysilanes, are used for fiber modification, there is a lowering effect of the fiber surface energy which increases compatibility with the matrix by matching the polarities [26]. A practical and quick way of estimating the surface energy of surfaces is by measuring the contact angle of the surface. However, in many NFPC applications, the size of the fibers used is in the range of micrometers, as shown in the granulometry in Figure 2, for the case of sugarcane bagasse fibers.
\nGranulometry of milled sugarcane fibers after lignin removal and silanization with hexadecyl triethoxysilane.
Measuring contact angle on rough surfaces, such as those formed for a bed of microsized fibers, can be challenging since the observed angle is the manifestation not just of the molecular interactions at the solid/liquid interphase but also of the microstructure of the surface (Figure 3).
\nSEM photograph of a silanized sugarcane fiber. Inset shows the rough surface of a bed of fibers used to measure contact angle of the fibers.
This effect is counted by the model of Wenzel which predicts that if a molecularly hydrophobic surface is rough, the appearance is that of an even more hydrophobic surface [27, 28]. An interesting evidence of this phenomenon in particulate fibers is shown in Figure 4, where the contact angle of several groups of sugarcane bagasse fibers treated with solutions of variable silane concentrations appears almost invariant (between 122° and 129°), even though the absorbed amount of silane changes nearly in one order of magnitude (5.5 × 10−5 to 1.04 × 10−4 mol of silane per gram of fiber).
\nEffect of silane absorbed on microsized sugarcane bagasse fibers and their contact angle.
Another critical factor to achieve a good reinforcing material made of small natural fibers is the generation of anchoring points on the rough surface in order to produce enough silanization. Natural fibers in contrast with fiberglass, for example, do not have a well-defined geometry and instead lack of the advantage of having a highly energetic surface prone to react during silanization. Fiberglass has a surface populated with free hydroxyl groups from Si-OH functionality, but natural fibers are usually covered by a nonreactive lignin layer. That is why for natural fibers, the surface area that will react with silanes is determined by a good process of lignin removal, using alkaline or oxidative solutions, which will expose cellulose at the natural fiber’s surface [29, 30].
\nThe performance of the delignification treatment can be estimated from thermal gravimetric analysis (TGA) of fibers. Figure 5 shows the TGA of sugarcane bagasse fibers before and after delignification with alkaline treatment and after silanization.
\n(a) TG and (b) DTG curves of bagasse fibers at heating rates of 10°C/min.
When a good delignification is carried out, fibers gain some thermal stability. As shown in Figure 5, the T0 for sugarcane bagasse goes from 262 to 282°C when lignin has been removed. Also, as noted in the DTG, the typical signal of hemicellulose around 290°C disappears [31]. Furthermore, after most of lignin goes away, it is possible to observe a cleaner DTG signal with no shoulders that make evident the presence of residual compounds in the fibers. With only cellulose, the maximum degradation (Tmax) in DTG occurs around 325°C. Silane presence also increases Tmax to 335°C, mostly due to the formation of refractory siloxane network after silanization. Additionally, as observed in the TGA results, when surface modification by silanization with hydrophobic moieties has occurred, there is a clear decrease of water evaporation after 60°C, since fibers absorb less water when silanized. Changes in water uptake can go from 5 to 1%. This result indicates that silanization process reduces water absorption of the fibers and may give resistance against fungal decay [32].
\nAnother factor that plays a role in the reinforcing ability of fibers is the distribution of silane on their surface. Few works have detailed how silane gets distributed in the rough surface of natural fibers. Using chemical maps from scanning electron microscopy coupled with energy dispersive X-ray spectroscopy (SEM-EDS), it is possible to survey the surface and locate silicon at specific locations on the fiber [33]. Figure 6 shows the chemical maps for oxygen and silicon, as an example of chemical mapping of silanized fibers.
\nSEM–EDX chemical maps of sugarcane bagasse fiber treated with hexadecyltrimethoxysilane after delignification with NaOH 8%.
From SEM–EDX spectra, the concentration of surface atoms can be estimated using the intensity of signals at the specific energies of each atom. In this case, sugarcane bagasse fibers modified with hexadecyltrimethoxysilane were interrogated for the content of oxygen, silicon, and carbon before and after silanization. Figure 7 reviews the results. Spectra revealed that atomic oxygen content changed from 28.57 to 17.44%, carbon from 70.86 to 81.67%, and silicon from 0.29 to 0.57% before and after silanization, respectively. The variations in the atomic content are in agreement with the process performed. For example, the increment in carbon content after silanization is due to the additional carbons brought to the surface by the long chains (C16) of hexadecyltrimethoxysilane. For oxygen instead, the surface atomic concentration becomes lowered since a not-significant amount of oxygen is added by silanes. Silicon, as expected, almost doubles his surface concentration.
\nPercentage of total atomic content of carbon, oxygen, and silicon at fiber surface (black) and after (gray) silanization, measured by SEM–EDX.
In general, thermal, morphological, and chemical characterization of fibers is necessary when lignocellulosic materials are prepared as reinforcing fillers. The knowledge of important factors like the degree of lignin removal, distribution of silane, and hydrophobic character of fibers are very important to ensure that the material will behave successfully during compounding with polymeric matrices and then to obtain suitable mechanical properties of biocomposites.
\nThe influence of bagasse fiber addition on the PP flexural and impact properties was evaluated. Table 1 presents the values of the flexural modulus, flexural strength, and impact strength of the materials. Biocomposites showed different mechanical properties, indicating that the treatments affect the fiber-matrix interaction.
\nSample | \nImpact and flexural properties* | \n||
---|---|---|---|
Flexural properties | \nImpact properties | \n||
Modulus (MPa) | \nStrength (MPa) | \nImpact strength (kJ/m2) | \n|
PP | \n1296 ± 70a | \n40.0 ± 0.7a | \n4.4 ± 0.5a | \n
PP-Bag | \n2069 ± 30b | \n48.0 ± 1.1b | \n4.2 ± 0.2a | \n
PP-Bag+NaOH | \n1847 ± 114c | \n43.3 ± 0.5c | \n5.1 ± 0.5a | \n
PP-Bag+NaOH+Silane | \n1505 ± 94d | \n38.6 ± 1.9a | \n6.2 ± 0.1b | \n
Flexural and impact properties of PP and PP-bagasse biocomposites.
Mean of five replications ± standard deviation.
Different letters (a–d) in the same column indicate significant differences (p < 0.05).
The results show that bagasse fiber incorporation induces a significant improvement of flexural properties of PP. For PP-bagasse and PP-Bag+NaOH biocomposite flexural modulus (FM) increased 60 and 42%, respectively. On the same way, flexural strength (FS) reached improvements of 20 and 8% compared to neat PP. For PP-Bag+NaOH+Silane, FM was enhanced 16%, respectively, in comparison with PP. However, the FS value was not significantly different (p ≥ 0.05). Similar results were reported by Cerqueira et al. [34] when they studied the morphology and mechanical properties of PP-bagasse biocomposites. The authors reported that biocomposites present higher FM and FS values in comparison with neat PP and suggested a good interaction under the compressive stresses developed in part of the transverse section of the biocomposite specimens during bending.
