Extraction and characterization of gelatin from camel skin (potential halal gelatin) and production of gelatin nanoparticles

Gelatin is used as an ingredient in both food and non-food industries as a gelling agent, stabilizer, thickener, emulsifier, and film former. Porcine skins, bovine hides, and cattle bones are the most common sources of gelatin. However, mammalian gelatins are rejected by some consumers due to social, cultural, religious, or health-related concerns. In the present study, gelatin was obtained from camel skin as an alternative source using a combination of processing steps. Central composite design combined with response surface methodology was used to achieve high gelatin yields under different extraction conditions: temperatures of 40, 60, and 80 °C; pH values of 1, 4, and 7; and extraction times of 0.5, 2.0, and 3.5 min. Maximum gelatin yield from camel skin (29.1%) was achieved at 71.87 °C and pH 5.26 after 2.58 min. The extracted gelatin samples were characterized for amino acid profile, foaming capacity, film formation, foam stability, and gel strength (Bloom value). Gelatin nanoparticles were produced, and their morphology and zeta potential were determined. Bloom value of the camel skin gelatin was 340 g. Amino acid analysis revealed that the extracted gelatin showed high glycine and proline contents. Analysis of camel skin gelatin nanoparticle and functional properties revealed high suitability for food and non-food applications, with potential use in the growing global halal food market.     

Preparation and characterizations of self-assembled PEGylated gelatin nanoparticles

The PEGylated gelatin nanoparticles were prepared by self-assembling method and characterized. The gelatin drug carrier was proposed as a targeting drug delivery system with the hypothesis that the gelatin carrier could be degraded by the matrix metalloprotease (MMP) and release the anticancer drug loaded inside carriers around the cancer site. The gelatin nanoparticles proposed in this study were composed of deoxycholic acid (DOCA), monomethoxy polyethylene glycol (MPEG), and gelatin. The carboxyl groups of DOCA and carboxylated MPEG were coupled with amine group of gelatin by dichlorohexylcarbodiimide (DCC) method. One molecule of gelatin coupled with 205 molecules of MPEG and 275 molecules of DOCA. The synthesized gelatin/DOCA/MPEG conjugates (GDM) were ultrasonicated to produce self-assembled nanoparticles. DOCA acted as the hydrophobic core, thereby aggregating gelatin molecules and hydrophilic MPEG chains located at the surface of the nanoparticles. The concentration of GDM, intensity of sonication, sonication time and temperature, all affected to control the particle size in the ultrasonication. The optimum condition was obtained as 1.0 mg/mL of GDM, 28 W for sonication intensity, 3 min of sonication time, and room temperature. The size distribution of particle was found to be 100–1000 nm in this condition. The particles which had a broad size distribution were filtered by 0.2 μm membrane. The product yield of particles having below 200 nm of size was about 30%. After filtration, an average diameter of GDM nanoparticle was 176 nm (155–200 nm).     

Synthesis of metallic nanoparticles using plant extracts

Biomolecules present in plant extracts can be used to reduce metal ions to nanoparticles in a single-step green synthesis process. This biogenic reduction of metal ion to base metal is quite rapid, readily conducted at room temperature and pressure, and easily scaled up. Synthesis mediated by plant extracts is environmentally benign. The reducing agents involved include the various water soluble plant metabolites (e.g. alkaloids, phenolic compounds, terpenoids) and co-enzymes. Silver (Ag) and gold (Au) nanoparticles have been the particular focus of plant-based syntheses. Extracts of a diverse range of plant species have been successfully used in making nanoparticles. In addition to plant extracts, live plants can be used for the synthesis. Here we review the methods of making nanoparticles using plant extracts. Methods of particle characterization are reviewed and potential applications of the particles in medicine are discussed.     

