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1.
Lotus corniculatus and Lotus uliginosus are agronomically important forage crops used in ruminant livestock production. The condensed tannin (CT) content, dry matter (DM) production, and persistence of these species are key characteristics of interest for future exploitation of these crops. Here we present field data on 19 varieties of L. corniculatus, 2 varieties of L. uliginosus and, additionally, a glasshouse experiment using 6 varieties of L. corniculatus and 2 varieties of L. uliginosus. Current methods for the quantification of condensed tannins in crop species are slow and labor intensive and are generally based upon polymer hydrolysis following the extraction of chlorophyll in a liquid phase. Presented here is a high-throughput protocol for condensed tannin quantification suitable for microtiter plates based upon the precipitation of condensed tannin polymers in complex with bovine serum albumin (BSA) with subsequent hydrolysis of precipates using butan 1-ol/ hydrochloric acid.  相似文献   

2.
Glomalin is described in the literature as a N-linked glycoprotein and the putative gene product of arbuscular mycorrhizal fungi (AMF). Since the link between glomalin and various protein fractions in soil is not yet clearly defined, glomalin-related soil protein (GRSP) more appropriately describes glomalin's existence in natural organic matter (NOM). The objective of this study was to examine the chemical characteristics of GRSP present in several mineral and organic soils of varying organic carbon content. GRSP was isolated using high temperature sodium citrate extraction followed by either trichloroacetic acid (TCA) or hydrochloric acid (HCl) precipitation. GRSP was characterized by quantitative solid-state 13C DPMAS NMR, infrared (IR) spectroscopy, elemental analysis, and the Bradford assay for protein content. GRSP accounted for 25% and 52% of total C in the mineral soils and organic soil, respectively. Molar C/N and H/C ratios reveal that GRSP has less nitrogen than bovine serum albumin (BSA), and that GRSP extracted from the Pahokee peat soil possessed a more unsaturated, and thus aromatic character relative to the mineral soil GRSP, respectively. GRSP's high aromatic (42-49%) and carboxyl (24-30%) carbon contents and low aliphatic (4-11%) and carbohydrate-type carbon contents (4-16%) suggests that GRSP does not resemble a typical glycoprotein. In fact, the NMR spectra of GRSP closely resemble that of humic acid. GRSP extracted from mineral and organic soils possessed the same NMR fingerprint regardless of the precipitation method used (i.e., either TCA or HCl). It is likely that the current GRSP extraction methods, because of their similarity to the method used to extract humic acid, are coextracting both materials.  相似文献   

3.
There is increasing evidence that tannins affect degradation of soil organic matter and nitrogen cycling. It is assumed that the influence of tannins on biochemical processes is partly related to their ability to precipitate proteins. However, there is almost no information about precipitation of organic nitrogen-containing compounds other than proteins. A few studies indicate that tannins can precipitate arginine, choline or chitosan. In this study we used commercial tannic acid and condensed tannins extracted from Norway spruce (Picea abies (L.) Karst) needles to investigate precipitation of a wide range of organic nitrogen compounds including amino acids (all 20 proteinaceous), peptides (insulin, oxidized glutathion, reduced glutathion, AlaAla, GlyGlu, GlyPhe, GlyGlyGly), proteins (bovine serum albumin, Rubisco i.e. d-ribulose 1,5-diphosphate carboxylase), nitrogen bases, polyamines and aminosugars (N-acetyl-d-glucosamine, chitin and chitosan). Our results showed that tannins can precipitate a subset of these compounds - of the amino acids only arginine, of the peptides studied only insulin, all the proteins, polyamines, nitrogen bases, chitin and chitosan, but not N-acetyl-d-glucosamine. Concentrations of organic nitrogen compound and tannins affected amount of these compounds in precipitates. Moreover, pH value affected precipitation. The amount of precipitated organic nitrogen compound and the amount of precipitated tannins showed positive correlation across different pH. Precipitation of organic N-containing compounds other than protein by tannins can potentially affect reactions in all biochemical mixtures including tannins and these organic nitrogen compounds, and affect soil N cycling.  相似文献   