\nOn the other hand, the impact tests did not show significant differences between the PP matrix and the biocomposites PP bagasse and PP-Bag+NaOH. However, for PP-Bag+NaOH+Silane an increase of 40% was observed. This result shows that silanization increases the capacity of PP to absorb energy. This phenomenon can be explained by a possible energy absorption promoted by fracture mechanisms, which involve detachment, slippage, and fragmentation of the fiber. These mechanisms do not occur in neat PP and PP biocomposites without silanization.
\nFigure 8 shows the results of the strain sweep tests of the PP matrix. Images of the PP specimen are included at a strain of 0.01% (linear region) and 0.6% which corresponds to the nonlinear zone. In this zone it is observed that the specimen has been highly deformed. From these results a strain of 0.01% was used for subsequent temperature ramp tests.
\nStrain sweep test curves for neat PP at 0, 30, and 60°C.
Figure 9 shows the thermograms obtained in the DMA for the PP matrix and its biocomposites. In these graphs the values of the storage modulus (E’), loss modulus (E”), and tan delta are shown. Neat PP tan delta plot shows two relaxations located near 6°C (β relaxation or Tg) and 60°C (α relaxation) [35]. It is also observed that the values of E´ are temperature dependent. At 25°C the value of E´ is 2708 MPa, while at 75°C, this value is 1199 MPa, which represents a decrease of 55%.
\nDynamic mechanical analysis (DMA) curves of neat PP.
In the tan delta plot of the biocomposite PP-Bag (Figure 10), a Tg of 5.3°C is observed, while the α relaxation increased 17.5°C compared to the neat PP. Also, E’values at 25°C is 2454 MPa, while at 75°C, this value is 1169 MPa. E’values’ decrease in this temperature range was 52%. This result shows that the addition of bagasse fiber improves the stability of the storage module of the PP matrix with the temperature. The increase in the value of α relaxation and the stability of E´ with the temperature was also observed in biocomposites PP-Bag + NaOH and PP-Bag + NaOH+Silanes (Figures 11 and 12).
\nDynamic mechanical analysis (DMA) curves of PP-Bag biocomposite.
Dynamic mechanical analysis (DMA) curves of PP-Bag + NaOH biocomposite.
Dynamic mechanical analysis (DMA) curves of PP-Bag + NaOH+Silane biocomposite.
Figure 13 shows the summary of the temperature sweep tests for biocomposites. The tan delta graphics show that there are no significant changes in the Tg of the biocomposites compared to the PP matrix. The values of Tg range between 3.7°C and 6.5°C. Also, these graphs show a reinforcing effect in the PP-bag biocomposite. Tan delta values varied 19.8% compared to the PP matrix. In the case of biocomposite PP-Bag + NaOH, this variation was 32.64% while in the biocomposite PP-Bag + NaOH+ Silane was 32.95%.
\nDynamic mechanical analysis (DMA) curves of neat PP and its composites.
With temperature increase, a second peak is observed around 60°C for neat PP. This peak can be related to an alpha transition. In the case of biocomposites, this alpha transition can be spotted at higher temperatures. This suggests that the service temperature of the biocomposites with alkaline and silanized treatments would allow a better performance of the material. In this experiment observed that the addition of silane to bagasse does not generate an improvement in the viscoelastic properties compared to the alkalinization treatment. It is emphasized that the alkalization treatment generates an improvement against the damping. This improvement can be positive for biocomposite applications that require an enhanced mechanical performance against stresses produced by bending loads.
\nThe first heating runs of PP and PP-bagasse biocomposite were shown in Figure 14. Both samples exhibit an endothermic peak between 163 and 165°C corresponding to the melting of the PP matrix. These results indicate that the addition of the bagasse fibers does not disturb the melting processes of the PP matrix.
\nFirst heating DSC curves for neat PP and PP-bagasse biocomposites.
TG and DTG curves for PP and PP-bagasse biocomposites are shown in Figure 15a and b, respectively. Neat PP degradation occurs in a single-step process with an onset temperature (To) located at 371°C and a Tmax of 449°C. Regarding biocomposites, TG and DTG show that degradation occurs in a two-step process. The first degradation step is associated with the decomposition of fiber constituents with a To located between 264 and 311°C for neat bagasse and silane-modified bagasse, respectively. This result indicates that chemical treatments improve the thermal stability of bagasse fibers.
\n(a) TG and (b) DTG curves of neat PP and PP-bagasse biocomposites.
The second degradation step corresponds to the decomposition of PP matrix. As shown in Table 2, To increases between 51 and 53°C. Also, Tmax increase between 4 and 9°C in comparison to neat PP. This increment in the thermal stability of the biocomposites has been previously observed in different studies [36, 37], indicating that the incorporation of fibers in the material induces spherulite nucleation points, increasing the crystallinity of the polymer and improving its thermal properties.
\nSample | \nDegradation stage | \nT0 (°C) | \nTmax (°C) | \n
---|---|---|---|
PP | \n1 | \n371 | \n449 | \n
PP-Bag | \n1 | \n264 | \n353 | \n
2 | \n423 | \n455 | \n|
PP-Bag + NaOH | \n1 | \n310 | \n355 | \n
2 | \n422 | \n453 | \n|
PP-Bag + NaOH + Silane | \n1 | \n311 | \n355 | \n
2 | \n424 | \n458 | \n
Thermal degradation data of the samples at 10°C/min in nitrogen atmosphere.
To: onset of inflection of each stage in TG curves.
Tmax: peak of the maximum degradation rate in DTG curves.
Figure 16 shows SEM images of fractured surfaces from PP-Bag and PP-Bag+NaOH+Silane biocomposites. Gaps between the bagasse fibers and the surrounding PP matrix can be clearly observed inFigure 16a, which indicates a poor interfacial adhesion between the PP matrix and the bagasse fibers [38]. For Figure 16b, with the chemical treatments, we can see that the gaps between bagasse and PP were reduced significantly and exhibited improved interface for the composite. This result confirms that chemical treatments expose the bagasse fibers and provided links between the cellulosic fibers and the surrounding polymer long chains, which improved the interfacial property of the hydrophobic PP matrix and the hydrophilic bagasse.
\nSEM pictures for a) PP-Bagasse and b) PP-Bag + NaOH+Silane biocomposites.
