Extraction and characterisation of gelatine from Atlantic salmon (Salmo salar) skin

Gelatine was extracted from Atlantic salmon and Atlantic cod skin by the acid extraction process. After filtration and ion exchange treatment the extracts were colourless and free from fishy odour. In three separate experiments the average yields of gelatine from salmon and cod skins were 39.7% (+/-2.2%) and 44.8% (+/-0.2%) respectively, on a dry matter basis. Gelatine from salmon contained slightly more hydroxyproline and proline (16.6%) than cod gelatine (15.4%), whereas the content of serine was lower (4.6% versus 6.3%). Salmon gelatine expressed slightly higher gelling temperature (12 degrees C) than cod gelatine (10 degrees C), and higher initial gel strength. During storage at 10 degrees C, gel strengths were increased and more so with gels made from cod than from salmon gelatine. Hence, gels made from cod and salmon gelatines extracted at 56 degrees C achieved the same gel strength (195g) after 7days of storage. Gelatines extracted at a higher temperature (65 degrees C) gave lower gel strengths.     

Optimization of gelatine extraction from grass carp (Catenopharyngodon idella) fish skin by response surface methodology

To establish the optimum gelatine extraction conditions from grass carp fish skin, response surface methodology (RSM) was adopted in this study. The effects of concentration of HCl (%, A), pre-treatment time (h, B), extraction temperature ( degrees C, C) and extraction time (h, D) were studied. The responses were yield (%) and gel strength (g). A=1.19%, B=24 h, C=52.61 degrees C and D=5.12h were determined as the optimum conditions while the predicted responses were 19.83% yield and 267 g gel strength. Gelling and melting points were 19.5 degrees C and 26.8 degrees C, respectively. Moreover, grass carp gelatine showed high contents of imino acids (proline and hydroxyproline) 19.47%. RSM provided a powerful tool to optimize the extraction parameters and the results may be adapted for industrial extraction of gelatine from grass carp fish skins.     

An alternative real-time PCR method for the detection of thermotolerant Bacillus sensu lato contaminants in naturally-contaminated gelatine

Aims:  Comparison of an internally-controlled real-time PCR assay with the current plate-based assay for the detection of Bacillus sensu lato contaminants in gelatine.
Methods and Results:  A comprehensive TaqMan^® probe was designed allowing the real-time PCR assay to be fully inclusive for the gelatine-contaminating Bacillus s.l. species. An internal amplification control was implemented at 500 copies per reaction without impact on target detection. Specific and selective detection of target cells was achieved with a quick and simple DNA preparation procedure. No significant difference (Kappa value = 0·94) was observed between the performance of the real-time PCR and the current plate-based method on naturally contaminated gelatines ( n  = 162). Relative accuracy, relative sensitivity and relative specificity were 97·5%.
Conclusions:  The real-time PCR assay is an adequate alternative of the current plate-based assay.
Significance and Impact of the Study:  The real-time PCR assay decreased the time between sample collection and result from 2 days to 2 h. The gelatine-producing industry can ensure gelatine quality in a much faster way.     

An alternative real-time PCR method to detect the Bacillus cereus group in naturally contaminated food gelatine: a comparison study

Aims:  Comparison of an internally controlled real-time PCR assay with the standard plate-based assay (ISO 21871) for the detection of Bacillus cereus group cells in gelatine.
Methods and Results:  A comprehensive TaqMan probe was designed allowing the real-time PCR assay to be fully inclusive and exclusive. An internal amplification control was designed and implemented at 500 copies per reaction without impact on target detection. Specific and selective detection of target cells was achieved with a quick and simple DNA preparation procedure. No significant difference (κ = 0·99) was observed between the performance of the real-time PCR and the standard plate-based method on naturally contaminated gelatines ( n  = 197). Relative accuracy, relative sensitivity and relative specificity were ≥99%.
Conclusions:  The real-time PCR assay is a valid alternative of the standard plate-based assay.
Significance and Impact of the Study:  The real-time PCR assay decreased the time between sample collection and result from 2 days to 2 h, while analysis cost did not increase. The gelatine-producing industry can ensure gelatine safety and quality in a much faster way.     