4.
Polyphenols are capable of binding to proteins and form polyphenol-protein complexes thus reducing the release of N from decomposing plant materials. The objective of this work was to test if under polyphenol-rich vegetations adapted microbial communities had developed capable of breaking down recalcitrant polyphenol-protein complexes. Soils used for this investigation were from different 10-year-old tropical agricultural systems (maize, sugarcane plots and Gliricidia sepium or Peltophorum dasyrrachis woodlots) and natural systems (secondary forest and Imperata cylindrica grassland). TA (tannic acid, hydrolysable tannin), QUE (quebracho, condensed tannin), BSA (bovine serum albumin, protein) or TA/BSA and QUE/BSA polyphenol-protein complexes were incubated at 28 °C in these soils. CO2-C and 13C evolution were periodically monitored and mineral N release, microbial biomass N and phospholipid fatty acid (PLFA) profiles measured at the end.QUE was able to bind about 25% more protein than TA. In all systems the individual uncomplexed substrates were more easily degraded than the complexes. On average, net cumulative CO2-C evolution from TA/BSA complexes was more than 5 times higher than from QUE/BSA complexes, indicating higher C availability and/or lower protection capability of TA compared to QUE. However, net N release was higher from QUE/BSA than from TA/BSA probably due to their higher protein-binding capacity and associated larger degradation of partly unprotected protein as suggested by 13C-CO2 signatures. Microbial respiration patterns indicated that polyphenol complexes were initially degraded more quickly in the maize cropping system than in soils from under polyphenol-rich communities (Peltophorum and natural forest) but this pattern reversed with time. Long-term incubation of QUE/BSA complexes even caused a negative effect on microbial respiration in agricultural soils with low polyphenol contents (e.g. maize and sugarcane).Incubation of polyphenol complexes in soil depressed microbial biomass N in maize, sugarcane, Imperata and forest systems and led to reduced soil pH. However, microbial biomass was increased under the polyphenol-rich vegetation of Peltophorum. The PLFA group 18:2w6,9 was highly enhanced by condensed tannin-protein complexes additions as compared to control and hydrolysable polyphenol-protein complexes in soils with high polyphenol contents. Polyphenol complexes increased the fungi:bacteria ratio in systems with a high polyphenol content, particularly with condensed tannin complexes. The results indicated that systems with a high polyphenol content favoured development of fungal communities that are highly adaptable to phenol-rich soil conditions and high acidity, particularly with regards to the more recalcitrant condensed tannin-protein complexes.  相似文献   

5.
Some nutritional and functional properties of defatted wheat germ protein   总被引：24，自引：0，他引：24  
Defatted wheat germ protein (DWGP) was isolated by alkaline extraction at pH 9.5 and subsequent isoelectric precipitation at pH 4.0, and its nutritional and functional properties were studied. The results showed that the amino acid content of defatted wheat germ was as high as 26.793 g/100 g, and the contents of eight essential amino acids were all relatively high. The isoelectric point of DWGP was 4.0. When pH >6.0, the DWGP had high solubility with a nitrogen solubility index of 70%. The emulsifying activity and emulsifying stability of DWGP were similar to those of bovine serum albumin and a little higher than those of casein. DWGP had good foaming capacity, but its foaming stability (FS) was not very good. However, the FS of DWGP can be improved through physical, chemical, or enzymatic methods. Moreover, DWGP had excellent water retention (WR); especially at pH 8.0 and a temperature of 70 degrees C, the WR of DWGP was the highest at 229.4%. DWGP offers is a potential source of functional protein isolate for possible food applications.  相似文献   

6.
The objective of the present investigation was to examine the oral astringency and protein-binding activity of four structurally well-defined tannins, namely, procyanidin [epicatechin16(4-->8)catechin], pentagalloyl glucose (1,2,3,4,6-penta-O-galloyl-beta-D-glucopyranose), castalagin, and grandinin, representing the three main structural categories of tannins, the proanthocyanidins, the gallotannins, and the ellagitannins. Astringency threshold and dos/response were determined by the half-tongue test using a trained human panel. Protein-binding stoichiometry and relative affinity were determined using radioiodinated bovine serum albumin in precipitation or competitive binding assays. Procyanidin and pentagalloyl glucose were perceived as highly astringent compounds and had relatively steep dose/response curves, but castalagin and grandinin had a lower mass threshold for detection. In vitro, procyanidin was the most effective protein-precipitating agent and grandinin the least. Increasing the temperature increased protein precipitation by the hydrolyzable tannins, especially grandinin. All four polyphenols had higher relative affinities for proline-rich proteins than for bovine serum albumin.  相似文献   