The chemical composition and thermal behavior of neat and chemically modified sugar bagasse fibers were studied. The biocomposites of bagasse fiber incorporated into a PP matrix were prepared by a melt-extrusion, injection, and thermocompression processes. The effects of bagasse fibers and chemical modification on the properties of the biocomposites were explored. Flexural characterization showed that bagasse fiber incorporation induces a significant improvement of flexural properties of PP. Also, the impact tests showed that the addition of silanized bagasse increases the capacity of PP to absorb energy. The DMA experiments show that bagasse fiber addition improves the maximum service temperature of the PP matrix. It was also observed that silanization process didn’t improve the viscoelastic properties compared to the alkalinization treatment. However, the alkalization treatment generates an improvement against the damping of the PP matrix. Thermal studies show that bagasse fiber addition did not disturb the melting process and improves the thermal stability of the PP matrix. This study offers an environmentally friendly alternative for utilizing waste bagasse fiber generated by the sugar industry for the production of biocomposites.
\nThe authors acknowledge the Autónoma de Occidente University, Cali-Colombia, for the technical and financial support; the nanocharacterization center of Virginia Commonwealth University, Virginia-United States, for EDX and SEM spectra; Santiago de Cali University, Cali-Colombia, for its support in the use of FAAS; Servicio Nacional de aprendizaje (SENA), Cali-Colombia, for the financial support through the System of Research, Technological Development and Innovation (SENNOVA). In addition, we wish to thank the company “Sucromiles” Colombia, for providing the sugarcane bagasse.
\nThe authors of this manuscript declare that they do not hold any conflicts of interest that might have any bearing on research reported in their submitted manuscript.
Antimicrobial photodynamic therapy (aPDT) is becoming a treatment option in dental medicine in different areas such as the diagnosis of malignant transformation of oral lesions, the treatment of head and neck cancer, as well as the treatment of bacterial and fungal infections [1, 2].
\nIn periodontology, the conventional therapeutic approach for treating periodontal diseases consists of mechanical cleaning combined with chemical decontamination, or the use of antimicrobial therapy which can be applied systemically or locally. The mechanical debridement has its own limitations in removing all the infections, such as the difficulties in reaching deep pockets and, as a result, the etiological factors continue to damage the periodontal ligament. Also, when mechanical debridement is used frequently, it can cause damage of the root surface [3]. The limitations of the conventional periodontal therapy have shifted the focus towards aPDT, as an effective alternative treatment for periodontal diseases [4, 5, 6, 7, 8]. aPDT is having many advantages over conventional therapy. The main advantage is the fact that photosensitizer can be placed directly into the periodontal pocket and then activated with an optical fiber tip in order to kill microbial cells, without damaging the host tissues. This makes aPDT a safe procedure against periodontal microbiota [9, 10].
\nMany studies have demonstrated potential improvements after the use of aPDT in conjunction with mechanical debridement [11, 12, 13]. However, there are several studies that report different results [5, 14, 15, 16]. Atieh suggested as a result of his meta-analysis, potential improvements after aPDT combined with scaling and root planning in probing periodontal pocket depth (PPD) reduction and greater clinical attachment level (CAL) gain [13]. Similarly, in their study Sgolastra et al. reported that the combination of aPDT and conventional treatment provides additional benefits by reducing the PPD and increasing the CAL [11].
\nIn endodontics, aPDT is used for the disinfection of the root canal. Conventional endodontic treatment consists of a combination of mechanical cleaning and shaping of the canals, the use of disinfecting solutions for irrigation and the placement of medicaments in between appointments. Sometimes, due to the root canal anatomy it is difficult to completely disinfect the canals by using only mechanical and chemical decontamination methods [17, 18]. aPDT demonstrated promising results as an adjunct therapy for the root canal disinfection in many studies. Raymond et al. [17] evaluated the efficacy of the combination of conventional treatment with photodynamic therapy in vitro. Their results showed that the combination of both therapies is more effective than the use of traditional treatment alone. Rios et al. [19] in their study used a combination of light-emitting diode (LED) as a light source and toluidine blue O dye as a photosensitizer. They suggested that photodynamic therapy can be used as an adjunctive antimicrobial procedure in endodontics. Similarly in their clinical study, Bago et al. [20] demonstrated that aPDT when used as an addition to the conventional mechanical and chemical root canal cleaning, can lead to significantly more reduction of the bacteria and in some samples the total elimination of the bacteria.
\nPhotodynamic therapy is used also in oral and maxillofacial surgery due to its potential to be used as an anti-cancer treatment and its antimicrobial potential. Oral squamous cell carcinomas (SCC) are the most frequent tumors in the oral cavity [21]. Up to date the traditional methods for treating SCC have not been very successful in increasing the 5-year survival rate. Furthermore they cause different side effects such as mouth sore, jaw pain and difficulties in chewing or swallowing [22].
\nOne of the developing factors of oral SCC are considered to be the pre-malignant lesions such as erythroplakias and dysplastic leukoplakias. Around half of oral SCC cases are associated with leukoplakias [23]. The potential therapeutic possibilities of photodynamic therapy are not limited only for oral SCC and other head and neck cancers, but also against pre-malignant, primary, recurrent and metastatic lesions [24, 25]. PDT when compared to conventional treatments of these lesions, has an advantage due to its selective tumor destruction and minimal invasiveness without affecting the healthy tissues. In addition, it can combined with conventional therapy to increase the overall treatment success [26].
\nThe PDT antimicrobial potential in oral and maxillofacial surgery, is mostly used for the disinfection of soft tissue or bone during surgical interventions, as a preventive measure. In a study done by Neugebauer et al. [27] it was demonstrated that use of aPDT caused significantly lower incidence of alveolar ostitis. In a another study it was concluded that the effect of photodynamic therapy is almost the same as the effect of 2.5% NaOCl without causing adverse effects on surrounding tissues on periapical lesion model in vitro [28]. Batinjan et al. [29] showed that aPDT causes reduced postoperative wound swelling and decreased wound temperature after the removal of the impacted third mandibular molar.
\nPDT has recently also been used as an adjuvant therapy for the treatment of medication-related osteonecrosis of jaws (MRONJ), that is highly related to bisphosphonate-related osteonecrosis of the jaw (BRONJ). In a study done by Minamisako et al. [30], it was suggested that both low level laser therapy (LLLT) and PDT are beneficial in the clinical management of the MRONJ. Similarly, Rugani et al. [31] concluded that photodynamic therapy can be used as treatment option in the early stages of BRONJ or as an adjunct therapy when surgical intervention is indicated.
\nIn 1978. Brånemark presented two-stage threaded titanium implants in a root-form [32]. The concept of osseointegration of the implants was first brought during the 1950s and 1960s after observing bone growth in contact with titanium. Brånemark defined osseointegration as: “A direct connection between living bone and a load-carrying endosseous implant at the light microscopic level.” [33]. Since then, dental implants have become a long-term reliable treatment option for replacing missing teeth [34]. An ideal implant should have the following properties: biocompatibility, adequate toughness, strength, corrosion resistance, facture and wear resistance [35, 36, 37].
