Tea Polysaccharide Conjugates Research Articles

Effects of Tea-Polysaccharide Conjugates and Metal Ions on Precipitate Formation by Epigallocatechin Gallate and Caffeine, the Key Components of Green Tea Infusion.

The mechanism of tea-infusion precipitation is not clear. Epigallocatechin gallate (EGCG) and caffeine (CAF) are the core components of green tea infusion precipitate. The critical precipitation concentration ( CCPC) of EGCG-CAF mixed aqueous solutions was assayed through a turbidity assay performed at 25 °C and verified by a differential-scanning calorimeter (DSC) for reliability. Different concentrations of the green tea polysaccharide conjugate fraction (gTPC-D), with a molecular weight of 2.32 × 105 Da, and of metal ions (K+, Mg2+, Ca2+, and Al3+) were respectively introduced into the mixed solution to study their effects on CCPC. On the basis of the changed CCPC, the clarification-precipitation phase diagrams of the mixed aqueous solutions were constructed. By calculating the integral area of the clarification phase, it was determined that the four metal ions and gTPC-D could accelerate precipitate formation with a dose-response relationship; among the metal ions, the effect of Al3+ is the strongest, followed by that of Ca2+.

Comparative analysis of physicochemical characteristics of green tea polysaccharide conjugates and its decolored fraction and their effect on HepG2 cell proliferation

During plant polysaccharide extraction, polyphenols are usually oxidatively polymerized to produce quinone pigments adsorbed the macromolecular such as protein and polysaccharides. Whether these derived pigments affect the properties of polysaccharides remains unknown. In this study, green tea polysaccharide conjugates (gTPC) were prepared from low-grade green tea by means of combined-enzyme extraction followed by decoloring with anion-exchange column chromatography to obtain the corresponding fraction (gTPC-D). gTPC comprised two components, with molecular weights of 1.71 × 107 Da and 2.56 × 105 Da in mass proportions of 0.09% and 99.91%, respectively. gTPC-D consisted of one component of molecular weight 2.32 × 105 Da. gTPC UV–vis spectrum presented a shoulder-shaped peak, with an inflection point at 257 nm in the 240–300 nm range, whereas the spectrum of gTPC-D displayed a very weak wave-like curve in the 255–300 nm region associated with quinone pigments rather than their covalently bound protein. Quinone pigments increased the zeta potential of gTPC and reduced its water-holding capacity at high relative humidity (RH) (above 80%). gTPC-D had a significant dose-dependent inhibitory effect on HepG2 cells (p < 0.01) with an IC50 of 500 μg/mL. These results will contribute to the quality control and large-scale production of tea polysaccharide conjugates (TPC).

Analysis of Protein Moiety of Polysaccharide Conjugates Water-extracted from Low Grade Green Tea

Tea polysaccharide conjugates(TPCs) were water-extracted and purified from low-grade green tea. Al-though TPC contains 1.97% covalent binding proteins, Coomassie brilliant blue G250 and R250 could not detect this protein. The "should-shape" absorption peak observed at 250–280 nm in the UV-Vis spectrum was caused by quinone compounds derived from polyphenol oxidation during TPC extraction. The UV-Vis spectrum could be used to detect the decolorizing degree of TPC. TPC aqueous solution merely presented the negative charge properties of its polysaccharides instead of the acid-base property of its protein section, and exhibited higher stability at pH greater than 5.0. No precipitation or haze occurred in three TPC/epigallocatechin gallate(EGCG) aqueous mixtures during 12 h storage. TPC had emulsifying activity, which indicated that its protein moiety formed hydrophobic groups. It is proposed that some of the physicochemical properties of the TPC protein moiety are shielded by its polysaccharide chain.

Some Physical Properties of Protein Moiety of Alkali-Extracted Tea Polysaccharide Conjugates Were Shielded by Its Polysaccharide.

Polysaccharide conjugates were alkali-extracted from green tea (TPC-A). Although it contained 11.80% covalently binding proteins, TPC-A could not bind to the Coomassie Brilliant Blue dyes G250 and R250. TPC-A had no expected characteristic absorption peak of protein in the UV-vis spectrum scanning in the range of 200–700 nm. The UV-vis wavelength of 280 nm was not suitable to detect the presence of the protein portion of TPC-A. The zeta potential of TPC-A merely presented the negative charge properties of polysaccharides instead of the acid–base property of its protein section across the entire pH range. Furthermore, TPC-A was more stable when the pH of solution exceeded 4.0. In addition, no precipitation or haze was generated in the TPC-A/(−)-epigallocatechin gallate (EGCG) mixtures during 12 h storage. TPC-A has emulsifying activity, which indicated that its protein moiety formed hydrophobic groups. Thus, it was proposed that some physical properties of TPC-A protein were shielded by its polysaccharide, since the protein moiety was wrapped by its polysaccharide chains.

