Entertainment & Media

Bovine Serum Albumin-Loaded Chitosan/Dextran Nanoparticles: Preparation and Evaluation of Ex Vivo Colloidal Stability in Serum

Description
Bovine Serum Albumin-Loaded Chitosan/Dextran Nanoparticles: Preparation and Evaluation of Ex Vivo Colloidal Stability in Serum
Published
of 9
All materials on our website are shared by users. If you have any questions about copyright issues, please report us to resolve them. We are always happy to assist you.
Related Documents
Share
Transcript
  Hindawi Publishing CorporationJournal o NanomaterialsVolume 󰀲󰀰󰀱󰀳, Article ID 󰀵󰀳󰀶󰀲󰀹󰀱, 󰀹 pageshttp://dx.doi.org/󰀱󰀰.󰀱󰀱󰀵󰀵/󰀲󰀰󰀱󰀳/󰀵󰀳󰀶󰀲󰀹󰀱 Research Article Bovine Serum Albumin-Loaded Chitosan/DextranNanoparticles: Preparation and Evaluation of   Ex Vivo  ColloidalStability in Serum Haliza Katas, Zahid Hussain, and Siti Asarida Awang  Centre for Drug Delivery Research, Faculty of Pharmacy, Universiti Kebangsaan Malaysia, Kuala Lumpur Campus, Jalan Raja Muda Abdul Aziz, 󰀵󰀰󰀳󰀰󰀰 Kuala Lumpur, Malaysia Correspondence should be addressed to Haliza Katas; haliz󰀱󰀲@hotmail.comReceived 󰀱 January 󰀲󰀰󰀱󰀳; Revised 󰀲󰀲 March 󰀲󰀰󰀱󰀳; Accepted 󰀷 April 󰀲󰀰󰀱󰀳Academic Editor: Xing-Jie LiangCopyright © 󰀲󰀰󰀱󰀳 Haliza Katas et al. Tis is an open access article distributed under the Creative Commons Attribution License,which permits unrestricted use, distribution, and reproduction in any medium, provided the srcinal work is properly cited.Chitosan (CS) nanoparticles have several distinct intrinsic advantages; however, their  in vivo  colloidal stability in biological 󿬂uidswas not ully explored especially when carrying proteins. Te present study aimed to investigate their colloidal stability using an ex vivo  physiological model o etal bovine serum (FBS) and human serum (HS). Te stability o bovine-serum-albumin (BSA-)loaded nanoparticles was relatively higher in FBS than that in HS. Particle size o unloaded and BSA-loaded nanoparticles wasstatistically unchanged up to 󰀲󰀴h afer incubation in FBS. However in HS, a signi󿬁cant increase in particle size rom 󰀱󰀴󰀴  ±  󰀱󰀷 to󰀷󰀱󰀱 ± 󰀲󰀲nm was observed or unloaded nanoparticles and by 󰀲.󰀵-old or BSA-loaded nanoparticle, at 󰀲󰀴h afer incubation in HS.Zeta potential o both nanoparticles was less affected by the components in FBS compared to those in HS. A remarkable swellingextent was experienced or unloaded and BSA-loaded nanoparticles in HS, up to 󰀵󰀴  ±  󰀴% and 󰀴󰀴  ±  󰀵%, respectively. Morphology o unloaded and BSA-loaded nanoparticles was varied rom smooth spherical and rod shape to irregular shape when incubated inFBS;however,ormagglomerateswhenincubatedinHS.Tese󿬁ndingsthereoresuggestthatHSismorereactivetocausecolloidalinstability to the chitosan nanoparticles compared to FBS. 1. Introduction Despite several intrinsic and distinct advantages o nanopar-ticles, the concerns about the health risks o polymericnanoparticles have been escalating nowadays due to thehigher incidence o instability o nanoormulations. Numer-ous  in vitro  studies reported that the nanosized particlesare biologically more potent than the equivalent micron-sized particles o same chemical composition [󰀱, 󰀲]. Te potency and biological activity o polymeric nanoparticleshave been connected with several physicochemical (col-loidal) characteristics