\nThe “gold standard” of dental implants are considered to be the implants produced from titanium and its alloys. Titanium has excellent biocompatibility and it was shown that long term surgical rates of titanium implants are excellent [38, 39]. However, due to their dark gray color sometimes the implants can reflect through the peri-implant soft tissue. This poses an esthetic challenge especially when a thin biotype of gingiva is present or when there is a resorption of the buccal lamina [39, 40]. Due to these reasons, many scientists have shifted their focus into producing ceramic implants [41].
\nThe infection around dental implants can be presented as peri-implant mucositis or peri-implantitis. Peri-implant mucositis is a reversible inflammatory process and it affects only the soft tissues around the dental implant. This is followed by reddening, swelling and bleeding on probing [42]. Peri-implantitis on the other hand affects both soft and hard tissues around the implant and as a result loss of supporting bone occurs [43]. The microbial etiology of peri-implantitis is well documented in literature [44]. The microorganisms found in peri-implantits are very similar to those found in advanced periodontitis [45, 46]. Most of them are spirochetes and non-motile anaerobic Gram-negative bacteria such as: Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans,, Prevotella intermedia, Tannarella forsythia, Treponema denticola etc. [47]. In the oral cavity the implant surfaces are colonized very rapidly by the bacteria, which leads to the formation of a bacterial biofilm on the implant surface. When peri-implanitits is in its early stages, there are no significant symptoms and most of the time it is diagnosed during routine dental check-up. It is of great importance to diagnose peri-implanitits in its early stages in order to prevent the progression of the disease and increase the chances for good osseointegration [48].
\nAccording to Teughels et al. [49], the quantity and quality of plaque formation and bacterial adhesion on implant surfaces is influenced by the chemical composition, and the surface roughness of the implant. Rough surfaces and those with greater surface free energy, accumulate more plaque. Furthermore, initial bacterial adhesion is attracted more to surfaces with high wettability and pits and grooves in the roughened surfaces. The formation of bacterial plaque in these surfaces is difficult to remove.
\nTo date, many treatment methods have been proposed for treating peri-implantitis. They can be grouped in two categories: resective and regenerative therapies [50]. The main goal of resective treatments is to eliminate the etiological factors of peri-implanittis and maintain optimal conditions. These treatments are mainly done by cleaning and decontaminating implant surfaces. Regenerative treatments aim to reconstruct the pre-existing hard and soft tissues by using bone substitute grafts, membranes and growth factors [50, 51]. Resective treatment of peri-implanitits is similar to the treatment of periodontitis and it consists of mechanical cleaning of the biofilm from the implant surface. This is of the utmost importance when treating peri-implanitits. During resective treatment, plastic curettes, air-powder abrasive or ablative lasers and ultrasonic scalers are used [52]. The main objective is to clean the surface which can stop the progression of the disease and increase the chances for re-osseointegration of the implant. However, due to the implant surface roughness, the bacterial adhesion and colonization is very difficult to remove and sometimes mechanical debridement alone is not very effective [53]. It has been suggested by some authors that the mechanical elimination of the implant threads and then smoothing the implant surface (implantoplasty) should be done, in order to improve the decontamination of the implant surface. In addition this procedure allows better maintenance and oral hygiene when threads are exposed to the oral environment [54]. When decontaminating the implant surface, the use of metallic curettes is not recommended due to the fact that they can alter the surface roughness of the implant and favor bacterial colonization. As an alternative, plastic curettes should be used because they do very little damage or none at all [55, 56].
\nRecently, as a treatment alternative, many scientists have shifted their focus towards the laser decontamination of the implant surfaces. In a study done by Kreisler et al. [57] the mechanical effects of Nd:YAG (Neodymium: yttrium-aluminum-garnet), Ho:YAG (Holmium: yttrium-aluminum-garnet), Er:YAG (Erbium: yttrium-aluminum-garnet), CO2 (Carbon dioxide) and GaAlAs (Gallium-Aluminum-Arsenide) lasers were evaluated, on different types of implant surfaces.. According to their results, Nd:YAG and Ho:YAG lasers cause significant damage to the implant surfaces, while CO2 and Er:YAG lasers when used in specific power settings do not cause any damage. GaAlAs laser did not damage the implant surface in any power settings. As an adjunct therapy to mechanical methods for treating peri-implantitis, the use of chemical decontamination and antibiotic therapy are being used with the aim of improving the treatment outcome. The most commonly used antimicrobial solutions are chlorhexidine, hydrogen peroxide, tetracycline or minocycline, citric acid, and phosphoric acid [58].
\nRecently aPDT has emerged as a new treatment option or adjuvant treatment to the conventional treatment of peri-implantitis. Its potential to decontaminate the implant surfaces without any damage to the implant or the surrounding tissues has generated a lot of interest in the scientific community. In addition aPDT is more effective than the use of lasers alone [53]. In their study, Hayek et al. [59] demonstrated that aPDT is both effective and non-invasive method when compared to traditional therapy during surgical treatment of peri-implantitis with elevated mucoperiosteal mucosal flaps. These beneficial characteristics of aPDT make it as promising novel and non-invasive method which can be used alone or as an adjunct therapy of peri-implantitis. [2].
\nThere are many in vitro studies evaluating the efficacy of photodynamic therapy against causative bacteria of peri-implantitis. The aim of our research was to evaluate the efficacy of aPDT on titanium and zirconia dental implants. For this purpose three different devices in combination with photosensitive dye were used. In addition, our aim was to evaluate if aPDT causes damage and alteration to the implant surfaces which would interfere with the re-osseoinegration of the implants in the clinical conditions.
\nThe study sample consisted of 144 sterile dental implants (72 titanium dental implants and 72 zirconia dental implants) (Bredent®, Senden, Germany). Both, titanium and zirconia dental implants were with a diameter of 4.0 mm and 12 mm of length, with sandblasted and acid etched surface. Each of the implants was in an unopened sterile packaging.
\nA bacterial suspension was prepared from three bacteria species: Prevotella intermedia, Aggregatibacter actinomycetemcomitans, and Porphyromonas gingivalis. These bacteria are commonly found in peri-implant diseases.
\nThe bacteria were cultivated separately in Columbia Agar for 72 hours and then, using thioglycolate broth, a bacterial suspension was prepared for each of the bacteria. The suspension of each of the bacteria was then mixed together in a joint suspension.
\nIn a single use tubes 300 μl of the bacterial suspension was put and then each implant was put separately in single use tubes (Figure 1). The tubes were incubated in anaerobic conditions for 72 hours.
\nImplants placed in Eppendorf tubes containing bacterial suspension. Implants covered in their entire length by the bacterial suspension.
Wavelength: 660 nm | \n
Fiber tip: 320 μm optical fiber tip | \n
Power output: 100 mW | \n
Power density: 124.3 W/cm2\n | \n
Irradiation Time: 60 seconds | \n
Distance from the implant: 5 mm | \n
PDT1 treatment parameters.