Physicochemical characteristics of polysaccharide conjugates prepared from fresh tea leaves and their improving impaired glucose tolerance

Hot-water extracts were prepared from fresh tea leaves and fractionated by DEAE-cellulose DE-52 column chromatography to yield one unexplored polysaccharide-conjugate fraction TPC-L (tea polysaccharide conjugates). Chemical components, molecular weight and its distribution, water vapor sorption properties, zeta potentials and optical characteristics of TPC-L were investigated. As compared with injured cell group, the two dosages of TPC-L (150 and 300μg/mL) were discovered to possess remarkably protective effect on human umbilical vein endothelial cells against impairments induced by high glucose in a dose-dependent manner (p<0.05, p<0.001, respectively). Compared with group NC (normal control), the ingestion of 40mg/kg of TPC-L could significantly reduce blood glucose levels of normal mice ingesting starch, and significant difference of AUC (area under the curve of blood glucose) and ΔAUC (p<0.05, p<0.01) at the postprandial time point of 0.5 and 1.0h were observed. The three dosages of TPC-L (10, 40 and 160mg/kg) did not significantly lower postprandial blood glucose levels of normal mice ingesting glucose. TPC-L could improve starch tolerance to prevent impaired glucose tolerance (IGT) from developing into diabetes as well as protective effects on HUVE cells against impairments induced by high glucose It was suggested that TPC-L improved IGT through its capability of inhibition on digestive enzymes.

Thermal Treatment on Tea Polysaccharide Conjugates Prepared from Green Tea by Extraction of Decoction and Complex Enzyme

Tea polysaccharide conjugates(TPC) named TPC-1 were prepared from low-grade green tea by extraction through complex enzymes method and purified by diethylaminoethyl(DEAE) cellulose(DE-52) column chromatography,while TPC-2 was prepared by extraction with boiling water and purified by Sephadex G-200 column chromatography.TPC-1 and TPC-2 were incubated respectively at 98 ℃ for 5.0 h.A single symmetrical peak corresponding to the main homogeneous component,TPC-1a in the chromatogram of high performance gel permeation chromatography(HPGPC) was split into two adjacent peaks representing two different components of TPC-1a-1 and TPC-1a-2.Circular dichroism(CD) spectra showed an additional positive Cotton effect for the peak at 216 nm along with the peak at 194 nm.HPGPC showed that incubation in water at 98 ℃ for 5.0 h has no effects on TPC-2 that contained two homogeneous components of TPC-2a and TPC-2b.The CD spectra of the treated sample revealed that the positive Cotton effects for peaks at 203 and 215 nm disappeared,while it retained for the one at 272 nm.The solution conformation and homogeneous component distribution of TPC-1 had been changed by heat treatment.But only solution conformation had been changed for TPC-2.

Suppression of diabetes in non-obese diabetic (NOD) mice by oral administration of water-soluble and alkali-soluble polysaccharide conjugates prepared from green tea

Water-soluble tea polysaccharide conjugates (TPC-W) and alkali-soluble tea polysaccharide conjugates (TPC-A) were extracted from green tea by hot and alkali water respectively. Physicochemical properties of TPC-W and TPC-A were analyzed. Non-obese diabetic (NOD) mice were used to evaluate antidiabetic bioactivities of TPC-W and TPC-A. The daily oral administration of 150 mg kg −1 TPC-W can significantly decrease the level of blood glucose in NOD mice. The anti-glutamic acid decarboxylase (GAD) antibody level in NOD mice treated with 150 mg kg −1 TPC-W decreased 27% ( P < 0.05). To the end of trial, only 2 out of 10 mice in NOD groups treated with TPC-W or TPC-A exhibited diabetic symptoms compared with model control group, in which 7 of 10 mice developed diabetes. The result of organ index showed that both TPC-W and TPC-A can protect thymus from shriveling to some extent. In sum, our studies demonstrated that both TPC-W and TPC-A can suppress spontaneous diabetes mellitus in NOD mice.

