Electrogenerated bromine as a coulometric reagent for the estimation of the bioavailability of polyphenols

A method is proposed for the estimation of the bioavailability of polyphenols using electrogenerated bromine as a coulometric titrant. The titration of model solutions of casein and bovine serum albumin (BSA) shows that casein does not interact with electrogenerated bromine, while BSA reacts with the titrant in the ratio 1: 63. The proteins bind rutin and quercetin (from 14 to 90%) at a high rate and thus reduce the bioavailability of polyphenols. The concentration of free polyphenol is reduced with an increase in the concentration of protein in the mixture. The total antioxidant capacity (AOC) of tea is determined. Green tea is shown to possess higher AOC than the black one because of the partial oxidation of polyphenols to respective thearubigins and theaflavins at the fermentation step in the production of green tea. The total AOC of tea drops from 7 to 85%, in proportion to the increase in the amount of milk in the mixture. Milk proteins bind tea polyphenols into complexes because of intermolecular interactions and thus reduce their bioavailability. The observed effect of milk is independent of the brand of black tea. The degree of reduction of the total AOC of tea in going from one tea to another remains virtually constant.     

茶多酚在水和N,N-二甲基甲酰胺中的溶解行为及动力学模型

用电导方法研究了茶多酚在水和二甲基甲酰胺(DMF)溶剂中的溶解行为,并导出了溶解动力学模型:dC/dt=k2C2+k1C+k0。结果显示,茶多酚溶液的浓度、溶解温度以及溶剂都会影响其溶解行为。温度升高,有利于茶多酚的溶解;增大茶多酚在溶液中的浓度,反应速率加快,而且茶多酚在H2O中比在DMF中更容易溶解,这跟溶剂本身的酸碱性有关。通过建立溶解动力学模型发现,所有的溶解过程基本上都是零级反应,其次是一级反应,而基本上不发生二级反应。     

Immobilised tyrosinase-based biosensor for the detection of tea polyphenols

An amperometric principle-based biosensor containing immobilized enzyme tyrosinase has been used for detection of polyphenols in tea. The immobilized tyrosinase-based biosensor could detect tea polyphenols in the concentration range 10–80 mmol L⁻¹. Immobilization of the enzyme by the crosslinking method gave good stable response to tea polyphenols. The biosensor response reached the steady state within 5 min. The voltage response was found to have a direct linear relationship with the concentration of polyphenols in black tea samples. Enzyme membrane fouling was observed with number of analyses with a single immobilised enzyme membrane. The tyrosinase-based biosensor gave maximum response to tea polyphenols at 30°C. The optimum pH was 7.0. This biosensor system can be applied for analysis of tea polyphenols. Variation in the biosensor response to black tea infusions gave an indication of the different amounts of theaflavins in the samples, which is an important parameter in evaluating tea quality. A comparative study of the quality attributes of a variety of commercially available brands of tea were performed using the biosensor and conventional analytical techniques such as spectrophotometry.     

NMR of casein hydration and activity in solutions with ions

Oxygen-17 NMR studies of caseins isolated from fresh milk of several species were carried out in concentrated electrolyte solutions as a function of both protein and electrolyte concentration. Molecular dynamics simulations of ion and water binding to caseins are compared with the experimental observations by 0–17 NMR of caseins in solutions with ions. Protein activities are also determined by utilizing Wyman's theory of linked functions with a detailed model of ion binding to casein.     

Speciation of aluminium in tea infusions by use of SEC and FPLC with ICP–OES and ES–MS–MS detection