After the incubation period, the implants were taken out of anaerobic conditions and they were randomly divided into four study groups and two control groups, each group containing twelve implants (n = 12).
\nThe implants were treated with a diode laser (Laser HF®, Hager Werken, Duisburg, Germany) and a toluidine blue-based dye (155 μg/ml, LaserHF® Paro-PDT solution). The laser parameters are presented in 1.
\nA combination of a diode laser (Helbo® Therapielaser, Helbo Photodynamic Systems GmbH & Co KG, Grieskirchen, Austria) and a phenothiazine chloride dye (Helbo® Blue photosensitizer) was used for the treatment of the implants belonging to this group. The laser parameters are presented in Table 2.
\nWavelength: 660 nm | \n
Fiber tip: 3D pocket probe | \n
Power output: 100 mW | \n
Power density: 35.37 W/cm2\n | \n
Irradiation Time: 60 seconds | \n
Distance from the implant: 5 mm | \n
PDT2 treatment parameters.
The implants belonging to this group, were treated with LED curing light (Optilight Ld®, Gnatus, Brazil). A red LED light, (Ledengin,Inc.®, San Jose, USA) was used in combination with a toluidine blue solution (Biognost®, Zagreb, Croatia). The laser parameters are presented in Table 3.
\nWavelength: 660 nm | \n
Fiber tip: 6 mm LED composite curing tip | \n
Power output: 200 mW | \n
Power density: 0.71 W/cm2\n | \n
Irradiation Time: 60 seconds | \n
Distance from the implant: 5 mm | \n
PDT3 treatment parameters.
The first step was coating the implants with the respective photosensitive dye according to the treatment group. After 60 seconds the implants were rinsed with sterile saline solution. For standardization of the treatment protocols for every treatment group, the implants, were placed in a rotating electric motor (Shenzhen Powerful Electronics, Shajing, China), with a rotating speed of 10 rpm.
\nThe treatment time was 60 seconds for every group from a distance of 5 mm from the implant. The treatment procedures for titanium and zirconia implants are shown in Figures 2 and 3.
\nA titanium implant treated from a distance of 5 mm for 60 seconds while rotating on the electric motor.
A zirconia dental implant placed in a rotational motor and treated with PDT2 for 60 seconds.
The implants belonging to this group were placed in photosensitive dye (toluidine blue; Biognost®, Zagreb, Croatia) solution (1 mg/ml) for 60 seconds. Then they were rinsed with sterile saline solution.
\nTwo control groups were included. The negative control group (NC) did not receive any treatment. After removing the implants from the bacterial suspension and keeping them in room conditions for 60 seconds, microbiologycal analysis followed.
\nThe implants belonging to the positive control group (PC) were put in 0.2% chlorhexidine gluconate solution (Curasept ADS® Curaden International AG, Kriens, Switzerland) for a duration of 60 seconds. After their removal from the chlorhexidine solution, the implants were only rinsed with sterile saline to remove the remaining solution.
\nAfter treatment procedures each implant was placed in a tube containing 500 μl of phosphate buffered saline (PBS). The tubes were vortexed for 60 seconds (Vortex, Genius 3, IKA, Germany). This was done to remove the remaining bacterial cells from the implant surfaces.
\nFrom each tube, 100 μl were taken and using a 96-well microtiter plates a ten-fold dilution was performed and 30 μl of suspension from each well was then inoculated to Brucella agar plates.
\nThe plates were placed in anaerobic conditions and after 72 hours and the colony forming units per milliliter (CFU/ml) were counted (Figure 4). MALDI Biotyper (Bruker Daltonics, Germany) was used to macroscopically differentiate distinctive colonies.
\nVisible colonies of bacteria on Brucella agar plate.
Scanning electron microscopy (SEM) was performed on one randomly selected implant from each of the treatment groups and one sterile non-treated implant. The implants for SEM analysis were stored for 2 hours in 2% paraformaldehyde and, later on dehydrated in increasing concentrations of ethanol (60%, 75% and 95%), for 30 minutes in each and dried overnight. The surfaces of the prepared implants were analyzed by SEM (Vega TS5136MM, Tescan, Brno, Czech Republic). The SEM images were taken at 1:250 magnifications under high vacuum (HiVac) with a high voltage (HV) of 30 kV. All the images were taken between the fourth and the fifth thread of the implants. As for the zirconia implants, they are non-conducting material and in order to make the samples conductive and avoid charging of the sample surface, the implants were coated with gold and palladium sputter (SC7620 Mini Sputter Coater, Quorum Technologies Ltd., UK).
\nThe differences between the groups for each bacterial species separately and for the total count of bacteria, were compared by analysis of variance test (ANOVA) and Tukey test, as a post hoc. The level of significance was set at 5%. The statistical package SAS system for Windows (Release 8.02, SAS Institute Inc., Cary, NC, USA) was used.
\nTo determine the difference among the groups and between the two types of implants, multivariate analysis of variance test was applied.
\nThe comparison between the two types of implants: titanium and zirconia, regardless of the study groups, showed that there was a significantly lower number of bacteria on zirconia implants for all three types of bacteria separately, as well as for the total number of bacteria (Table 4).
\nFor the comparison among the study groups regardless of the type of implant, Tukey test was applied. Regarding the total number of bacteria, the least bacteria were found in PDT1 and PDT2. These two groups were followed by PDT3 and PC without significant difference among them. The negative control group (NC) as expected, had the largest number of bacteria compared to the other groups. The same results were obtained for the number of each bacteria separately.
\nThe total number of bacteria for every group and both implant types are shown in Figure 5 in schematic form. The difference between zirconia implants and titanium implants was not the same for all groups. The smallest difference between both types of implants in the number of bacteria was for the control group. The impact was almost the same for PDT1, PDT2, PC and TB, while the largest difference between titanium and zirconia implants were in the PDT3 group. The results for each of the bacteria separately are shown in Figures 6–8.
\nThe total number of bacteria in logarithmic form, for both types of implants and for each study groups.
The number of A. actinymycetemcomitans in logarithmic form for both types of implants and the study groups.
The number of P. gingivalis in logarithmic form for both types of implants and the study groups.
The number of P. intermedia in logarithmic form for both types of implants and the study groups.
There were statistically significant differences among the groups for each of the bacteria separately and also for the total number of bacteria (p = 0.0022). These results are presented in Table 5 in logarithmic form. Regarding the total number of bacteria, the largest reduction was observed in the PDT1 (98.3%) and PDT2 (97.8%) groups. These two groups had statistically significant difference when compared to NC (p < 0.05). In the PDT3 group there was a 68.7% bacterial reduction, without statistically significant difference when compared to NC (Table 5).