Physical and chemical properties of selected agricultural byproduct-based activated carbons and their ability to adsorb geosmin

The objectives of this study were to evaluate selected physical and chemical properties of agricultural byproduct-based activated carbons made from pecan shells and sugarcane bagasse, and compare those properties to a commercial coal-based activated carbon as well as to compare the adsorption efficiency of these carbons for geosmin. Comparison of the physical and chemical properties of pecan shell- and bagasse-based carbons to the commercial carbon, Calgon Filtrasorb 400, showed that pecan shell carbon, but not the bagasse carbon, compared favorably to Filtrasorb 400, especially in terms of surface area, bulk density, ash and attrition. A carbon dosage study done in a model system showed the amount of geosmin adsorbed to be greater for Filtrasorb 400 and the bagasse-based carbon at low carbon concentrations than for the pecan shell carbons, but geosmin adsorption was similar in all carbons at higher carbon dosages. Application of the Freundlich isotherm model to the adsorption data showed that carbons made by steam activation of pecan shells or sugarcane bagasse had geosmin adsorption characteristics most like those of the commercial carbon. In terms of physical, chemical and adsorptive properties, steam-activated pecan shell carbon most resembled the commercial carbon and has the potential to replace Filtrasorb 400 in applications involving removal of geosmin from aqueous environments.     

Agar Processed from Gracilaria foliifera of Florida

Agar producing a firm gel with specific rheological properties as well as a good commercial value was processed from Gracilaria foliifera (Forsskal) Børgesen, a red alga harvested along the Florida west coast. The alkali pretreatment of the seaweed based on the Funaki-Kojima’s method was applied before the process of agar extraction. The chemical and physical properties of the processed agar were compared with other typical agar experimentally processed.     

Molecular weight distribution of chitosan isolated from Mucor rouxii under different culture and processing conditions

The isolation of chitosan from a fungal source offers the potential of a product with controlled physicochemical properties not obtainable by the commercial chemical conversion of crustacean chitin. A variety of culture and processing protocols using Mucor rouxii were studied for their effects on biomass yield and chitosan molecular weight. Weight-averaged molecular weight determined by gel permeation chromotography ranged from 2.0 x 10(5) to approximately 1.4 x 10(6) daltons. The chitosan yield ranged from 5% to 10% of total biomass dry weight and from 30% to 40% of the cell wall. Of the culture parameters studied, length of incubation and medium composition effected biomass production and molecular weight. Modification of the processing protocol, including the type and strength of acid, and cell wall disruption in acid prior to refluxing were used to optimize the efficiency of chitosan extraction.The degree of deacetylation of fungal and commercial chitosans was compared using infrared spectrometry, titration, and first derivative of UV absorbance spectrometry. The chitosan obtained directly from the fungal cell wall had a higher degree of deacetylation than commercial chitosan from the chemical conversion process.     

Comparison of the structural and physical properties of human hair eumelanin following enzymatic or acid/base extraction

Eumelanin was isolated from a sample of black, Indonesian human hair using three different published procedures: two different acid/base extractions and an enzymatic extraction. The morphology and spectroscopic properties of the isolated pigments differ significantly. The acid/base procedures both yield an amorphous material, while enzymatic extraction yields ellipsoidal melanosomes. Amino acid analysis shows that there is protein associated with the isolated pigments, accounting for 52, 40 and 14% of the total mass for the two acid/base extractions and the enzymatic extraction, respectively. The amino acid compositions do not correlate with those of keratin or tyrosinase. Metal elemental analysis shows that the acid/base extraction removes a majority of many metal ions bound to the pigment. Chemical degradation analysis by KMnO4/H+ and H2O2/OH- indicates significant differences between the pigments isolated by acid/base and enzymatic extraction. After correction for the protein mass in the two pigments, the lower yields of both pyrrole-2,3,5-tricarboxylic acid and pyrrole-2,3-dicarboxylic acid, eumelanin degradation products, indicate acid/base extraction modifies the chemical structure of the melanin, consistent with the result of Soluene solubilization assay. While the optical absorption spectra of the bulk pigments are similar, the spectra of the molecular weight less than 1000 mass fractions differ significantly. The data clearly indicate that pigment obtained from human hair by acid/base extraction contains significant protein, exhibits destruction of the melanosome, and possesses altered molecular structure. The acid/base extracted hair melanin is not representative of the natural material and is a poor model system for studying the physical and biological properties of melanins. The enzymatically extracted hair melanin, on the contrary, retains the morphology of intact melanosomes and is an excellent source of human melanin.     