such as the shape o the particles,theirsuracearea,hydrodynamicparticlesize,agglomerationor 󿬂occulation rate, surace potential (zeta potential), andthe surace chemistry o nanoparticles [󰀳]. Tese physico- chemical characteristics o polymeric nanoparticles are inturn highly affected by the medium in contact such asbiological 󿬂uids (plasma, serum, saliva, sweat, sebum, tears,etc.). Tus, it was argued that the physicochemical propertiesand unique kinetics o nanomaterials in biological solutionsshould be considered prior to various pharmaceutical andpharmacodynamic testings [󰀳–󰀵]. Besides, the adsorption anddesorptionaffinitiesoreactivecomponentsobiological󿬂uids (such as proteins) onto the nanoparticles surace, type,amount, and conormation o the adsorbed proteins arenecessary to be considered when analyzing the biologicalresponses o the nanoparticles [󰀶, 󰀷]. Among the biological 󿬂uids, etal bovine serum (FBS), acommonly used supplement or cell culture, and the humanserum (HS), the main component o human blood [󰀸], were used in the current research to assess the colloidal stability o nanoparticles. Serum is a complex mixture o differentactors and contains a large number o components likegrowthactors,proteins,vitamins,hormones,traceelements,and other essential and nonessential components [󰀹]. Serum  󰀲 Journal o Nanomaterialscomponents would tend to alter the colloidal characteris-tics (particle size, PDI, and zeta potential) o polymericnanoparticles and carry their own surace charges (e.g.proteins with negative charges while some growth hormonesexhibit positive charges) which may in󿬂uence the overallphysicochemical characteristics o polymeric nanoparticles[󰀱󰀰,󰀱󰀱].Tereore,determinationoproteinstabilityinserum constitutes a powerul and important screening assay.  In vivo testing o peptide stability is obviously o more relevancethan  in vitro  but blood samples should be as steriled aspossible to ensure maximum activity o proteolytic enzymesandminimalintererencewiththeassay [󰀱󰀲].Besidesthatthe blood is a biohazardous material, it must also be heparinisedand this may interere with the assay.Recently, or preparing the polymeric nanoparticles, chi-tosan (CS) has been extensively ocused on. CS is a naturalbiopolymer derived rom chitin deacetylation [󰀱󰀳, 󰀱󰀴]. It has attained a remarkable attention due to its biologicalproperties such as excellent biocompatibility, hydrophilicity,biodegradability, and antibacterial activity  [󰀱󰀵]. Gan and Wang [󰀱󰀶] had recommended the CS as the superior partic- ulate polymer or  in vivo  administration due to its nontoxicnature and degradation by the action o lysozyme in thebody. Besides, CS is a good candidate or  in vivo  and  ex vivo applications because it would not accumulate in body tissues[󰀱󰀷].aken together, the selection o cross-linking agent usedin the particle preparation is also an imperative actor to ur-therimprovethecolloidalstabilityoCSnanoparticles.Tus,inthepresentstudy,CSnanoparticleswerepreparedbyionic-cross linking o CS with dextran sulphate (DS). Te criterionor using DS as the cross-linking agent is that it had beenreported to produce mechanically more stable nanoparticlescompared to the penta-sodium tripolyphosphate (PP) [󰀱󰀸,󰀱󰀹]. Moreover, CS/DS nanoparticles