\n\n\n | Aggregatibacter actynomycetemcomitans | \nPorphyromonas gingivalis | \n||||||||
---|---|---|---|---|---|---|---|---|---|---|
Factor | \nN | \nmean | \nst.d. | \n\n | p* | \nN | \nmean | \nst.d. | \n\n | p* | \n
Implants | \n\n | \n | \n | \n | \n | \n | \n | \n | \n | \n |
Zirconia | \n72 | \n1.9 | \n(2.1) | \n\n | <0.0001 | \n72 | \n1.6 | \n(2.1) | \n\n | <0.0001 | \n
Titanium | \n72 | \n4.9 | \n(2.5) | \n\n | 72 | \n4.9 | \n(2.6) | \n\n | ||
Group | \n\n | \n | \n | \n | \n | \n | \n | \n | \n | \n |
PDT1 | \n24 | \n1.9 | \n(2.2) | \nb | \n<0.0001 | \n24 | \n2.0 | \n(2.5) | \nb | \n<0.0001 | \n
PDT2 | \n24 | \n1.8 | \n(2.0) | \nb | \n24 | \n1.6 | \n(2.1) | \nb | \n||
PDT3 | \n24 | \n3.1 | \n(2.9) | \nab | \n24 | \n2.9 | \n(2.8) | \nab | \n||
TB | \n24 | \n4.3 | \n(2.7) | \na | \n24 | \n4.0 | \n(2.7) | \na | \n||
PC | \n24 | \n2.9 | \n(2.7) | \nab | \n24 | \n2.8 | \n(2.6) | \nab | \n||
NC | \n24 | \n6.2 | \n(1.3) | \n\n | 24 | \n6.2 | \n(1.6) | \n\n | ||
\n | \nPrevotella intermedia\n | \nTotal | \n||||||||
Factor | \nN | \nmean | \nst.d. | \n\n | p* | \nN | \nmean | \nst.d. | \n\n | p* | \n
Implants | \n\n | \n | \n | \n | \n | \n | \n | \n | \n | \n |
Zirconia | \n72 | \n2.0 | \n(2.1) | \n\n | <0.0001 | \n72 | \n2.3 | \n(2.3) | \n\n | <0.0001 | \n
Titanium | \n72 | \n5.3 | \n(2.7) | \n\n | 72 | \n5.8 | \n(2.5) | \n\n | ||
Group | \n\n | \n | \n | \n | \n | \n | \n | \n | \n | \n |
PDT1 | \n24 | \n2.6 | \n(2.5) | \nb | \n<0.0001 | \n24 | \n2.8 | \n(2.6) | \nb | \n<0.0001 | \n
PDT2 | \n24 | \n2.0 | \n(2.3) | \nb | \n24 | \n2.3 | \n(2.4) | \nb | \n||
PDT3 | \n24 | \n3.1 | \n(3.2) | \nab | \n24 | \n3.6 | \n(3.2) | \nab | \n||
TB | \n24 | \n4.5 | \n(2.9) | \na | \n24 | \n5.0 | \n(2.7) | \na | \n||
PC | \n24 | \n3.2 | \n(2.7) | \nab | \n24 | \n3.6 | \n(2.7) | \nab | \n||
NC | \n24 | \n6.4 | \n(1.8) | \n\n | 24 | \n7.1 | \n(1.5) | \n\n |
Number of bacteria by implant type and treatment groups in logarithmic form.
p-value for MANOVA test.
abc - result of post-hoc comparison (Tukey test). Having the same letter means that there is no statistically significant difference.
\n | Aggregatibacter actynomycetemcomitans | \nPorphyromonas gingivalis | \nWilks’ lambda | \n||||||||
---|---|---|---|---|---|---|---|---|---|---|---|
Group | \nN | \nmean | \nst.d. | \n\n | p* | \nN | \nmean | \nst.d. | \n\n | p* | \np | \n
PDT1 | \n12 | \n3.3 | \n2.2 | \nb | \n0.0006 | \n12 | \n3.7 | \n2.5 | \nbc | \n0.0003 | \n0.0026 | \n
PDT2 | \n12 | \n3.1 | \n2.0 | \nb | \n\n | 12 | \n2.8 | \n2.4 | \nc | \n\n | \n |
PDT3 | \n12 | \n5.4 | \n2.3 | \nab | \n\n | 12 | \n5.2 | \n2.2 | \nabc | \n\n | \n |
TB | \n12 | \n6.2 | \n2.3 | \na | \n\n | 12 | \n6.2 | \n2.0 | \nab | \n\n | \n |
PC | \n12 | \n4.7 | \n2.7 | \nab | \n\n | 12 | \n4.7 | \n2.3 | \nabc | \n\n | \n |
NC | \n12 | \n6.5 | \n1.7 | \na | \n\n | 12 | \n6.8 | \n1.9 | \na | \n\n | \n |
\n | \nPrevotella intermedia\n | \nTotal number of bacteria | \nWilks’ lambda | \n||||||||
Group | \nN | \nmean | \nst.d. | \n\n | p* | \nN | \nmean | \nst.d. | \n\n | p* | \np | \n
PDT1 | \n12 | \n4.3 | \n2.4 | \nab | \n0.0096 | \n12 | \n4.7 | \n2.3 | \nbc | \n0.0022 | \n0.0026 | \n
PDT2 | \n12 | \n3.6 | \n2.4 | \nb | \n\n | 12 | \n3.9 | \n2.3 | \nc | \n\n | |
PDT3 | \n12 | \n5.4 | \n3.1 | \nab | \n\n | 12 | \n6.1 | \n2.5 | \nabc | \n\n | |
TB | \n12 | \n6.7 | \n2.4 | \na | \n\n | 12 | \n7.0 | \n2.2 | \nab | \n\n | \n |
PC | \n12 | \n4.9 | \n2.7 | \nab | \n\n | 12 | \n5.4 | \n2.6 | \nabc | \n\n | \n |
NC | \n12 | \n7.0 | \n2.2 | \na | \n\n | 12 | \n7.4 | \n1.8 | \na | \n\n | \n |
Results of ANOVA and Tukey’s post hoc test for the titanium implants.
p-value for ANOVA test.
abc - result of post-hoc comparison (Tukey test). Having the same letter means that there is no statistically significant difference.
When each bacteria was compared separately, the PDT1 and PDT2 groups also showed the largest bacterial reduction. PDT1 group, was significantly more effective in the eliminating A. actinomycetemcomitans and P. gingivalis (p < 0.05). As for P.intermedia the PDT1 group showed no significant difference compared to NC group. On the other hand the PDT2 group was significantly more effective in the elimination of each of the bacteria separately when compared to the NC group (p < 0.05).
\nThe least effective among the groups, when compared to the NC group, was the TB group (62.4%). Compared to NC there was no significant difference neither for the total number nor for each bacteria separately.