7.
小麦Rubisco抗体的制备和旗叶中Rubisco含量的ELISA测定   总被引：2，自引：0，他引：2  
Rubisco(核酮糖-1,5-二磷酸羧化酶/加氧酶)为光合生物中的关键性酶。以小麦(Triticum aestivum L. cv.)叶片为材料，在(NH4)2SO4盐析和DEAE-Sepharose F F层析纯化的基础上添加了Sephacryl S 300分子筛，进一步纯化了Rubisco。 将纯化的Rubisco免疫大白兔制备了Rubisco的多克隆抗体， 经Western blot检测，Rubisco抗体与Rubisco呈免疫反应特异性。并运用Rubisco抗体，采用ELISA方法直接测定小麦旗叶生长发育过程中Rubisco含量的变化，结果表明：不同生长阶段，旗叶的Rubisco含量与光合速率大小呈正相关。该方法比免疫沉淀法更为快捷、准确。  相似文献   

8.
土壤可培养真菌RAPD扩增条件的优化   总被引：4，自引：0，他引：4  
李钧敏  金则新 《土壤通报》2004,35(3):295-298
以改进的CTAB-溶菌酶-蛋白酶K裂解法抽提土壤可培养真菌总DNA,直接进行随机扩增多态DNA(RAPD)分析。分别测试了镁离子浓度,4×dNTP浓度,模板DNA量,引物用量,Taq酶用量和牛血清白蛋白浓度对反应结果的影响,通过各因子的组合研究,确定了土壤可培养真菌遗传多样性分析的稳定的RAPD反应体系:15μlPCR反应体积,1×Taq酶配套缓冲液(10mmolL-1Tris·HClpH9.0,50mmolL-1KCl,0.1%TritonX-100),1.5mmolL-1MgCl2,2UTaq酶(上海华美公司),20ng模板DNA,15pmol引物(上海Sangon公司);dATP、dCTP、dGTP、dTTP各0.2mmolL-1;1mgmL-1牛血清白蛋白。  相似文献   

9.
Two diffusion-based NMR techniques are presented and used to investigate the binding of selected flavor compounds to macromolecules. A pulsed field gradient NMR (PFG-NMR) method was applied to measure the apparent diffusion coefficients of four alkanone compounds as they associated with bovine serum albumin (BSA). The change in the apparent diffusion coefficient as a function of the BSA/alkanone ratio was fitted to yield binding constants (K(a)()) and binding stoichiometry (n) for each alkanone. The results showed that the apparent diffusion coefficients of alkanones increased with a decrease in the BSA/alkanone ratios, and the measured values of K(a)() and n were comparable with those obtained with other methods and depended on the alkanone structure. A diffusion-based nuclear Overhauser effect (called diffusion NOE pumping) method was also applied to screen mixtures of flavor compounds and identify those that have a binding affinity to complex macromolecules. Using this technique benzaldehyde and vanillin were observed to bind with bovine serum albumin, whereas 2-phenylethanol was identified as a nonbinding or weakly binding ligand with BSA. The diffusion NOE pumping method was also applied to a hydro alcoholic solution of cacao bean tannin extracts to which a mixture of ethylbenzoate, benzaldehyde, and 2-phenylethanol was added. The diffusion NOE pumping technique clearly indicated that ethylbenzoate had a stronger binding affinity to the polymeric (-)-epicatechin units of the cacao bean tannin extracts than the other two flavor compounds. The results successfully demonstrate the potential applications of diffusion-based NMR techniques for studying flavors and nonvolatile food matrix interactions.  相似文献   

10.
A simple, fast, and more selective approach is presented in this study for the identification of haze-active proteins. Grape seed proteins were unfolded by 1% SDS and then interacted with different amounts of tannin at 4 degrees C, followed by gel electrophoresis. It was found that the intensity of the band at 45 kDa was decreased as tannins increased. The amino acid composition of this isolated 45-kDa protein was higher in proline (9.49%) than the average proline content of total grape seed proteins (4.85%). To verify the selectivity of the proposed method, a globular protein (bovine serum albumin, BSA) and a proline-rich protein (gelatin) were selected and used in the model system. As expected, gelatin was removed as it reacted with the increasing added tannins, whereas BSA did not. These results showed that it is possible to identify haze-active proteins by modulating the accessibility of protein to tannins, suggesting this new method can be used by the beverage industry to trouble-shoot haze problems and for quality control.  相似文献   