Speciation of Al in tea infusions was studied by size exclusion chromatography (SEC) and anion-exchange fast protein liquid chromatography (FPLC). Fractions were collected throughout the chromatographic separations and Al was determined “off line” by inductively coupled plasma optical emission spectroscopy (ICP–OES). Black, green, and red tea samples were investigated. The total concentration of Al in tea infusions was determined by ICP–OES and ranged between 0.5 and 4 mg dm⁻³. The pH of tea infusions ranged between 5.3 and 5.5. Data from SEC–ICP–OES analysis indicated that 10–35% of total Al in tea infusions was eluted at a retention volume corresponding to a molecular mass of approximately 3800 Da. The remaining Al was adsorbed on the column resin. The same tea infusions were also analysed by anion-exchange FPLC–ICP–OES. It was found experimentally that the same percentage of total Al as from the SEC column was eluted at a retention volume that corresponded to negatively charged Al-citrate. The remaining Al was adsorbed on the column resin. Identification of Al-binding ligands eluting under the chromatographic peak was performed by electrospray ionisation tandem mass spectrometry (ES–MS–MS) analysis. It was proven that ionic Al species in tea infusions (10–35% of the total Al) corresponded to negatively charged Al-citrate. The remaining species that was adsorbed on the SEC or FPLC columns was most probably bound to phenolic compounds. Speciation of Al in tea with milk or lemon was also studied. Results for tea with milk indicated that Al-citrate was not transformed and that approximately 60% of total Al was transformed into high-molecular-mass Al species. This fraction was subjected to sodium dodecyl sulfonate polyacryl gel electrophoresis (SDS–PAGE). The results indicated that Al was occluded by milk proteins (mostly caseins). When citric acid was added to tea infusions the percentage of negatively charged Al-citrate remained either the same or increased to 40% of total Al.     

Aqueous two-phase partitioning of milk proteins

Milk of transgenic pigs secreting recombinant human Protein C (rHPC) was used as a model system to determine the utility of aqueous two-phase extraction systems (ATPS) for the initial step in the purification of proteins from milk. The major challenges in purification of recombinant proteins from milk are removal of casein micelles (that foul processing equipment) and elimination of the host milk proteins from the final product. When milk was partitioned in ATPS composed of polyethylene glycol (PEG) and ammonium sulfate (AS), the phases were clarified and most of the caseins precipitated at the interphase. The partition coefficients of the major milk proteins and rHPC were dependent upon the molecular weight of the PEG used in the ATPS. Higher-partition coefficients of the major whey proteins, Β-lactoglobulin, and α-lactalbumin were observed in ATPS made up of lower molecular-weight PEG (1000 or 1450) as compared to systems using higher molecular-weight PEG. Lowering the pH of the ATPS from 7.5 to 6.0 resulted in increased precipitation of the caseins and decreased their concentration in both phases. rHPC had a partition coefficient of 0.04 in a system composed of AS and PEG 1450. The rHPC in pig milk was shown to be highly heterogenous by two-dimensional gel electrophoresis. The heterogeneity was owing to inefficient proteolytic processing of the single chain to the heterodimeric form and differences in glycosylation and other post-translational processing. Differential partitioning of the multiple forms of purified rHPC in the ATPS was not observed. rHPC after processing in ATPS was recovered in a clear phase free of most major milk proteins. ATPS are useful as the initial processing step in the purification of recombinant proteins from milk because clarification and enrichment is combined in a single step.     

天然抗氧化剂茶多酚

抗氧化剂是指能防止或延缓食品成分氧化变质的食品添加剂.茶多酚是目前尚不能人工合成的纯天然、多功能、高效能的抗氧化剂和自由基净化剂,可用超临界CO2萃取法、离子沉淀提取法、树脂吸附分离法、有机溶剂萃取法提取.采用酒石酸亚铁比色法测定茶多酚含量.     

茶多酚的表面性能研究

通过分别测试二碘甲烷、水、甲酰胺等液体在茶多酚上的接触角,并根据van Oss-Chaudhury-Good的组合理论对茶多酚的表面性能进行研究。结果表明茶多酚的表面能随着茶多酚的含量增加而提高;且随着茶多酚的含量增加,儿茶素类物质即含酚羟基类物质的含量进一步增加,形成氢键的几率进一步增大,导致分子之间作用力和氢键力加大,因而使茶多酚的表面能提高。     

茶业风云——茶多酚

Tea polyphenols are the main active substances in pu-erh tea. They play an important role in anti-tumor, anti-virus, and antioxidant activity, as well as preventing senile cardiovascular and cerebrovascular diseases. In this paper, we, in a hint fiction, introduced the basic properties, physiological functions of tea polyphenols and described the effect of production technology on the content of tea polyphenols.     

