The methionine precursor dl-2-hydroxy-(4-methylthio)butanoic acid protects intestinal epithelial barrier function

dl-2-hydroxy-(4-methylthio)butanoic acid (HMTBA) is a source of dietary methionine (Met) that is widely used in poultry nutrition. We have previously shown that HMTBA is preferentially diverted to the transsulfuration pathway, which gives antioxidant metabolites such as taurine and glutathione. Therefore, here we hypothesize that this Met source can protect epithelial barrier function in an in vitro model of intestinal inflammation of Caco-2 cells. The results show that HMTBA prevents the increase in paracellular permeability induced by H₂O₂ or tumour necrosis factor-α. This effect can be attributed to the increased production of taurine and reduced glutathione. Similar results were obtained for dl-Met, although the protective role of the amino acid was less pronounced than that of the hydroxy analogue. In conclusion, the diversion to the transsulfuration pathway means that this Met precursor is of greater value than previously thought, due to its capacity to improve intestinal homeostasis and the quality of poultry products destined for human consumption.     

灵芝片抗疲劳作用实验研究

目的研究林中宝灵芝片抗疲劳的作用。方法按小鼠体重分为高、中、低剂量组。分别灌胃给予1.03、0.36、0.18g/kg的灵芝片30d,对照组给予生理盐水。通过小鼠负重游泳实验测定小鼠耐缺氧时间、小鼠肝糖原含量改变及小鼠血清尿素氮含量变化等。观察小鼠运动性疲劳时林中宝灵芝片对体内相关指标的影响及其机制。结果林中宝灵芝片组小鼠游泳时间、耐缺氧力、肝糖原含量、血清尿素氮含量降低均与生理盐水组有显著差异。结论林中宝灵芝片有明显的抗疲劳作用。     

聚乙二醇与蛋白质相互作用的荧光和共振散射光谱研究

用荧光光谱和共振散射光谱(RLS)研究了聚乙二醇(PEG)与牛血清白蛋白(BSA)在溶液中的相互作用.通过考察PEG浓度对体系荧光光谱和RLS光谱的变化规律,探讨了两者之间的相互作用机理.结果表明PEG与BSA之间的相互作用较弱,对BSA的结构和构象影响较小.然而,铜离子的加入可以有效的促进PEG与BSA之间的相互作用,不仅引起体系荧光有规律的猝灭,而且使得体系共振散射光谱出现持续的增强.     

聚醚砜微孔膜对牛血清蛋白吸附性能的研究

主要研究了聚醚砜(PES)微孔膜对牛血清蛋白(BSA)的吸附性能.用BCA(bicinchoninincacid)法测定了膜孔中吸附的蛋白量.运用扫描电子显微镜和电子单纱强力仪分别对PES膜结构和力学性能进行表征.随着聚乙二醇20000(PEG20000)与聚乙二醇400(PEG400)比值的增大牛血清蛋白的吸附量先减小后增大,水通量先增大后减小,截留率始终维持在97.93%～99.39%之间,结果表明,PEG20000与PEG400比例为2时PES膜的亲水性最好,对BSA的吸附量最小.     

Electroseparation of an antibacterial peptide fraction from snow crab by-products hydrolysate by electrodialysis with ultrafiltration membranes

Recently, a snow crab by-products hydrolysate has demonstrated antibacterial properties due to a peptide with a molecular weight of about 800 Da, but only at high concentration. Consequently, peptide hydrolysate has been fractionated to obtain peptides in a more purified form. The aim of this work was to separate a snow crab by-products hydrolysate by electrodialysis with ultrafiltration membranes (EDUF). EDUF, which allows separation of molecules according to their charges and molecular weights, was used to recover and concentrate the active antibacterial fraction. Two different ultrafiltration membranes (20 and 50 kDa) and two electrical field strengths (2 and 14 V/cm) were used as separation parameters. After EDUF separation, the 300-600 Da peptide molecular weight range was the most recovered with an abundance of 94%. Moreover, fractionation at 14 V/cm with ultrafiltration membranes of 50 kDa allowed the recovery of an anionic fraction which showed antibacterial properties on Escherichia coli ATCC 25922 and Listeria innocua HPB 13.     