\nStatistically significant difference was observed among the groups. This was the case for both the total number of bacteria and each bacteria separately (p < 0.0001). Every group showed vast bacterial reduction with statistically significant difference when compared to the negative control (NC). These results are shown in Table 6 in logarithmic form.
\n\n | Aggregatibacter actynomycetemcomitans | \nPorphyromonas gingivalis | \nWilks’ lambda | \n||||||||
---|---|---|---|---|---|---|---|---|---|---|---|
Group | \nN | \nmean | \nst.d. | \n\n | p* | \nN | \nmean | \nst.d. | \n\n | p* | \np | \n
PDT1 | \n12 | \n0.4 | \n0.8 | \na | \n0.0001 | \n12 | \n0.4 | \n0.8 | \nb | \n0.0001 | \n0.0001 | \n
PDT2 | \n12 | \n0.4 | \n0.6 | \na | \n\n | 12 | \n0.3 | \n0.5 | \nb | \n\n | \n |
PDT3 | \n12 | \n0.8 | \n1.1 | \na | \n\n | 12 | \n0.6 | \n0.7 | \nb | \n\n | \n |
TB | \n12 | \n2.4 | \n1.3 | \n\n | \n | 12 | \n1.9 | \n1.1 | \na | \n\n | \n |
PC | \n12 | \n1.2 | \n1.0 | \na | \n\n | 12 | \n0.9 | \n1.2 | \nab | \n\n | \n |
NC | \n12 | \n5.9 | \n0.7 | \n\n | \n | 12 | \n5.7 | \n1.0 | \n\n | \n | \n |
\n | \nPrevotella intermedia\n | \nTotal number of bacteria | \nWilks’ lambda | \n||||||||
Group | \nN | \nmean | \nst.d. | \n\n | p* | \nN | \nmean | \nst.d. | \n\n | p* | \np | \n
PDT1 | \n12 | \n0.8 | \n0.9 | \nb | \n0.0001 | \n12 | \n0.9 | \n1.0 | \nb | \n0.0001 | \n0.0001 | \n
PDT2 | \n12 | \n0.5 | \n0.7 | \nb | \n\n | 12 | \n0.7 | \n0.8 | \nb | \n\n | |
PDT3 | \n12 | \n0.8 | \n0.9 | \nb | \n\n | 12 | \n1.1 | \n1.2 | \nb | \n\n | |
TB | \n12 | \n2.3 | \n1.2 | \na | \n\n | 12 | \n2.9 | \n1.2 | \na | \n\n | \n |
PC | \n12 | \n1.5 | \n1.5 | \nab | \n\n | 12 | \n1.8 | \n1.5 | \nab | \n\n | \n |
NC | \n12 | \n5.9 | \n1.3 | \n\n | \n | 12 | \n6.7 | \n0.9 | \n\n | \n | \n |
Results of ANOVA and Tukey’s post hoc test for the zirconia implants.
p-value for ANOVA test.
abc - result of post-hoc comparison (Tukey test). Having the same letter means that there is no statistically significant difference.
The PDT1, PDT2 and PDT3 had the largest bacterial reduction for each bacterium separately, as well as for the total count of bacteria. There was a reduction of more than 99% in comparison to NC. However, between these three groups the differences in bacterial reduction were not statistically significant difference neither for each of the bacteria separately nor for the total number of bacteria (p > 0.05).
\nThe lowest bacterial reduction for each bacteria separately and also for the total number of bacteria was observed in the TB group. The PC group had lower bacterial reduction compared to PDT1, PDT2 and PDT3 without statistically significant differences among them. It also did not differ significantly compared to the TB in terms of the total bacterial count, P. gingivalis and P. intermedia. It had a significant difference compared to the TB only for A. actinomycetemcomitans.
\nThe SEM images from the PDT1, PDT2, and PDT3 groups did not show any surface alterations, cracks, or damage when compared to the images obtained for the sterile implants. Visually, their surface appeared to be very similar to the surface of the sterile implant, for both titanium and zirconia implants (Figures 9–12).
\nSterile titanium implant; magnification 1:250 (left). Titanium implant treated with PDT1; magnification 1:250 (right).
Titanium implant treated with PDT2; magnification 1:250 (left). Titanium implant treated with PDT3; magnification 1:250 (right).
Sterile zirconia implant; magnification 1:250 (left). Zirconia implant treated with PDT1; magnification 1:250 (right).
Zirconia implant treated with PDT2; magnification 1:250 (left). Zirconia implant treated with PDT3; magnification 1:250 (right).
The clinical efficacy of PDT against peri-implantitis has been demonstrated in several clinical studies. In a randomized controlled trial study by Wang et al. [60] it was shown that PDT combined with mechanical debridement significantly improved pocket depth, clinical attachment loss, plaque index and sulcus bleeding index compared with baseline and the control groups in participants with peri-implantitis. Similar results were obtained in a 3 months randomized clinical trial done by Rakasevic et al. [61].
\nSince the main goal when treating peri-implantitis is to eliminate the bacteria from the soft tissues and the implant surface, in order to create conditions for grafting and re-osseointegration, the use of photodynamic therapy is mostly used as an adjunct therapy during the treatment of peri-implanitits with the purpose of eliminating bacteria from the rough surfaces of the implants. The treatment of peri-implantitis can be non-surgical and surgical. During the non-surgical treatment of peri-implantitis the photosensitive dye is applied on the pocket around the infected implant and the light source is applied. This procedure is shown in Figure 13.
\n(Left) application of the dye. (Right) application of the light source.
However, photodynamic therapy is mostly used in conjunction with surgical treatment of peri-implantitis as an adjunct therapy after implantoplasty, mechanical debridement and chemical decontamination of the implant surface. The surgical approach is presented in Figures 14–17.
\nSurgical treatment of peri-implantits. Visible bone resorption around the implants.
Implantoplasty procedure.
Application of the photosensitive dye.
Application of the light source.
One of the main reasons of implant failure is peri-implantitis. The prevalence rates of peri-implantitis differs among different studies and this is due to the different reporting methods and characteristics [62, 63, 64]. Van Velzen et al. [65] in their 10 years prospective cohort study reported a prevalence of 7%. Meijer et al. [66] reported that after 10 years 29.7% of patients were affected by peri-implantitis. Fardal et al. [67] report a rate of 53.5% at the patient level and 31.1% at the implant level, which is much higher than the data from other studies.
\nThe treatment of peri-implantitis is complex and it often includes combination of conventional therapy with the addition of antimicrobials. However, use of antimicrobials does not have a long term effects and it can lead to antimicrobial resistance and development of superinfections [68]. Therefore, alternative antimicrobial approaches for achieving implant disinfection have been sought.
\nPhotodynamic therapy is a promising alternative when treating periodontal diseases and peri- implant diseases. Up to date there have been many in vitro [69, 70, 71] and clinical studies [60, 61] evaluating the effect of photodynamic therapy in treating peri-implantitis.