offer many therapeuticadvantagesoverCS/PPnanoparticles.ParticlesizeoCS/DSnanoparticles was comparatively small which directly affectthe colloidal stability o particulate dispersion [󰀲󰀰]. Besides the particle size, zeta potential is also the vital indicatorto predict and control the stability o colloidal dispersion[󰀲󰀱]. Hence, the present study was aimed to investigate the colloidal stability o CS/DS nanoparticles in HS and FBS.Te colloidal stability was assessed in terms o particle size,zetapotential,PDI,swellingcharacteristics,andaswellasthemorphology o CS/DS nanoparticles. 2. Materials and Methods 󰀲.󰀱. Materials.  Low molecular weight chitosan (deacety-lation degree, 󰀷󰀵–󰀸󰀵%, M.wt, 󰀷󰀰kDa), glacial acetic acid(CH 3 COOH), dextran sulphate (DS), bovine serum albu-min (BSA) (M.wt, 󰀴󰀶kDa), Bradord reagent, etal bovineserum, and human serum (obtained rom human male ABplasma) were purchased romSigma-Aldrich, USA. All otherchemicals were o analytical grades and used without urtherpuri󿬁cation. 󰀲.󰀲. Preparation of Unloaded CS/DS Nanoparticles.  CS/DSnanoparticles were prepared  via  ionic-gelation method, pre- viously developed by Calvo et al. [󰀲󰀶] with some modi󿬁ca- tion.CSwasdissolvedin󰀱%v/vglacialaceticacidtoproduce󰀰.󰀰󰀷󰀵% w/v CS solution. Tree different concentrations o DS (󰀰.󰀰󰀷󰀵, 󰀰.󰀱, and 󰀰.󰀱󰀲󰀵% w/v) were prepared by dissolvingDS in distilled water. CS/DS nanoparticles were preparedsimultaneously by adding 󰀴󰀰mL o DS solution dropwise in󰀱󰀰󰀰mLo󰀰.󰀰󰀷󰀵%w/vCSsolutionunderaconstantmagneticstirring at 󰀷󰀰󰀰rpm or 󰀳󰀰min. Tereafer, CS/DS nanoparti-cles were harvested byultracentriugation(󰀲󰀵󰀰󰀰󰀰rpm)usingan Optima L-󰀱󰀰󰀰 XP Ultracentriuge (Beckman-Coulter,USA) with a rotor NV 󰀷󰀰.󰀱 i (Beckman-Coulter, USA) at󰀱󰀰 ∘ C or 󰀱󰀵min. 󰀲.󰀳. Preparation of BSA-Loaded CS/DS Nanoparticles.  Forpreparing BSA-loaded CS/DS nanoparticles, BSA was dis-solvedinPBSsolution(pH,󰀷.󰀴)toproducetheconcentrationo󰀱mg/mL.TepHoCSsolutionwasthenadjustedto󰀵.󰀵by adding either 󰀰.󰀵M NaOH or 󰀰.󰀵M HCl. BSA solution wasthen premixed with 󰀰.󰀰󰀷󰀵% w/v CS solution and incubatedor 󰀳󰀰min at room temperature. DS solution (󰀰.󰀰󰀷󰀵, 󰀰.󰀱and 󰀰.󰀱󰀲󰀵% w/v) was then added dropwise in the reactionmixture under a continuous magnetic stirring (󰀷󰀰󰀰rpm) or󰀳󰀰min to produce BSA-loaded CS/DS nanoparticles. Teresultantnanoparticleswereharvestedbyultracentriugationat 󰀲󰀵󰀰󰀰󰀰rpm at 󰀱󰀰 ∘ C or 󰀱󰀵min. 󰀲.󰀴. Determination of Particle Size, PDI, and Zeta Potential. Mean particle size, PDI, and zeta potential o unloaded andBSA-loaded CS/DS nanoparticles were measured by usinga ZS-󰀹󰀰 Zetasizer (Malvern Instruments, UK). For particlesize analysis, measurements were perormed at 󰀲󰀵 ∘ C with adetection angle o 󰀹󰀰 ∘ . All measurements were perormedin triplicate and results were reported as mean  ±  standarddeviation (S.D). 󰀲.󰀵. Entrapment Efficiency (EE).  o determine EE, theBSA-loaded CS/DS nanoparticles were harvested by ultra-centriugation at 󰀲󰀵󰀰󰀰󰀰rpm using Optima L-󰀱󰀰󰀰 XP Ultra-centriuge (Beckman-Coulter, USA) with a rotor NV 󰀷󰀰.