Interactions of different polyphenols with bovine serum albumin using fluorescence quenching and molecular docking

Polyphenols are responsible for the major organoleptic characteristics of plant-derived foods and beverages. Here, we investigated the binding of several polyphenols to bovine serum albumin (BSA) at pH 7.5 and 25 °C: catechins [(−)-epigallocatechin-3-gallate, (−)-epigallocatechin, (−)-epicatechin-3-gallate], flavones (kaempferol, kaempferol-3-glucoside, quercetin, naringenin) and hydroxycinnamic acids (rosmarinic acid, caffeic acid, p-coumaric acid). Fluorescence emission spectrometry and molecular docking were applied to compare experimentally determined binding parameters with molecular modelling. Among these polyphenols, (−)-epicatechin-3-gallate showed the highest Stern–Volmer modified quenching constant, followed by (−)-epigallocatechin-3-gallate. Similarly, (−)-epicatechin-3-gallate had the highest effect on the Circular Dichroic spectrum of BSA, while the changes induced by other polyphenols were negligible. Molecular docking predicted high binding energies for (−)-epicatechin-3-gallate and (−)-epigallocatechin-3-gallate for the binding site on BSA near Trp213. Our data reveal that the polyphenol structures significantly affect the binding process: the binding affinity generally decreases with glycosylation and reduced numbers of hydroxyl groups on the second aromatic ring.     

Mass spectrometry of peptides and proteins from human blood

It is difficult to convey the accelerating rate and growing importance of mass spectrometry applications to human blood proteins and peptides. Mass spectrometry can rapidly detect and identify the ionizable peptides from the proteins in a simple mixture and reveal many of their post‐translational modifications. However, blood is a complex mixture that may contain many proteins first expressed in cells and tissues. The complete analysis of blood proteins is a daunting task that will rely on a wide range of disciplines from physics, chemistry, biochemistry, genetics, electromagnetic instrumentation, mathematics and computation. Therefore the comprehensive discovery and analysis of blood proteins will rank among the great technical challenges and require the cumulative sum of many of mankind's scientific achievements together. A variety of methods have been used to fractionate, analyze and identify proteins from blood, each yielding a small piece of the whole and throwing the great size of the task into sharp relief. The approaches attempted to date clearly indicate that enumerating the proteins and peptides of blood can be accomplished. There is no doubt that the mass spectrometry of blood will be crucial to the discovery and analysis of proteins, enzyme activities, and post‐translational processes that underlay the mechanisms of disease. At present both discovery and quantification of proteins from blood are commonly reaching sensitivities of ～1 ng/mL. © 2010 Wiley Periodicals, Inc., Mass Spec Rev 30:685–732, 2011     

A reliable and sensitive time-resolved fluorescent immunochromatographic assay (TRFICA) for ochratoxin A in agro-products

Immunochromatographic assays (ICAs) are considered as a suitable diagnostic tool for the detection of mycotoxins. Mycotoxins and especially, ochratoxin A are analytes with more demanding sensitivity requirements. To enhance the sensitivity of current immunochromatographic assays for ochratoxin A (OTA), a novel sensitive ICA was developed in this study. In the assay, microspheres enclosing fluorescent europium (III) [Eu(III)] nanoparticles (EuNPs) were used as a label for OTA monoclonal antibody (OTA-mAb) conjugation. Accordingly, assay was called time-resolved fluorescent immunochromatographic assay (TRFICA). The test strip was composed of three parts: a sample pad, nitrocellulose membrane and an absorbent pad. As for detection, a proper concentration of conjugated microspheres was pipetted into the microtube and sample extract was added to it. Then the strip was inserted into the tube and the fluid flow along the strip. The TRFICA results were obtained in 8 min and read by a portable TRFICA strip reader. The established method allows quantitative determination of OTA with limit of detection as low as 1.0 μg kg⁻¹ in the samples. For validation, spiked samples including wheat, maize, soybean and rice were respectively assayed by TRFICA and a standard high performance liquid chromatography equipped with a fluorescence detector (HPLC-FLD), and good agreement of results was obtained between two methods.     

Isolation of recombinant antibody fragments (scFv) by phage display technology for detection of almond allergens in food products

Tree nut allergies are considered an important health issue in developed countries. To comply with the regulations on food labeling, reliable allergen detection methods are required. In this work we isolated almond-specific recombinant antibody fragments (scFv) from a commercial phage display library bypassing the use of live animals, hence being consistent with the latest policies on animal welfare. To this end an iterative selection procedure employing the Tomlinson I phage display library and a crude almond protein extract was carried out. Two different almond-specific scFv (named PD1F6 and PD2C9) were isolated after two rounds of biopanning, and an indirect phage ELISA was implemented to detect the presence of almond protein in foodstuffs. The isolated scFvs demonstrated to be highly specific and allowed detection of 40 ng mL⁻¹ and 100 ng mL⁻¹ of raw and roasted almond protein, respectively. The practical detection limit of the assay in almond spiked food products was 0.1 mg g⁻¹ (110–120 ppm). The developed indirect phage ELISA was validated by analysis of 92 commercial food products, showing good correlation with the results obtained by a previously developed real-time PCR method for the detection of almond in foodstuffs. The selected phage clones can be affinity maturated to improve their sensitivity and genetically engineered to be employed in different assay formats.