Synthesis and characterization of cobalt oxide nanoparticles by thermal treatment process

The simple preparation of Co₃O₄ nanoparticles from a solid organometallic molecular precursor N-N′-bis(salicylaldehyde)-1,2-phenylenediimino cobalt(II); Co(salophen) has been achieved via two simple steps: firstly, the Co(salophen) precursor was precipitated from the reaction of cobalt(II) acetate and N-N′-bis(salicylaldehyde)-1,2-phenylenediimino; H₂salophen; in propanol under nitrogen condition; then, cubic phase Co₃O₄ nanoparticles with the size of mostly 30-50 nm could be produced by thermal treatment of the Co(salophen) in air at 773 K for 5 h. The as-synthesized products were characterized by powder X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS), and scanning electronic microscopy (SEM). These results confirm that the resulting oxide was pure single-crystalline Co₃O₄ nanoparticles. The optical property test indicates that the absorption peak of the nanoparticles shifts towards short wavelength, and the blue shift phenomenon might be ascribed to the quantum effect. The hysteresis loops of the obtained samples reveal the ferromagnetic behaviors the enhanced coercivity (H_c) and decreased saturation magnetization (M_s) in contrast to their respective bulk materials.     

Studies on the characteristics of drug-loaded gelatin nanoparticles prepared by nanoprecipitation

The morphology of gelatin nanoparticles loaded with three different drugs (Tizanidine hydrochloride, Gatifloxacin and Fluconazole) and their characteristics of entrapment and release from gelatin nanoparticles were investigated by the analysis on nanoparticle size distribution, SEM and FT-IR in this study. The particles were prepared by nanoprecipitation using water and ethanol as a solvent and a nonsolvent, respectively. The exclusion of a crosslinking agent from the procedure led the system to have an irregularly-shaped morphology. Nonetheless, the uncrosslinked case of Gatifloxacin loading generally led to a more homogeneous population of nanoparticles than the uncrosslinked case of Tizanidine hydrochloride loading. No loading was achieved in the case of Fluconazole, whereas both Tizanidine hydrochloride and Gatifloxacin are observed of being capable of being loaded by nanoprecipitation. Tizanidine hydrochloride-loaded, blank and Gatifloxacin-loaded nanoparticles yielded, under crosslinked condition, 59.3, 23.1 and 10.6% of the used dried mass. The crosslinked Tizanidine hydrochloride-loaded particles showed the loading efficiency of 13.8%, which was decreased to 1.1% without crosslinking. A crosslinker such as glutaraldehyde is indispensable to enhance the Tizanidine hydrochloride-loading efficiency. To the contrary, the Gatifloxacin-loading efficiency for crosslinked ones was lower by a factor of 2-3 times than that for uncrosslinked ones. This is due to the carboxylic groups of Gatifloxacin and the aldehyde groups of glutaraldehyde competing with each other during the crosslinking process, to react with the amino groups of gelatin molecules. The loading efficiency of gelatin nanoparticles reported by other investigators greatly varies. Nevertheless, the loading efficiency reported by us is in good agreement with the drug-loading data of gelatin nanoparticles reported by other investigators. The 80% of loaded Tizanidine hydrochloride was released around 15 h after start-up of the release experiment, while the 20% of loaded Gatifloxacin was released more rapidly, as free Gatifloxacin, than the loaded Tizanidine hydrochloride and it showed the trend of sustained slow release during the remaining period of its release experiment. Furthermore, the result of comparative FT-IR analysis is consistent to that of the corresponding drug release study.     

Therapeutic gene delivery and transfection in human pancreatic cancer cells using epidermal growth factor receptor-targeted gelatin nanoparticles