󰀱 i(Beckman-Coulter, USA) at 󰀱󰀰 ∘ C or 󰀱󰀵min. Supernatantsrecovered by ultra-centriugation were then decanted, andthe BSA contents were analyzed using Bradord proteinassay as per manuacturer’s instructions. Te samples werethen subjected to U.V/Vis spectrophotometer (U.V-󰀱󰀶󰀰󰀱, Shi-madzu, Japan) and analyzed at 󰀵󰀹󰀵nm (  max ), and U.Vabsorbance was recorded. Te percent EE o BSA was thencalculated indirectly rom remaining supernatant using theollowing equation [󰀲󰀲]:EE  = ( otal initial amount o BSA added −  Free amount o BSA in supernatant 􀀩× ( otal initial amount o BSA added ) −1 × 100 (󰀱)  Journal o Nanomaterials 󰀳 󰀲.󰀶. Stability Studies of CS/DS Nanoparticles in Serum.  oassess the colloidal stability o unloaded and BSA-loadedCS/DS nanoparticles, FBS was 󿬁rst incubated at 󰀳󰀷 ∘ C in awater bath incubator to simulate physiological media. FBSwas then added in the CS/DS nanoparticles dispersion withthe volume ratio o 󰀱:󰀱 in which the 󿬁nal concentration o 󰀵󰀰% v/v o serum was yielded. Subsequently, the resultingmixture was incubated in the water bath incubator at 󰀳󰀷 ∘ Cor󰀲󰀴h.Temeanparticlesize,PDI,andsuracechargeweremeasured at predetermined time points (󰀰, 󰀱󰀰, 󰀳󰀰min, 󰀱, 󰀲, 󰀳,󰀱󰀲, and 󰀲󰀴h). Same experimental protocol was also repeatedto determine the colloidal stability o CS/DS nanoparticles inHS. CS/DS nanoparticles were suspended in PBS (pH, 󰀷.󰀴)prior to mixing with FBS or HS. 󰀲.󰀷. Swelling Analysis.  o investigate the swelling character-istics o CS/DS nanoparticles, 󰀱󰀰󰀰mg o lyophilized powdersample o either unloaded or BSA-loaded CS/DS nanopar-ticles was immersed in 󰀱󰀰󰀰mL o FBS and/or HS at variousDS concentrations (󰀰.󰀰󰀷󰀵, 󰀰.󰀱, and 󰀰.󰀱󰀲󰀵) or 󰀲󰀴h at roomtemperature until a swollen equilibrium was achieved. Teswollensampleswerethencollectedby󿬁ltration,blottedwith󿬁lter paper or the removal o the surace adsorbed water,and weighed immediately. Ten, the swelling ratios o CS/DSnanoparticles were calculated using the ollowing equation[󰀲󰀳]:Swelling ratio  ( % ) = 󰀨󽠵   − 󽠵 󽠵 󽠵 󽠵 󰀩 × 100,  (󰀲)where  󽠵   and  󽠵 󽠵  are the average weights o swollen and dry samples, respectively. Results were reported as mean  ±  S.D. 󰀲.󰀸. Morphological Examination.  With a view to evaluatethe effect o FBS and HS on the morphological charac-teristics o CS/DS nanoparticles, the unloaded and BSA-loadednanoparticleswereincubatedinbothseraor󰀲󰀴handwereviewedundertransmissionelectronmicroscope(EM).Prior to EM analysis, the incubated dispersions o CS/DSnanoparticles (in FBS or HS at 󰀲󰀴h afer incubation) werediluted to 󰀱:󰀱󰀰 ratio with phosphate buffered saline (PBS,󰀰.󰀰󰀱M). Te resulting dilution was carried out in order toobtain the clear images (microscopic micrographs) o CS/DSnanoparticles under EM. o perorm the microscopic(EM) analysis, a drop o diluted nanoparticles dispersionin FBS or HS was placed onto the copper microgrid thatwas natively stained with phosphotungstic acid and allowedto evaporate and dry at room temperature ( 25 ± 2 ∘ C).Te dried microgrids were then viewed at different EMresolutions to assess the morphology o unloaded and BSA-loaded nanoparticles beore and afer incubation in FBS andHS. 󰀲.󰀹. Statistical Analysis.  Te data was presented as mean  ± standard deviation (S.D). Data was analyzed using SPSS 󰀱󰀷.󰀰(paired  t  -test and independent  t  -test and ANOVA, ollowedby ukey’s post hoc analysis). For paired  t  -test,  􍠵 < 0.05 showed the signi󿬁cant difference between the mean o testedgroups. For independent  t  -test,  􍠵 < 0.05  showed signi󿬁cantdifference between the mean o two independent testedsamples. 