\nRegarding the in vitro evaluation, the present study aimed to evaluate the efficacy of photodynamic therapy on dental implants contaminated under in vitro conditions. The implants were contaminated in order to try to recreate the adhesion stage of biofilm formation on the implant surface. Many in vitro studies have used similar methodology to achieve titanium implant contamination [62, 63, 71].
\nThe main focus of our study was to evaluate if photodynamic therapy is efficient in eradicating the bacteria from the implant surface when compared to the negative control group (NC) and to the conventional treatment with chlorhexidine solution (PC). Furthermore, the focus was to evaluate different types of devices and with different parameters and photosensitizers and the reaction of different bacteria to aPDT.
\nThe results from our study showed that PDT1 and PDT2 groups were more eliminated 98.3% and 97.8% of the total number of bacteria when compared to NC group. These groups were the most effective among the study groups. Both PDT1 and PDT2 groups were a combination of a diode laser with a wavelength of 660 nm and a photosensitizer.
\nThe results of this study are similar to other in vitro, in vivo and clinical studies [64, 71, 72]. Marotti et al. [71] in their study demonstrated that aPDT is effective against the bacteria present in peri-implantitis. The irradiation time did not influence the results. Similar results were obtained from both the groups (3 minute and 5 minutes irradiation time) and there was no significant difference between them. The effect of aPDT did not differ significantly from the disinfection with 0.12% chlorhexidine solution. Our results were similar to this study and even though we used a higher concentration of chlorhexidine (0.2%), both PDT1 and PDT2 had no significant difference when compared to PC.
\nIn a study done by Haas et al. [64] it was demonstrated that 60 seconds of light exposure in combination with photosensitizer can effectively eradicate A. actinomycetemcomitans, P. gingivalis and P. intermedia. One of the goals of the present study was to evaluate aPDT against each bacteria separately. The results obtained for A. actinomycetemcomitans and P. gingivalis were similar to the results obtained for the total bacterial count. Both PDT1 and PDT2 had significant difference when compared to the NC. However, the results of P. intermedia showed that PDT2 was more effective against this bacteria when compared to the other groups.
\nThe least effective treatment group was PDT3 without statistically significant difference compared to NC or PC groups regarding the total bacterial count. It must be noted that for the PDT3 group we used a modified dental LED light and not a diode laser. This was done to evaluate and compare the LED light against diode lasers as a light source.
\nThe efficacy of LED lights as a light source in photodynamic therapy has been tested in many studies however, only a few studies have tested its efficacy on titanium implant surfaces. The results from these studies are conflicting since the study design and light source parameters differ greatly. In a study conducted by Cho et al. [73] the efficacy of a green LED light was tested. The LED light was combined with erythrosine dye and was evaluated against A. actinomycetemcomitans. Their results showed that this combination is effective and reduces the bacteria attached titanium surfaces up to 92.4%. The irradiation time in this study was 60 seconds and the treatment was done on only one surface of titanium discs. This provides uniform distribution of the light source. In contrast, in the present study we applied the light source in a rotating motion in order to emulate clinical application of aPDT around a contaminated implant. This might be the reason why our results showed differ with the aforementioned study [73].
\nIn contrast to the in vitro study by Cho et al. [73], in a clinical study done by De Angelis et al. [74] the use of LED light showed no significant difference after 4 months of follow up when compared to mechanical debridement and scaling.
\nIn our study we evaluated the efficacy of aPDT on two types of implants: titanium and zirconia dental implants. The efficacy of aPDT on zirconia implant surfaces has not been evaluated in other studies up to date.
\nThe results obtained from our study showed that each test group was very effective in eliminating the bacteria from the zirconia surface and all had significantly lower bacterial count when compared to NC. However, in between the groups there was no significant difference. The higher efficacy of aPDT against zirconia surfaces when compared to titanium surfaces might be due to the surface properties of zirconia which might lead to a lower affinity of the bacteria to be attached to zirconia surfaces. Zirconia surfaces are smoother, have a lower surface roughness and lower surface free energy [75, 76]. In a study done by Scarano et al. [75] titanium and zirconia oxide discs were placed in the mouths of patients in order to evaluate in which surface the bacteria adhere less. After 24 h it was shown that there were significantly less bacteria on the zirconium oxide surfaces. Al-Radha et al. [76] showed similar results. In their study titanium blasted with zirconia and the zirconia material showed better results when compared to the titanium surface regarding the adhesion of bacteria after coating the surfaces with saliva pellicle.
\nPDT1, PDT2 and PDT3 in addition to the significant difference compared to NC, they also had significant difference from the TB group. The results of the PDT3 group for the zirconia dental implants were comparable to PDT1 and PDT2. This can suggest that LED light with additional improvements in light distribution and parameters can have an antimicrobial effect. As mentioned before there are conflicting results regarding the antimicrobial effect of using LED. Several studies reported beneficial results following use of LED lights, as a light source [77, 78]. On the other hand, several studies reported insignificant improvement in the treatment outcomes using LED light for PDT [74]. However, it is difficult to compare the results of present study with the previous ones, mainly due to the differences in the study protocols and lack of studies conducted on zirconia implant surfaces.
\nThe use of diode lasers in many studies has been shown to be safe in regard to the implant surface, compared to Nd:YAG, Er:YAG, CO2 and Ho:YAG lasers, which can damage the implant surfaces [57]. Castro et al. [79] concluded that 980 nm diode laser irradiation does not damage titanium implant surfaces and seems to be safe irrespective of power output used.
\nIn the present research, no structural changes on the implant surfaces following therapy was observed. PDT1, PDT2 and PDT3, did not cause visible damage on titanium or zirconia implant surface at a magnification of 1:250.
\nRegarding the clinical use of PDT, several studies reported conflicting results. Many studies demonstrated improvement in clinical outcomes of patients with peri-implantitis when aPDT was combined with mechanical debridement [60, 80, 81]. Romeo et al. [82] suggested that PDT is a useful adjunct therapy but it could not replace the mechanical and surgical treatment of peri-implantitis. Similarly other studies suggest that the PDT improves the outcomes of peri-implantitis [60, 83, 84].
\nOn the other hand, there are several studies that report no added benefit from using PDT when compared to conventional treatment modalities for peri-implantitis [85, 86].
\nThe results of this in vitro study should be considered preliminary, since it cannot be generalized to in vivo and clinical conditions. The biggest concern related to future in vivo and clinical applications is stability of achieved in vitro results (short term beneficial effects in reducing the number of periopathogens). Also, the presence of plaque formation on implants, degree of salivation and host-immune response is very important.
\nIt is of utmost importance that further clinical trials be conducted in order to clarify the potential efficacy of PDT as an adjunct therapy to peri-implantitis and clear and effective treatment protocols should be established in order to benefit the most from the properties of PDT.
\nThe authors declare no conflicts of interest.
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