11.
In vitro digestions were performed on pearl millet flours with decreased phytate contents and on two dephytinized or nondephytinized pearl millet grain fractions, a decorticated fraction, and a bran fraction with low and high fiber and tannin contents, respectively. Insoluble residues of these digestions were then incubated with buffer or enzymatic solutions (xylanases and/or phytases), and the quantities of indigestible iron and zinc released by these different treatments were determined. In decorticated pearl millet grain, iron was chelated by phytates and by insoluble fibers, whereas zinc was almost exclusively chelated by phytates. In the bran of pearl millet grain, a high proportion of iron was chelated by iron-binding phenolic compounds, while the rest of iron as well as the majority of zinc were chelated in complexes between phytates and fibers. The low effect of phytase action on iron and zinc solubility of bran of pearl millet grain shows that, in the case of high fiber and tannin contents, the chelating effect of these compounds was higher than that of phytates.  相似文献   

12.
Protein oxidation in meat is considered to decrease meat tenderness due to protein disulfide cross-link formation of thiol-containing amino acid residues. An LC-MS method for detection of thiol-quinone adducts (RS-QH(2)) in myofibrillar proteins was developed to investigate the interaction between phenols, as protective antioxidants, and proteins from meat under oxidative conditions using aqueous solutions of (i) cysteine (Cys), (ii) glutathione (GSH), (iii) bovine serum albumin (BSA), or (iv) a myofibrillar protein isolate (MPI). The aqueous solutions were incubated at room temperature (30 min) with 4-methyl-1,2-benzoquinone (4MBQ) prepared from oxidation of 4-methylcatechol (4MC) by periodate resin or incubated at room temperature (5 h) with 4MC and Fe(II)/H(2)O(2). GSH, BSA, and MPI were hydrolyzed (6 N HCl, 110 °C, 22 h) after incubation, and the cysteine-quinone adduct, Cys-QH(2) (m/z 244.2) was identified according to UV and mass spectra after separation on an RP-C18 column. The thiol-quinone adduct was present in all thiol systems after incubation with 4MBQ or 4MC oxidized by Fe(II)/H(2)O(2). Direct reaction with 4MBQ resulted in each case in increased Cys-QH(2) formation compared to simultaneous oxidation of thiol source and 4MC with Fe(II)/H(2)O(2). The covalent bonds between quinones and thiol groups may act as a potential antioxidant by inhibiting disulfide protein cross-link formation.  相似文献   

13.
小麦花药蛋白质组双向电泳技术体系的优化   总被引：15，自引：1，他引：14  
以小麦单核期花药为材料,比较了两种不同蛋白质提取方法TCA/丙酮法和酚提取法及不同的蛋白质裂解液组成对双向电泳的影响,并在蛋白质上样量和SDS-PAGE胶浓度等方面也进行了探索与优化。结果表明,采用TCA/丙酮提取法比用酚提取法提取的蛋白质所得的2-DE胶图谱上蛋白质点数增加,图谱背景也比较清晰;样品溶解于含有硫脲和TBP的蛋白质裂解液Ⅱ中,可显著提高蛋白质的溶解性,在2-DE图谱上可分辨出554个蛋白质点,比用蛋白质裂解液Ⅰ提取的蛋白多39个点。以TCA-丙酮法提取小麦花药组织中的蛋白,用蛋白质裂解液Ⅱ[7mol/L尿素、2mol/L硫脲、4%CHAPS、2%TBP、65mmol/L DTT、0.2%载体两性电解质(其中0.1%pH 3~10,0.1%pH4~6)]溶解蛋白,以pH4~7线性17cm的IPG胶条进行双向凝胶电泳,在上样量为800μg,13%SDS-PAGE胶浓度下,蛋白质得到了更好的分离,2-DE图谱上可分辨出631个蛋白点,图谱质量最佳。优化后的双向电泳技术体系,适合于小麦花药全蛋白质的双向电泳分析。  相似文献   