More than 32,000 patients are diagnosed with pancreatic cancer in the United States per year and the disease is associated with very high mortality (1). Urgent need exists to develop novel clinically-translatable therapeutic strategies that can improve on the dismal survival statistics of pancreatic cancer patients. Although gene therapy in cancer has shown a tremendous promise, the major challenge is in the development of safe and effective delivery system, which can lead to sustained transgene expression. Gelatin is one of the most versatile natural biopolymer, widely used in food and pharmaceutical products. Previous studies from our laboratory have shown that type B gelatin could physical encapsulate DNA, which preserved the supercoiled structure of the plasmid and improved transfection efficiency upon intracellular delivery. By thiolation of gelatin, the sulfhydryl groups could be introduced into the polymer and would form disulfide bond within nanoparticles, which stabilizes the whole complex and once disulfide bond is broken due to the presence of glutathione in cytosol, payload would be released (2-5). Poly(ethylene glycol) (PEG)-modified GENS, when administered into the systemic circulation, provides long-circulation times and preferentially targets to the tumor mass due to the hyper-permeability of the neovasculature by the enhanced permeability and retention effect (6). Studies have shown over-expression of the epidermal growth factor receptor (EGFR) on Panc-1 human pancreatic adenocarcinoma cells (7). In order to actively target pancreatic cancer cell line, EGFR specific peptide was conjugated on the particle surface through a PEG spacer.(8) Most anti-tumor gene therapies are focused on administration of the tumor suppressor genes, such as wild-type p53 (wt-p53), to restore the pro-apoptotic function in the cells (9). The p53 mechanism functions as a critical signaling pathway in cell growth, which regulates apoptosis, cell cycle arrest, metabolism and other processes (10). In pancreatic cancer, most cells have mutations in p53 protein, causing the loss of apoptotic activity. With the introduction of wt-p53, the apoptosis could be repaired and further triggers cell death in cancer cells (11). Based on the above rationale, we have designed EGFR targeting peptide-modified thiolated gelatin nanoparticles for wt-p53 gene delivery and evaluated delivery efficiency and transfection in Panc-1 cells.     

Synthesis of alkyl esters by cutinase in miniemulsion and organic solvent media

The main objective of this work was studying and testing the nature and influence of reaction media (organic solvent vs. miniemulsion system) on the synthesis of alkyl esters catalyzed by Fusarium solani pisi cutinase. Ester synthesis and cutinase selectivity for different chain length of acids and alcohols (ethyl and hexyl) were evaluated. In iso-octane, after 1 h of reaction, cutinase exhibits rates of esterification between 0.24 μmol x mg^–1 x min^–1 for ethyl oleate and 1.15 μmol x mg^–1 x min^–1 for ethyl butyrate, while in a miniemulsion system the rates were from 0.05 for ethyl heptanoate to 0.76 μmol x mg^–1 x min^–1 for ethyl decanoate. The reaction rate for the synthesis of hexyl esters in a miniemulsion system was from 0.19 for hexyl heptanoate to 1.07 μmol x mg^–1 x min^–1 for hexyl decanoate. High conversion yields of 95% at equilibrium after 8 h of reaction in iso-octane for pentanoic acid (C₅) with ethanol at equimolar concentration (0.1 M) was achieved. Additionally, this work showed that a significant and unexpected shift in cutinase selectivity occurred towards longer chain length carboxylic acids (C₈–C₁₀) in miniemulsion system as compared to organic solvent (iso-octane) and previous studies in reverse micellar systems. The possibility of working with higher concentration of substrates, without inhibitory effect on the enzyme, was another advantage of the miniemulsion system.     

Experimental optimization of a real time fed-batch fermentation process using Markov decision process

This article describes a methodology that implements a Markov decision process (MDP) optimization technique in a real time fed-batch experiment. Biological systems can be better modeled under the stochastic framework and MDP is shown to be a suitable technique for their optimization. A nonlinear input/output model is used to calculate the probability transitions. All elements of the MDP are identified according to physical parameters. Finally, this study compares the results obtained when optimizing ethanol production using the infinite horizon problem, with total expected discount policy, to previous experimental results aimed at optimizing ethanol production using a recombinant Escherichia coli fed-batch cultivation. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 317-327, 1997.     

Synthesis of soluble coordination polymer nanoparticles using room-temperature ionic liquid

We report a new family of soluble cyano-bridged coordination polymer nanoparticles M₃[Cr(CN)₆]₂/[BMIM][BF₄] (where M²⁺ = Ni, Mn, V^IVO; BMIM = 1-butyl-3-methylimidazolium). These nanoparticles of ca. 6 nm were synthesised in the ionic liquid 1-butyl-3-methylimidazolium tetrafluoroborate, which acts both as a stabilizing agent and a solvent. The magnetic properties of frozen colloids containing the nanoparticles show that the relaxation of magnetisation is strongly influenced by interparticle interactions leading to the appearance of spin-glass-like dynamics in these systems.     