Thermal Effects on the Stability and Antioxidant Activity of an Acid Polysaccharide Conjugate Derived from Green Tea

A technique of high-performance gel permeation chromatography (HPGPC)-evaporative light-scattering detection and circular dichroism (CD) was developed for the measurement of thermal effects on the homogeneity and conformation of polymeric carbohydrate conjugates and was applied to an acid polysaccharide conjugate (GTa) isolated from the composite enzyme extract of green tea. Incubations in water at 40 and 70 degrees C for 1.0, 2.5, and 5.0 h have no effects on GTa. In contrast, when incubated in water for 1.0, 2.5, and 5.0 h at 98 degrees C, a single symmetrical peak corresponding to GTa in HPGPC was split into two adjacent peaks representing two different components formed, and CD spectra revealed an additional positive Cotton effect at 216 nm. To contribute toward our understanding of thermal effects of this polymeric carbohydrate conjugate on antioxidant activity, GTa and related heat-treated samples (GTa-HTI, GTa-HTII, and GTa-HTIII), the latter being obtained from 1.0, 2.5, and 5.0 h incubations at 98 degrees C, respectively, were subjected to the self-oxidation of 1,2,3-phentriol assay and found to have respective scavenging activities in a concentration-dependent manner. In comparison with GTa, the scavenging potency of heat-treated samples was similar at the dosage range of 50-300 microg/mL but became stronger with continually increasing concentration. Moreover, the present study also provides further insights into the optimal preparation of tea polysaccharide conjugates.

Antioxidant activities of different fractions of polysaccharide conjugates from green tea ( Camellia Sinensis)

Three fractions of water-soluble polysaccharide conjugates, coded as TPC-1, TPC-2, and TPC-3, were isolated and purified from low-grade green tea ( Camellia sinensis) by absorbent chromatography and ion exchange chromatography. Their chemical and physical characteristics were determined by chemical methods, gas chromatography and size exclusion chromatography with laser light scattering. Deoxyribose assay, photoreduction of Nitro Blue Tetrazolium (NBT) assay and lipid peroxidation inhibition assay were applied to test the antioxidant activities of tea polysaccharide conjugates in vitro. The results indicated that the three polysaccharide conjugates were heteropolysaccharides bounded with protein. TPC-1 was composed of l-arabinose (Ara), d-ribose (Rib), d-xylose (Xyl), d-glucose (Glc), d-galactose (Gal) and d-mannose (Man). TPC-2 was only composed of four monosaccharides: Ara, Rib, Glc and Man. There was no Man detected in TPC-3. The protein contents of TPC-1, TPC-2, and TPC-3 were 2.8%, 3.8% and 4.0% and the molecular weights were 26.8 × 10 4, 11.8 × 10 4, 4.2 × 10 4, respectively. TPC-3 showed the highest antioxidant activities among the three fractions of polysaccharide conjugates, with an IC 50 of 182 μg/ml for the deoxyribose assay, and 93 μg/ml for the photoreduction of NBT assay, values which were lower than those of TPC-1 and TPC-2 ( P < 0.01). The effects of the molecular weight and protein content of the polysaccharide conjugates on the improvement of the bioactivities appeared to be significant.

Quantification of uronic acids in tea polysaccharide conjugates and their antioxidant properties.

A technique of high-performance liquid chromatography (HPLC) was described for the measurement of total uronic acids in tea polysaccharide conjugates. This method was applied to polysaccharide conjugate extracts obtained from green tea after most of the components that produce interference were removed. The preliminary extraction process was according to the procedure of isolation of polysaccharide conjugates. The uronic acid content of different polysaccharide conjugate fractions was quantified by HPLC on a Sugar-Pak I column with a 1.0 x 10(-)(4) mol x L(-)(1) calcium disodium ethylenediaminetetraacetic acid solution as the mobile phase and refractive index detection. The validation study showed high recoveries (>97.0%) and low coefficients of variance (<3.0%). The minimum detectable limit concentration of uronic acid was 10 microg x mL(-)(1). The analysis of a standard range of galacturonic acid concentrations (100-4000 microg x mL(-)(1)) yielded linear results. The use of the method on different polysaccharide conjugate fraction samples confirmed its effectiveness. With the high content of uronic acids in polysaccharide conjugates, the stronger reactive oxygen species scavenging activities were found.