14.
Food allergies represent an important health problem in industrialized countries, such that detection and quantitative analysis of the protein considered to be the main allergen is crucial. A dot-blot fluorescent staining method for the detection and quantitative analysis of protein residues in food grade amino acids and nucleic acids is presented. This method combines fluorescence staining with dot-blotting onto PVDF membrane. Several standard proteins, such as bovine serum albumin (66 kDa), lysozyme (14 kDa), ubiquitin (8.6 kDa), bovine insulin (5.7 kDa), and oxidized insulin B chain (3.5 kDa), were detectable at 0.1 ppm using SYPRO Ruby blot stain. Twenty-five different amino acids and two different nucleic acids of food grade were analyzed using this method combined with an internal standard addition method using BSA as an internal standard. All amino acids and nucleic acids were dissolved in 3.6% aqueous HCl and dot-blotted onto PVDF membrane before a large amount of amino acids and nucleic acid were removed. Protein residues and the internal standard protein immobilized on the membrane were stained using SYPRO ruby blot stain. The internal standard in all samples was detectable at 0.1 ppm. Samples were dissolved at 120 or 70 mg/mL, according to their solubility under acidic conditions. The detection limit of protein residues per weight was 0.8-1.4 ppm in amino acids and nucleic acids; residual protein was not detected in any sample.  相似文献   

15.
Microsample deproteinization of bovine milk was carried out on-line using a series of restricted access medium (RAM) bovine serum albumin (BSA) columns: C8, C18, phenyl, and cyano. The four different columns prepared showed a high percentage of protein exclusion using water as the mobile phase and provided an appropriate retention profile for a series of five cephalosporin antibiotics (cefoperazone, cephacetril, cephalexin, cephapirin, and ceftiofur). Chromatographic conditions such as washing time, buffer pH, and type and percentage of organic modifier were fully evaluated with respect to the protein elution profile and retention of the antibiotic by RAM column. One of these columns was chosen to develop and validate a method for the determination of cefoperazone in bovine milk. The system used in this work was composed of a RAM-BSA phenyl column coupled to a C18 analytical column. The standard curve was linear over the range 0.100-2.50 microg/mL. The limits of quantification and detection were 0.100 and 0.050 microg/mL, respectively. The developed method showed high intermediate precision (CV of 2.37-2.63%) and accuracy (90.7-94.3%) with adequate sensitivity for drug monitoring in bovine milk samples.  相似文献   

16.
Simulations of gastro-intestinal digestion, used to estimate in vitro iron and zinc availability, were performed on two kinds of samples: (i) samples with decreased phytate contents from whole pearl millet flour and (ii) nondephytinized or dephytinized samples from two pearl millet grain fractions, a decorticated fraction with low fiber and tannin contents and a bran fraction with high fiber and tannin contents. Iron and zinc in vitro availabilities of whole pearl millet flour were significantly improved by phytate degradation, even if the IP6 were not all degraded. Total dephytinization of decorticated fraction led to a marked increase in iron and zinc in vitro availabilities, but that of bran fraction had no effect on either iron or zinc in vitro availability. Even if phytates are involved in reducing in vitro iron and zinc availability in pearl millet flour, fibers and tannins play an important role by chelating a high proportion of iron and zinc in grain hulls.  相似文献   

17.
Genetically modified crops, that produce Cry insecticidal crystal proteins (Cry) from Bacillus thuringiensis (Bt), release these toxins into soils through root exudates and upon decomposition of residues. The fate of these toxins in soil has not yet been clearly elucidated. Persistence can be influenced by biotic (degradation by microorganisms) and abiotic factors (physicochemical interactions with soil components, especially adsorption). The aim of this study was to follow the fate of Cry1Aa Bt toxin in contrasting soils subjected to different treatments to enhance or inhibit microbial activity, in order to establish the importance of biotic and abiotic processes for the fate of Bt toxin. The toxin was efficiently extracted from each soil using an alkaline buffer containing a protein, bovine serum albumin, and a nonionic surfactant, Tween 20. The marked decline of extractable toxin after incubation of weeks to months was soil-dependent. The decrease of extractable toxin with incubation time was not related to microbial degradation but mainly to physicochemical interactions with the surfaces that may decrease immunochemical detectability or enhance protein fixation. Hydrophobic interactions may play an important role in determining the interaction of the toxin with surfaces.  相似文献   