Synthesis of gold nanotriangles and silver nanoparticles using Aloe vera plant extract

Biogenic gold nanotriangles and spherical silver nanoparticles were synthesized by a simple procedure using Aloe vera leaf extract as the reducing agent. This procedure offers control over the size of the gold nanotriangle and thereby a handle to tune their optical properties, particularly the position of the longitudinal surface plasmon resonance. The kinetics of gold nanotriangle formation was followed by UV-vis-NIR absorption spectroscopy and transmission electron microscopy (TEM). The effect of reducing agent concentration in the reaction mixture on the yield and size of the gold nanotriangles was studied using transmission electron microscopy. Monitoring the formation of gold nanotriangles as a function of time using TEM reveals that multiply twinned particles (MTPs) play an important role in the formation of gold nanotriangles. It is observed that the slow rate of the reaction along with the shape directing effect of the constituents of the extract are responsible for the formation of single crystalline gold nanotriangles. Reduction of silver ions by Aloe vera extract however, led to the formation of spherical silver nanoparticles of 15.2 nm +/- 4.2 nm size.     

Purification of fibronectin using gelatin derivatives

This work compares two distinct methods, column and batch, for the purification of fibronectin, from different plasma fractions, using gelatin and one of its derivates (Hemoce, Behring). The yields of both techniques at quantitative as well as qualitative levels (levels of immunologically active fibronectin), are evaluated. The data indicate that better conservation of immunological immunological characteristics is obtained with the use of gelatin derivatives (Hemoce). The plasma fraction does not have significant influence on the process yields.     

基于微生物生物合成纳米颗粒机制的研究进展

纳米粒子的合成方法多种多样,包括物理法、化学法和生物合成法,其中生物合成法是以生物为基体的绿色合成方法。由于微生物易于培养、生长快、廉价易得,已成为纳米粒子生物合成法的重要生物类群。微生物和纳米材料的多样性决定了其合成机制的多样化。本文结合国内外的科研报道,着重介绍了目前纳米粒子生物合成机制,并对纳米粒子微生物合成技术未来发展趋势进行了展望。     

Synthesis of silver nanoparticles using haloarchaeal isolate Halococcus salifodinae BK3

Numerous bacteria, fungi, yeasts and viruses have been exploited for biosynthesis of highly structured metal sulfide and metallic nanoparticles. Haloarchaea (salt-loving archaea) of the third domain of life Archaea, on the other hand have not yet been explored for nanoparticle synthesis. In this study, we report the intracellular synthesis of stable, mostly spherical silver nanoparticles (AgNPs) by the haloarchaeal isolate Halococcus salifodinae BK₃. The culture on adaptation to silver nitrate exhibited growth kinetics similar to that of the control. NADH-dependent nitrate reductase was involved in silver tolerance, reduction, synthesis of AgNPs, and exhibited metal-dependent increase in enzyme activity. The AgNPs preparation was characterized using UV–visible spectroscopy, XRD, TEM and EDAX. The XRD analysis of the nanoparticles showed the characteristic Bragg peaks of face-centered cubic silver with crystallite domain size of 22 and 12 nm for AgNPs synthesized in NTYE and halophilic nitrate broth (HNB), respectively. The average particle size obtained from TEM analysis was 50.3 and 12 nm for AgNPs synthesized in NTYE and HNB, respectively. This is the first report on the synthesis of silver nanoparticles by haloarchaea.     

Synthesis of ethyl isovalerate using Rhizomucor miehei lipase: optimization

Immobilized lipase from Rhizomucor miehei (Lipozyme IM-20) was used to catalyze the esterification reaction between isovaleric acid and ethanol to synthesize ethyl isovalerate in n-hexane. Response surface methodology based on five-level four-variable central composite rotatable design was employed to optimize four important reaction variables such as enzyme/substrate E/S ratio, substrate concentration, incubation time, and temperature affecting the synthesis of ethyl isovalerate. The optimum conditions predicted for achieving maximum ester yield (500 mM) are as follows: E/S ratio, 48.41 g/mol; substrate concentration, 1 M; reaction time, 60 h; temperature, 60 degrees C. The predicted value matched well with experimentally obtained value of 487 mM.     