3. Results and Discussion 󰀳.󰀱. Colloidal Characteristics of CS/DS Nanoparticles󰀳.󰀱.󰀱. Particle Size, PDI, and Zeta Potential.  Figures 󰀱(a) and󰀱(b) present the results o particle size and zeta potentialo unloaded and BSA-loaded CS/DS nanoparticles preparedrom different concentrations o DS (󰀰.󰀰󰀷󰀵, 󰀰.󰀱, 󰀰.󰀱󰀲󰀵%w/v). Data clearly demonstrates that the particle size andzeta potential o CS/DS nanoparticles were not signi󿬁cantly affected by the DS concentrations ( 􍠵 > 0.05 , ANOVA,ukey’s post-hoc analysis), regardless o unloaded or BSA-loaded nanoparticles. Te results obtained were differentrom the previously published results [󰀲󰀴]. In present study,DS concentrations had less impact on the particle size, PDI,and zeta potential o CS/DS nanoparticles. Tis was expectedbecause CS/DS weight ratio o different ormulations wassmallandthereoreyieldednanoparticleswithalmostsimilarcolloidal characteristics. Te CS/DS weight ratios studied inthe present study were 󰀱:󰀱, 󰀱:󰀱.󰀳, and 󰀱:󰀱.󰀷 or 󰀰.󰀰󰀷󰀵, 󰀰.󰀱,and 󰀰.󰀱󰀲󰀵% w/v DS, respectively, while, the weight ratios o 󰀵:󰀳, 󰀵:󰀵, 󰀵:󰀱󰀰, and 󰀵:󰀲󰀰 were applied in the previous report.Despite that, all the ormulations were within the nanosizedrange (󰀲󰀰󰀰 ∼ 󰀳󰀰󰀰nm). On the other hand, the lower suracecharges o BSA-loaded CS/DS nanoparticles as compared tothe unloaded nanoparticles were expected to be due to theneutralization o positive charges ( − NH 3+ ) on the contouro CS by the negatively charged BSA molecules [󰀲󰀲, 󰀲󰀴, 󰀲󰀵], and this resulted in a decrease o the overall surace charge o nanoparticles as shown in Figure 󰀱(b). 󰀳.󰀱.󰀲. Entrapment Efficiency (EE).  Data revealed that the EEo BSA was observed to be affected by the DS concentrations.A higher EE was obtained when the CS/DS weight ratiowas reduced. Te lowest CS/DS weight ratio (󰀱:󰀱) producednanoparticles with the highest EE ( 95 ± 2 %) o BSA. EEor the other DS concentrations wase ound to be  91 ±2 % (at 󰀱:󰀱.󰀳) and  86 ± 2 % (at 󰀱:󰀱.󰀷). Tis was expectedas increase in the concentration o DS that would tendto increase the negative charge density on the surace o nanoparticleswhichmaysubsequentlyincreasetherepulsionenergy against the negatively charged BSA molecules whichmight not be avourable or the protein entrapment process.Furthermore, EE o BSA was also affected by pH o CSsolution. Te optimal pH was ound to be at 󰀵.󰀵. Te highestEE achieved in this study without adjusting the pH o CSsolution to 󰀵.󰀵 was only   46 ± 4 % (CS/DS weight ratio 󰀱:󰀱).Tis 󿬁nding was in agreement with Calvo et al. [󰀲󰀶] whichreportedthatthegreatestloadingefficiencycouldbeobtainedwhen protein was dissolved at a pH above its isoelectricpoint(pH󰀴.󰀸).AtthispH,BSAwouldpredominantlyexhibitits highest negative charge and could ionically interact withpositively charged  − NH 3+ groups on the backbone o CS.BSA also tends to be more negative as the pH o media  󰀴 Journal o Nanomaterials 01002003000.0750.10.125    P  a  r   t   i  c    l  e  s   i   z  e    (  n  m    ) Concentrations of DS (% w/v)Unloaded CS/DS nanoparticlesBSA-loaded CS/DS nanoparticles (a) 0204060800.0750.10.125    Z  e   t  a   p  o   t  e  n   t   i  a    l    (  m   V    ) Concentrations of DS (% w/v)Unloaded CS/DS nanoparticlesBSA-loaded CS/DS nanoparticles ∗∗∗ (b) F󰁩󰁧󰁵󰁲󰁥 󰀱: Particle size and zeta potential o unloaded and BSA-loaded CS/DS nanoparticles at various concentrations o DS (CS 󰀰.