Antioxidant activity of polyphenols from green and toasted mate tea

The production and distribution of toasted mate tea in Brazil has increased, which has resulted in its greater consumption. Mate tea is obtained by roasting non-fermented erva-mate in order to produce toasted erva-mate or toasted mate tea. However, although the product is much appreciated, studies of its chemical composition and the concentration of polyphenols, particularly flavonols present in toasted mate tea, are few and often controversial. This paper elucidates some misunderstandings involving the nomenclature of erva-mate and toasted mate, and mainly provides an overview of the composition of polyphenols and antioxidant capacity of toasted mate tea and its raw material, erva-mate, in comparison with other teas, the compositions of which were found in the literature.     

Improved method for the simultaneous determination of whey proteins, caseins and para-kappa-casein in milk and dairy products by capillary electrophoresis

A capillary electrophoresis method for the simultaneous determination of whey proteins, caseins and their degradation products, such as para-kappa-casein, was proposed. The effect of several parameters (pH, ionic strength and concentration of urea in the electrophoresis buffer and applied voltage) on the analysis time and on the separation efficiency of the major milk proteins was studied. Using a hydrophilically coated capillary, in combination with electrophoresis buffer 0.48 M citric acid-13.6 mM citrate-4.8 M urea at pH 2.3, and a separation voltage of 25 kV, a complete separation of beta-lactoglobulin and para-kappa-casein was achieved, permitting the quantification of both components.     

茶叶中的茶多酚荧光检测方法

以石墨烯量子点为荧光探针,基于二氧化锰纳米片与石墨烯量子点之间发生有效的荧光共振能量转移,构建了一种茶叶中茶多酚含量的荧光检测新方法。MnO2纳米片使石墨烯量子点的荧光淬灭,而茶多酚能与MnO2发生氧化还原反应,将其还原成Mn^2+,使体系荧光恢复。茶多酚含量与荧光强度增量在20~750μg/mL范围内成线性相关,线性方程为F=1.0574c-114.9,线性相关系数为R=0.9936。将该方法用于茶叶中茶多酚含量的检测,结果与国标法接近。方法有望应用于食品中其它抗氧化物质的实时检测。     

Modification of milk using immobilized enzymes

The preparation and use of immobilized enzyme systems for the modification of the principle components of milk—casein, lactose, and butterfat—are discussed. Design of a pilot-scale bioreactor is described. Also the use of tea polyphenols as a crosslinking agent is reported. A review of methods for hydrolyzing lactose and characteristics of a concentrated liquid sweetener derived from dairy byproducts is presented. Further, the application of immobilized esterases to modify milk, cream, and butterfat is reported.     

Quantification of proteins in dairy products using an optical biosensor

Two recent techniques using optical immunosensor technology were developed for the quantification of milk proteins in dairy products. The first application is the simultaneous quantification of the 3 major caseins (alpha(s1), beta, and kappa). This assay consists of a 2-step sandwich strategy, with 2 monoclonal antibodies directed against the N- and C-terminal extremities of each of the caseins, respectively. This strategy permits only intact caseins to be quantified, and not their degradation products. The technique is fast (10 min), sensitive (detection limit about 0.87 microg/mL), and has been applied successfully to raw and drinking milks. In the second application, the severity of the heat treatment sustained by a milk of unknown origin is determined by quantifying separately the native and heat-denatured forms of alpha-lactalbumin with specific monoclonal antibodies. The technique allows discrimination of the different heat treatments studied (pasteurization, direct and indirect ultra-high temperature, sterilization), is fast (4 min), repeatable, fully automated, and requires no pretreatment of the milk sample.     

Human urinary metabolite profile of tea polyphenols analyzed by liquid chromatography/electrospray ionization tandem mass spectrometry with data-dependent acquisition