赤楠果实红色素的性质及提取工艺

本实验探讨赤楠果实红色素的理化性质及提取条件。结果表明,该色素属花色苷类;其最佳提取工艺是:1%HCl-乙醇(1ml HCl:99ml 95%乙醇)溶液作为提取剂,料液比1/10,温度35℃,提取时间3h,提取次数3次。     

Comparison of customized spin-column and salt-precipitation finger-prick blood DNA extraction

gDNA (genomic DNA extraction from blood is a fundamental process in many diagnostic, identification and research applications. Numerous extraction methods have been reported and are available commercially. However, there is insufficient understanding of the impact of chemical buffers on DNA yield from either whole or nucleated blood. Moreover, these commercial kits are often costly, constraining less well-funded laboratories to traditional and more cost-effective salt-precipitation methods. Towards this, we compared a salt-precipitation and a customized cost-effective spin-column-based method, studying the impact of different chemical constituents on the yields. This customized method resulted in a shortening of the extraction process, higher gDNA yields, and more successful PCR amplification of gDNA genes compared with the salt-precipitation method. Optimizing different chemical buffers on whole- and nucleated blood materials further revealed that certain chemicals boosted extractions from whole- but not nucleated blood. These findings may be useful to laboratories that do not have ready access to commercial kits, and improve their nucleic acid extractions from blood economically.     

Glyphosate-induced stiffening of HaCaT keratinocytes, a Peak Force Tapping study on living cells

The skin is the first physiological barrier, with a complex constitution, that provides defensive functions against multiple physical and chemical aggressions. Glyphosate is an extensively used herbicide that has been shown to increase the risk of cancer. Moreover there is increasing evidence suggesting that the mechanical phenotype plays an important role in malignant transformation. Atomic force microscopy (AFM) has emerged within the last decade as a powerful tool for providing a nanometer-scale resolution imaging of biological samples. Peak Force Tapping (PFT) is a newly released AFM-based investigation technique allowing extraction of chemical and mechanical properties from a wide range of samples at a relatively high speed and a high resolution. The present work uses the PFT technology to investigate HaCaT keratinocytes, a human epidermal cell line, and offers an original approach to study chemically-induced changes in the cellular mechanical properties under near-physiological conditions. These experiments indicate glyphosate induces cell membrane stiffening, and the appearance of cytoskeleton structures at a subcellular level, for low cytotoxic concentrations whereas cells exposed to IC50 (inhibitory concentration 50%) treatment exhibit control-like mechanical behavior despite obvious membrane damages. Quercetin, a well-known antioxidant, reverses the glyphosate-induced mechanical phenotype.     

皮肤模型与微针穿刺性能评价的研究

微针阵列作为新型透皮给药技术,受到广泛关注。通常以刺入力、刺入率和刺入深度来评价微针刺入皮肤的程度和效率。皮肤是其性能评价的基础。皮肤的物理特性主要由角蛋白丝、胶原纤维、弹性纤维和皮下组织综合决定,并且从厚度、弹性、硬度和韧性等维度反映其对微针刺入的影响。机械的、渗透的、组织的和屏障的等皮肤模型被用于解读和模拟真实皮肤的该方面功能。同样,通过皮肤力学分析后建立的包括本构模型在内的各种皮肤力学模型也从物理维度解析皮肤的力学特征。真实皮肤具有复杂性,存在差异性大、不易获取和储存,以及伦理等问题,而皮肤模型可在一定程度上代替真实皮肤辅助微针递送系统设计、开发和性能评价。本文系统回顾分析了皮肤组织的物理特性、各种皮肤模型的制备及特点、真实皮肤和模拟皮肤在评价微针穿刺性能方面的应用,为开发及建立合适的皮肤模型提供借鉴。     

Comparison of suitable protein measurement for Thalassiosira weissflogii (Bacillariophyta) and Tetraselmis chuii (Chlorophyta)

To perform effective measurement of the protein contents, it is necessary to apply the most suitable extraction technique. We compared protein extraction techniques for two microalgal species with entirely different cell wall structures, such as diatom Thalassiosira weissflogii and green flagellate Tetraselmis chuii, using mechanical, physical, and chemical extraction methods. Our results showed that in Th. weissflogii, the control treatment without extraction and mechanical disruption using ultrasonication was sufficient to disrupt its brittle cell wall, resulting in the protein yield of 19.03 and 19.46% of dry weight (DW), respectively. In case of Te. chuii, the chemical extraction using 6% trichloroacetic acid at 95°C was the most suitable extraction method, resulting in the protein yield of 23.78% of DW. Therefore, the most suitable extraction technique must be individually selected for each species depending on their cell wall properties.     