Synthesis of chitosan-stabilized polymer dispersions,capsules, and chitosan grafting products via miniemulsion

The potential of chitosan as an emulsion stabilizer is combined with the miniemulsion technique to generate oil droplets, hollow capsules, and latex particles in the diameter range of 100-300 nm carrying a functional biopolymer surface. It turned out that chitosan alone, independent of its molecular weight, is just moderately efficient, which is speculatively attributed to its stiff polysaccharide structure. The addition of biocompatible costabilizers with higher flexibility either to the oil phase or to the water phase, such as Jeffamine or Gluadin (a peptide), eliminates these deficiencies, and very small nanocapsules made of biopolymer hybrids can be obtained. NMR analysis shows that the costabilizer is effectively grafted/cross-linked to the chitosan which makes the miniemulsion route also effective for the modification of hard-to-handle, amphiphilic biopolymers either from the water or from the oil phase.     

Rapid identification of gelatin and casein hydrolysis using TCA

Using TCA as the developing agent we have identified the hydrolysis of gelatin and casein within 3 h. When compared with conventional gelatin and casein hydrolysis techniques we have found the results of the TCA enhancement to be more rapid and sensitive than the conventional methods.     

Temperature-dependent selective purification of plasmid DNA using magnetic nanoparticles in an RNase-free process

The efficiency of transformation by electroporation has been known to be compromised by strain dependency. A high efficiency protocol is still lacking for distinct two-hybrid yeast strains of diverse genetic features. Here, we used 0.5 M lithium acetate (LiAc) and 50 mM Tris-HCl with 5 mM EDTA (pH 7.5), i.e., fivefold the standard concentrations, and voltage at 1.0 to develop a protocol which, for the first time, is able to effect an average efficiency of 1.84 × 10⁶ transformants/μg DNA for three commonly used yeast strains committed to two-hybrid screening experiments.     

Analysis of the freezing process across temperature gradients within gelatin gels.

Previous investigators of freezing rate distribution within the bulk of aqueous material have concluded that the fastest freezing occurs at the outermost, and in the central region of the sample, while the slowest occurs in the intermediate regions. The present work challenges the universality of the conclusion by presenting detailed experimental evidence on the distribution of the ice phase across gelatin gels and by furnishing related time-temperature recordings. The analysis of the ice structure together with the time-temperature record provides information required to reconstruct the distribution of the freezing rates. The results of the study indicate that the rate of freezing in the central region of the specimen varies, but most frequently is the lowest of all, and depends on the gel concentration, freezing temperature and the specimen size. The freezing curves for gelatin gels exhibit an unusual configuration, in that their freezing plateau assumes an increasingly saddle-shaped form as the concentration of gelatin increases from sample to sample. A possible explanation of this phenomenon is suggested. It is shown that supercooling within the gelatin samples can persist long after the onset of freezing. In order to explain how the specimen interior can remain in the supercooled state while being surrounded by the converging ice front, a graphical representation of the temperature changes within the specimen is presented. An alternative explanation is mentioned very briefly.     

Synthesis of biologically stable gold nanoparticles using imidazolium-based amino acid ionic liquids

A novel double-step reduction procedure for the synthesis of gold nanoparticles (AuNPs) using amino acid ionic liquids has been employed. 1-Dodecyl-3-methyl imidazolium tryptophan ([C(12)mim]Trp) and 1-ethyl-3-methyl imidazolium tryptophan ([C(2)mim]Trp) were used for this synthesis. The synthesized AuNPs were characterized by UV-vis spectroscopy, transmission electron microscopy and dynamic light scattering. The behavior of these AuNPs were also probed in a biological media. It was proven that AuNPs synthesized at [C(12)mim]Trp have more stability than AuNPs synthesized at [C(2)mim]Trp due to the longer alkyl chain of the imidazolium moiety. The solubility test shows that the resultant AuNPs have a hydrophilic nature. Finally, it was seen that due to the presence of a biomolecule, namely Trp, in the structure of AuNPs protecting shell, higher stability and biocompatibility was achieved in the biological media.