Tea is rich in polyphenols and has a variety of biological activities. In order to better understand the biological effects of tea constituents on human health, markers for their exposure and their metabolic fates are needed. Previously, we have characterized several catechin metabolites in the blood and urine, but more information on the metabolite profile of tea polyphenols is needed. In the present study, the human urinary metabolite profile of tea polyphenols was investigated using liquid chromatography/electrospray ionization tandem mass spectrometry with data-dependent acquisition. With data-dependent MS/MS analysis by collecting the MS2 and MS3 spectra of the most intense ions in the sample, we identified more than twenty metabolites of tea polyphenols from human urine samples. (-)-Epigallocatechin (EGC) glucuronide, methylated EGC glucuronide, methylated EGC sulfate, (-)-epicatechin (EC) glucruronide, EC sulfate, methylated EC sulfate, as well as the glucuronide and sulfate metabolites of the ring-fission metabolites of tea catechins, 5-(3',4',5'-trihydroxyphenyl)-gamma-valerolactone (M4), 5-(3',4'-dihydroxyphenyl)-gamma-valerolactone (M6) and 5-(3',5'-dihydroxyphenyl)-gamma-valerolactone (M6'), were the major human urinary metabolites of tea polyphenols. To our knowledge, this is the first report of the direct simultaneous analysis of the human urinary metabolite profile of tea polyphenols using single sample analysis. This method can also be used for thorough investigations of the metabolite profiles of many other dietary constituents.     

Synthesis, characterization and cytotoxicity studies of chitosan-coated tea polyphenols nanoparticles

Chitosan nanoparticles (CS-NPs) were prepared by ionic gelation method using carboxymethyl chitosan and chitosan hydrochloride as carriers of tea polyphenols. The characteristics of chitosan-coated tea polyphenols nanoparticles (CS-TP NPs) were determined by using transmission electron microscopy (TEM) and FT-IR spectroscopy. It was found that the synthesized CS-TP NPs were non-spherical in shape with an average size of 407±50nm. Meanwhile, the drug content and encapsulation rate of the nanoparticles was 8-16% and 44-83%, respectively. These CS-TP NPs also demonstrated sustained release of tea polyphenols in PBS. The antitumor of CS-TP NPs towards HepG2 cancer cells was investigated. The result showed that CS-TP NPs retained significant antitumor activities.     

Effect of pH and ionic strength on competitive protein adsorption to air/water interfaces in aqueous foams made with mixed milk proteins

Quantitative analysis of competitive milk protein adsorption to air/water interfaces in aqueous foam was performed by capillary electrophoresis (CE). Foams were made by whipping protein solutions, in which skim milk powder (SMP) and whey protein isolate (WPI) were mixed at 0.5% protein in different proportions at different pH values and NaCl concentrations. Preferential adsorption of beta-casein into foam phases occurred under most solution conditions, if partial dissociation of the casein micelles had occurred. Preferential adsorption of beta-casein was not observed with added Ca2+, due to the re-association of casein micelles. Enrichment of caseins into the foam phase was more apparent than that of whey proteins. The foamability of SMP demonstrated a continuous improvement due to the gradually increasing dissociation of casein micelles when the concentration of NaCl increased from 0 to 0.8 M. The foamability of WPI increased when NaCl concentration rose from 0 to 0.1 M, and decreased with further increase in NaCl concentration. NaCl at low concentration (I < or = 0.4) did not show a significant effect on the competitive adsorption among milk proteins, indicating that electrostatic interactions do not play a key role in competitive adsorption. NaCl at higher concentration, e.g., 0.6 M, caused less whey protein to be adsorbed to the air/water interfaces. The whippability of WPI was highest at pH 4.5 and lowest at pH 3, and that of SMP was the opposite. The proportions of beta-lactoglobulin and alpha-lactalbumin in the foam phase were lower at acidic pH and higher at basic pH, compared with that at natural pH of WPI.     

Isolation and Purification of a Novel Long－chain Acyl Catechin from Lipophilic Tea Polyphenols

Llpophilic tea polyphenols (LTP) was prepared by esterification of green tea polyphenols (GTP) with hexadecanoyl chloride. A novel long-chaln acyl catechin was isolated and purified from LTP by high-speed countercurrent chromatography (HSCCC).Its molecular structure was elucidated as epigallocatechin-3-O-gallate-4‘-O-hexadecanate by elemental analysis, IR, MS and ^1H NMR spectra.     

树脂对茶多酚与咖啡因的吸附分离

从低档绿茶末的水提取液中，利用离子交换，吸附树脂成功地分离出茶多酚与咖啡因，并详细考察了其交换与吸附性能。与其它方法相比，该树脂法具有产品收率高，成本低廉等特点。