Properties of polysaccharide produced by Azotobacter vinelandii cultured on 4-hydroxybenzoic acid

AIMS: Characterization of the exopolysaccharide produced by Azotobacter vinelandii grown on 4-hydroxybenzoic acid (EPS I), and the comparison between this exopolysaccharide and commercial alginate, constituted the main objective of this work. METHODS AND RESULTS: Total carbohydrates, uronic acids, acetyl and pyruvyl groups and proteins were determined by colorimetric methods and composition was confirmed by Nuclear Magnetic Resonance studies. Rheological properties were analysed under different physical and chemical conditions. Results showed differences between EPS I and commercial alginate, in relation to both composition and viscosity. Higher amount of guluronnosyl residues were found in EPS I, whereas commercial alginate contained the same proportion of mannuronosyl and guluronnosyl residues. In accordance with this result, EPS I gave rise to solutions of higher viscosity than commercial alginate, although solutions of this polysaccharide showed greater stability when conditions were altered. CONCLUSIONS: The exopolysaccharide produced by A. vinelandii grown on 4-hydroxybenzoic acid showed a different composition in comparison with commercial alginate, which leads to higher viscosity values for the aqueous solutions of EPS I. SIGNIFICANCE AND IMPACT OF STUDY: This work describes for the first time the characteristics of an exopolysaccharide produced by A. vinelandii from 4-hydroxybenzoic acid, a substrate rarely used as sole carbon source.     

Hibiscus vitifolius,a new fibre crop

This is a discussion of an investigation conducted on the morphology and anatomy ofHibiscus vitifolius, the physical and chemical properties of its fibre and yarn, and a preliminary study on reaction to dyes. The quality and physical and chemical properties of the fibre closely resemble those of jute. But further agronomic investigations are needed to assess more fully its potentialities as a commercial crop to supplement the growing demand for jute. Because of its short reed length and higher tex values, it may be best utilized, at present, as a mixture with jute.     

Study of the bacterial load in a gelatine production process focussed on Bacillus and related endosporeforming genera

Gelatine is an animal protein with many industrial applications. Previous studies pointed out that endosporeforming bacteria, belonging to the genus Bacillus or related genera, might contaminate and survive the production process of gelatine, leading to products of low quality and safety. The aim of this study is to determine the bacterial diversity of contaminants isolated from a gelatine production chain with emphasis on aerobic endosporeforming bacteria. Contaminants were isolated from samples taken at five crucial points along two different production lines of a gelatine production process and from water supplies used for extraction and cooling. Gaschromatographic methyl ester analysis of fatty acids was performed to differentiate isolates at the genus level. Apart from members of the genus Bacillus or related endosporeforming genera, also members of Salmonella, Kluyvera, Staphylococcus, Burkholderia, Enterococcus, Pseudomonas, Yersinia, Streptococcus and Brevundimonas could be detected. Isolates identified as belonging to Bacillus and related endosporeforming genera were further characterised by gelatinase tests, rep-PCR and 16S rDNA sequencing. All these isolates showed the ability to liquefy gelatine. Endosporeforming isolates were assigned to Bacillus licheniformis, B. fumarioli, members of the B. cereus group, B. badius, B. coagulans, B. subtilis, Brevibacillus agri, Alicyclobacillus acidocaldarius and a yet undescribed Paenibacillus species.     