󰀰󰀷󰀵%w/v, BSA 󰀱mg/mL, 󰀳󰀷 ∘ C, mean  ± S.D,  󝠵 = 3 ).  ∗ Particle surace charge o BSA-loaded was signi󿬁cantly different rom unloaded CS/DSnanoparticles. 010020030040050010 min30 min1 h2 h12 h24 h    P  a  r   t   i  c    l  e  s   i   z  e    (  n  m    ) Incubation period0 min ∗∗∗ (a) 0100200300400500    P  a  r   t   i  c    l  e  s   i   z  e    (  n  m    ) 10 min30 min1 h2 h12 h24 hIncubation period0 min ∗∗∗ (b) 0204060    Z  e   t  a   p  o   t  e  n   t   i  a    l    (  m   V    ) −20 10 min30 min1 h2 h12 h24 hIncubation period0 min ∗∗∗ 0.075% (w/v)0.1% (w/v)0.125% (w/v) (c) 010203040    Z  e   t  a   p  o   t  e  n   t   i  a    l    (  m   V    ) −10−20−30∗∗∗ 10 min30 min1 h2 h12 h24 hIncubation period0 min0.075% (w/v)0.1% (w/v)0.125% (w/v) (d) F󰁩󰁧󰁵󰁲󰁥 󰀲: Particle size o unloaded (a) and BSA-loaded (b) and zeta potential o unloaded (c) and BSA-loaded (d) CS/DS nanoparticles at various concentrations o DS when incubated in FBS (CS 󰀰.󰀰󰀷󰀵% w/v, BSA 󰀱mg/mL, mean  ±  S.D,  󝠵 = 3 ).  ∗ Signi󿬁cantly different rom beoreincubation in serum. increases above its isoelectric point [󰀲󰀷] which avours theentrapment o BSA into the CS nanoparticles. 󰀳.󰀲. Colloidal Stability of CS/DS Nanoparticles󰀳.󰀲.󰀱. Stability in FBS.  Figures 󰀲(a) and 󰀲(b) clearly highlight that the particle size o both unloaded and BSA-loadedCS/DS nanoparticles (prepared rom 󰀰.󰀰󰀷󰀵% w/v o DS) wassigni󿬁cantly increased ( 􍠵 < 0.05 , independent  t  -test) at 󰀱hafer incubation and declined rom this point, up to 󰀲󰀴h aferincubation. Similarly, the zeta potential showed a signi󿬁cantdecline( 􍠵 < 0.05 ,independent t  -test)rom +56±5 and +34±4 mV to  +33 ± 3  and  +14 ± 2 mV or unloaded (Figure 󰀲(c))and BSA-loaded nanoparticles (Figure 󰀲(d)), respectively, at󰀱󰀰min afer incubation in FBS. A urther decrease in zetapotential o both nanoparticles was also observed over timeand this was thought to be caused by their interactions  Journal o Nanomaterials 󰀵 0200400600800    P  a  r   t   i  c    l  e  s   i   z  e    (  n  m    ) 10 min30 min1 h2 h12 h24 hIncubation period0 min ∗∗ (a) 0200400600    P  a  r   t   i  c    l  e  s   i   z  e    (  n  m    ) 10 min30 min1 h2 h12 h24 hIncubation period0 min ∗∗ (b) 020406080    Z  e   t  a   p  o   t  e  n   t   i  a    l    (  m   V    ) Human serumFetal bovine serum10 min30 min1 h2 h12 h24 hIncubation period0 min −20∗ (c) 0204060    Z  e   t  a   p  o   t  e  n   t   i  a    l    (  m   V    ) −20 10 min30 min1 h2 h12 h24 hIncubation period0 min ∗ Human serumFetal bovine serum (d) F󰁩󰁧󰁵󰁲󰁥 󰀳: Particle size o unloaded (a), BSA-loaded (b), and zeta potential o unloaded (c) and BSA-loaded (d) CS/DS nanoparticles in HSand FBS (CS 󰀰.󰀰󰀷󰀵% w/v, BSA 󰀱mg/mL, mean  ±  S.D,  󝠵 = 3 ).  ∗∗ Particle size o nanoparticles incubated in HS was signi󿬁cantly different romthe ones beore incubation and incubated in FBS. Particle surace charge o nanoparticles incubated in HS was signi󿬁cantly different romthe ones beore incubation and incubated in FBS. withnegativelychargedalbuminmacromolecules( >−20 mV)in the incubation media. Te obtained data also suggestedthat the unloaded nanoparticles were more strongly affectedby the components in FBS than the BSA-loaded one. Tismight be due to stronger ionic interactions between serumalbumin and the unloaded nanoparticles which had rela-tively higher positive surace charges as compared to BSA-loaded nanoparticles. Te positive surace charge density o BSA-loaded CS/DS nanoparticles was relatively low dueto the neutralization action by the BSA during the loadingstage [󰀲󰀸]. Moreover, charge neutralization by the albuminmacromolecules in the incubating media urther reducedrepulsion energy between nanoparticles. Tis may also pro-gressively acilitate agglomeration process and promote theormation o larger particles [󰀹]. Te tendency o particlesto orm aggregates was less prominent in case o BSA-loaded nanoparticles because they possessed relatively low zeta potential (positive charges) which subsequently resultedin a lesser degree o interaction with negatively chargedcomponents in FBS. Tus, this suggested that BSA-loadednanoparticles had better colloidal stability in FBS comparedto the unloaded ones. Tis 󿬁nding is important becausecolloidal stability in serum or any nanoparticulate systemdetermines the successul delivery o protein/peptides as itprevents particle aggregation or embolism rom occurring inthe systemic circulation [󰀲󰀹].Te incubation time is also a critical actor to determinethe adsorption pattern o proteins on the solid suraces o nanoparticles [󰀳󰀰]. In accordance to that, the particle sizeandzetapotentialounloadedandBSA-loadednanoparticlesweresigni󿬁cantlyin󿬂uencedbytheincubationtime.Figure 󰀲clearly shows the changing patterns o the particle size andzeta potential over extended incubation time. A signi󿬁cantdifference ( 􍠵 < 0.05 , paired  t  -test) was observed amongparticle sizes at the different postincubation time points. Forexample, BSA-loaded CS/DS nanoparticles prepared rom󰀰.󰀰󰀷󰀵% w/v o DS were ound to be enlarged rom  225 ±15  to  416 ± 21 nm at 󰀱h afer incubation in FBS as shownin Figure 󰀲(b). However, the particle size was subsequently decreased to  303 ± 18 nm at 󰀲h afer incubation in FBS. Tisphenomenoncouldbeexplainedbytheprocessoassociationand dissociation o protein molecules on the suraces o nanoparticles during the course o incubation. Tese resultswere also in accordance with previously published study thatreportedtheadsorption,anddesorptioncouldalsotakeplaceduring  in vivo  circumstances [󰀳󰀱]. Likewise, the continuous󿬂uctuations o the nanoparticles zeta potential in response o continuous attraction-repulsion processes may indicate thepositive and negative charge shifing between the particlesurace and serum components in order to stabilize thenanoparticles in the serum medium as shown in Figure 󰀲. 󰀳.󰀲.󰀲. Stability in HS.  For  in vivo  correlation, the stability analysis o unloaded and BSA-loaded CS/DS nanoparticles(produced rom DS 󰀰.󰀰󰀷󰀵% w/v) was urther carried out intheHSaspresentedinFigure 󰀳.Teselectiononanoparticlesproduced rom DS 󰀰.󰀰󰀷󰀵% w/v was based on the criterion
Search
Related Search
We Need Your Support
Thank you for visiting our website and your interest in our free products and services. We are nonprofit website to share and download documents. To the running of this website, we need your help to support us.

Thanks to everyone for your continued support.

No, Thanks