18.
添加菌体蛋白和尿素对玉米秸秆厌氧发酵的影响   总被引：3，自引：2，他引：1  
王阳  林聪  侯锦  侯雨 《农业工程学报》2011,27(13):74-78
以玉米秸秆为原料的厌氧发酵,碳氮比（C/N）一直是影响其发酵的主要问题之一。该文针对玉米秸秆的C/N进行了研究,通过对比试验的方法,以尿素和菌体蛋白作为氮源添加剂配制成C/N分别为25/1和35/1的秸秆发酵料液,进行30d的厌氧发酵试验。结果表明,以尿素和菌体蛋白作为氮源调整C/N时,C/N为35/1时的产气量要好于C/N为25/1;与秸秆单独发酵相比,C/N为35/1的添加菌体蛋白试验组R2和添加尿素试验组R4的总产气量分别提高了31.43%和2.69%。此外,添加菌体蛋白和尿素能有效抑制秸秆体积的漂浮膨胀,对料液起到酸碱缓冲的作用。菌体蛋白作为秸秆发酵添加剂应用在沼气工程中可以变废为宝,提高经济效益。  相似文献   

19.
Buckwheat (Fagopyrum esculentum Moench) protein (BWP) exhibits hypocholesterolemic activity in several animal models by increasing fecal excretion of neutral and acidic sterols. In the current study, the ability of BWP to disrupt micelle cholesterol solubility by sequestration of cholesterol was investigated. When BWP (0.2%) was incubated with cholesterol and micelle lipid components prior to micelle formation, cholesterol solubility was reduced 40%. In contrast, cholesterol solubility was not decreased when BWP (0.2%) was incubated after micelle formation and incorporation of soluble cholesterol. Buckwheat flour, from which BWP was derived, had no significant effect on cholesterol solubility. Cholesterol uptake in Caco-2 cells from micelles made in the presence of BWP (0.2%) was reduced by 47, 36, 35, and 33% when compared with buckwheat flour, bovine serum albumin, casein, and gelatin, respectively. Reduction in cholesterol uptake in Caco-2 cells was dose-dependent, with maximum reductions at 0.1-0.4% BWP. In cholesterol-binding experiments, 83% of the cholesterol was associated with an insoluble BWP fraction, indicating strong cholesterol-binding capacity that disrupts solubility and uptake by Caco-2 cells.  相似文献   

20.
Chromatographic separation of soluble proteins from rice (Oryza sativa L.) yielded a major albumin protein (16 kDa), with the DHHQVYSPGEQ sequence in the N terminus, showing antioxidant action. The rice albumin was more potent than other rice proteins in preventing Cu2+-induced low-density lipoprotein (LDL) oxidation. Additionally, it also exhibited a remarkable suppression of HOCl oxidation. In a further study, albumin inhibited Cu2+-induced oxidation of LDL in a stoichiometric manner with an EC50 value of 4.3 microM, close to that of serum albumins. Moreover, after digestion with trypsin or chymotrypsin, it maintained its antioxidant action. In an experiment to see the involvement of the N terminus in antioxidant action, a synthetic tetrapeptide, equivalent to the N terminus DHHQ, was found to inhibit Cu2+-induced LDL oxidation or degradation of apolipoprotein B, similar to that of rice albumin. In mechanistic analyses, the action of rice albumin or tetrapeptide is primarily due to the removal of Cu2+, as suggested from its inhibitory effect on Cu2+/diphenylcarbohydrazide (DPCH) complex formation. However, despite its similar inhibitory effect on Cu2+-induced oxidation of LDL, rice albumin was less effective than serum albumin in inhibiting Cu2+/DPCH complex formation, suggesting that the number of Cu2+-binding sites in rice albumin may be less than that in serum albumins. Taken together, rice albumin exerts a potent preventive action against Cu2+-induced oxidations, which is due to the Cu2+ binding by DHHQ in the N-terminal sequence. Such a role as a Cu2+ chelator would add up to the application of rice albumin protein.  相似文献   

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