Characteristics and Gel Properties of Gelatin from Skin of Asian Bullfrog (<Emphasis Type="Italic">Rana tigerina</Emphasis>)

Characteristics and gel properties of gelatin from frog skin as influenced by extraction temperatures (45–75 °C) were investigated. Yield of gelatin increased as the extraction temperature increased (P < 0.05). All gelatins contained α- and β-chains as the predominant components and showed a high imino acid content (215 residues/1000 residues). Fourier transform infrared (FTIR) spectra indicated that all gelatin samples had major peaks in amide regions. Gelatin extracted at 55 °C exhibited the highest gel strength (P < 0.05), which was similar to that of commercial bovine gelatin (P > 0.05). Gelling and melting temperatures of frog skin gelatin were 23.47–24.87 and 33.22–34.66 °C, respectively. Gels became more yellowish with increasing extraction temperatures (P < 0.05). All gelatin gels were sponge or coral-like in structure but varied in patterns as visualized by scanning electron microscopy (SEM). Gelatin from frog skin could be used as a replacement for land animal counterpart.     

Reactive extraction of lactic acid using alamine 336 in MIBK: equilibria and kinetics

Lactic acid is an important commercial product and extracting it out of aqueous solution is a growing requirement in fermentation based industries and recovery from waste streams. The design of an amine extraction process requires (i) equilibrium and (ii) kinetic data for the acid-amine (solvent) system used. Equilibria for lactic acid extraction by alamine 336 in methyl-iso-butyl-ketone (MIBK) as a diluent have been determined. The extent to which the organic phase (amine +MIBK) may be loaded with lactic acid is expressed as a loading ratio, z=[HL](o)/[B](i,o). Calculations based on the stoichiometry of the reactive extraction and the equilibria involved indicated that more lactic acid is transferred to the organic phase than would be expected from the (1:1) stoichiometry of the reaction. The extraction equilibrium was interpreted as a result of consecutive formation of two acid-amine species with stoichiometries of 1:1 and 2:1. Equilibrium complexation constant for (1:1) and (2:1) has been estimated. Kinetics of extraction of lactic acid by alamine 336 in MIBK has also been determined. In a first study of its kind, the theory of extraction accompanied by a chemical reaction has been used to obtain the kinetics of extraction of lactic acid by alamine 336 in MIBK. The reaction between lactic acid and alamine 336 in MIBK in a stirred cell falls in Regime 3, extraction accompanied by a fast chemical reaction occurring in the diffusion film. The reaction has been found to be zero order in alamine 336 and first order in lactic acid with a rate constant of 1.38 s(-1). These data will be useful in the design of extraction processes.     

Presence of phosphorus in Nephila clavipes dragline silk.

Solid-state 31P-NMR of Nephila clavipes dragline silk indicates the presence of phosphorus in at least two chemically distinct environments. Amino acid analyses of acid-hydrolyzed silk confirm the presence of phosphotyrosine as one of the phosphorus-containing components. The unusual chemical shift (18.9 ppm downfield from 85% H3PO4), proton chemical shift, and acid lability of a second component suggest that it is part of a strained five-membered cyclic phosphate that might be found on a beta-D-ribose. The five-membered cyclic phosphate is not removed from the silk fibers by exhaustive aqueous extraction. It is absent in nascent silk fibroin from the glands, suggesting that its formation is part of the fiber processing that occurs in the ducts leading to the spinnerets. High-resolution NMR spectra of silk dissolved in propionic acid/12 N HCl (50:50 v/v) show five phosphorus sites assigned to phosphorylated tyrosine residues, phosphorylated serine residues, inorganic phosphate, and two hydrolysis products of the cyclic phosphate compound. The observed posttranslational phosphorylation may be important in the processing and modulation of the physical properties of dragline silk.     

Relations between extraction protocols for activated sludge extracellular polymeric substances (EPS) and complexation properties of Pb and Cd with EPS: Part II. Consequences of EPS extraction methods on Pb2+ and Cd2+ complexation

To study the effect of extraction protocols on extracellular polymeric substances (EPS) metal binding ability, EPS from two activated sludges were extracted by eight extraction protocols: three chemical treatments, four physical treatments and a control. Two pK_a, for each EPS, were determined: pK_a1 may be specific for carboxyl and phosphoric and pK_a2 may be attributed to phenolic and amino functional groups according to EPS composition and IR spectra. EPS pK_a values could be affected by the presence of extraction reagents and/or the modifications of EPS by extraction reagents.Complexation study performed at pH 7 by a polarographic method has always showed a greater affinity of EPS for Pb²⁺ than for Cd²⁺. The complexation properties of EPS extracted by chemical methods were greatly modified. Concerning EPS extracted by physical methods, their complexation properties were close except for EPS obtained by heating. Standardized extraction methods must be established as a function of the aims of the EPS study.