Non-fermented Tea Research Articles

Quality Characteristics and Content of Polysaccharides in Green Tea Fermented by Monascus pilosus

In this study, we designed a method to manufacture elevated fermented green tea by using Monascus pilosus, which is known as a functional microbe, and observe its antioxidant abilities and quality characteristics. The water-soluble substance (WSS) content of the fermented tea by M. pilosus (FTM) was lower than that of the non-fermented tea (NFT), although the alcohol-insoluble substance (AIS) content of the FTM was higher than that of NTM. On the other hand, the fractionated distilled water-soluble polysaccharide (DWSP), CDTA-soluble polysaccharides (CDSP), sodium carbonate-soluble polysaccharide (SCSP) and KOH soluble hemicellulose (HC) obtained from the AIS of the FTM was markedly higher than that of NFT. In the antioxidant parameters, the electron donating ability of all fractions, except HC, extracted from FTM was higher than that of NFT, and iron chelating ability of all fractions, except CDSP, extracted from FTM was higher than that of NFT. Whereas the DWSP and SCSP obtained from the FTM were higher than that of NFT, the activity of the HC fraction from both NFT and the FTM could not be detected. In addition, the xanthin oxidase (XO) inhibitory activities of the DWSP, CDSP and the SCSP obtained from the NFT were significantly higher than that of FTM, the aldehyde oxidase (AO) inhibitory activities of the DWSP and SCSP extracted from the FTM were markedly higher than that of the NFT. Meanwhile, the acceptance of NFT and FTM had no significant difference, while the quality of aroma, taste and mouthfeel of the FTM was higher than that of NFT. These results suggest that the post-fermented tea by Monascus microorgan-isms may be responsible for functional components as well as contribute to the improvement of the tea quality.

균주에 따른 청미래덩굴잎 발효차의 항산화 활성

* 명지대학교 농생명바이오식의약소재개발사업단, ** 명지대학교 생명과학정보학부 Abstract To evaluate the functional characteristic and availability for drinking of the fermented Smilax china leaf tea by using different microbial species, various fermented leaf tea was prepared by non-fermentation (C), or the fermentation of Saccharomyces cerevisiae (S), Bacillus sp. (B), Bifidobacterium bifidus (L), Monascus pilosus (M) and Aspergilus oryzae (A), and sensory and antioxidant parameter of each brewed tea was observed. The color of the A tea was red, but the other teas were yellow in color. Furthermore, the aesthetic quality of the A and M tea was 3.95 and 3.30 point, respectively, and other teas (2.55~ 2.28) were similar to that of the C tea. TP of fermented tea water extract was lower than that of the C, although TF was not significantly different between the fermented and non-fermented tea. Especially, TF of the A tea was significantly lower than those of the other teas. The range of EDA (1 ㎎/㎖) of water and ethanol extracts of tea C and the fermented teas was 19.25~22.48%; however, tea A was only 8.04~12.49%. In addition, FRAP, FICA and LPOIA of teas were not significantly different between the fermented and non-fermented teas. On the other hand, XOIA and AOIA of tea ethanol extracts were slightly higher than those of water extracts. XOIA of water extract derived from the teas was 4.83~9.20%, while ethanol extract of these was 9.00~19.00%. However, XOIA of B and L teas water extract was not detected. Furthermore, AOIA of fermented tea water extract (30.17~48.52%) were lower than those of ethanol extract (44.09~66.93%). In this study, interestingly, antioxidant parameters, such as FRAP, FICA, LPOIA and AOIA, of the A tea water extract (0.1%) was higher than that of the other tea in spite of high decreasing rate in the contents of TP and TF. Therefore, above results imply the possibility of fermented Smilax china leaf tea as a functional food.

Analysis of metabolic markers of tea origin by UHPLC and high resolution mass spectrometry

Tea is an infusion made from the dried leaves of Camellia sinensis L. and is the second most consumed beverage in the world. It has been shown that factors such as fermentation methods, cultivar, geographical origin and season can affect the biochemical composition of tea. In this study, the biochemical composition of green, oolong and black commercial tea samples from around the world was studied using a non-targeted method utilising reversed phase ultra high pressure liquid chromatography (UHPLC) and high resolution mass spectrometry. Principal component analysis of green, oolong and black tea extracts clearly showed that fermented tea can be resolved from non-fermented tea. When the non-targeted data were combined with the supervised multivariate technique, partial least squares discriminant analysis, the method was able to clearly distinguish ‘country of origin’ within green tea and to a lesser extent within a black tea sample set, plus provide indicative marker ions for the country of origin. Many of the significant components detected in this study are unknowns, emphasising the importance of un-biased non-targeted analytical techniques. This study highlights the potential efficacy of non-targeted UHPLC–mass spectrometry when combined with multivariate statistics to differentiate fermented from non-fermented tea and provide potential indicators of provenance of tea samples for further examination.

Comparison of Azorean tea theanine to teas from other origins by HPLC/DAD/FD. Effect of fermentation, drying temperature, drying time and shoot maturity

The objectives of the present study were the determination of theanine (a specific amino acid of tea leaves with repercussion in human neuronal activity) by HPLC with o-phthaldialdehyde derivatisation prior analysis and comparison of its level in samples from different origins or from the same origin (Azores Islands) but with different processing conditions (fermentation, drying temperature and drying time) and shoot maturity. The results revealed a higher level of theanine in the Azorean green tea (3.10%), the only one produced in Europe, as compared with the Chinese, Japanese, Korean and Taiwan green teas that have an average theanine content of 1.52%, 1.34%, 0.89% and 2.76% of the dry weight (dw) of tea leaves, respectively. The fermentation for a longer period of time results in less theanine remaining (2.28% and 3.10% of dw for fermented and non-fermented tea, respectively). The drying temperature and time of the drying process have an inversely strong effect (4.45% and 5.65% of dw for dried at 55°C during 8.50h and at 40°C during 7.10h, respectively). The younger tea shoots have a higher theanine content than mature leaves (7.02% and 6.10% of dw, respectively). Our data suggested that a new type of Azorean theanine-rich tea may be produced by modifying the manufacturing processes of the younger tea shoots.

ANTIBACTERIAL ACTIVITY OF BLACK TEA AGAINST STREPTOCOCCUS MUTANS AND ITS SYNERGISM WITH ANTIBIOTICS

Tea is the most common beverage after it is extracted from the leaves of Camellia sinenesis. It is classified in to fermented tea (Black Tea) and non-fermented tea (green tea). Methanolic extract of tea diluted in methanol and ethanol, which were tested for antibacterial activity against Streptococcus mutans. Methanol was found to be best antimicrobial solvent. The MIC value of methanolic extracts of Black Tea diluted in ethanol was 0.1 mg/ml. Synergistic results of antibacterial activity of methanolic extracts (diluted in ethanol) were found poor. Synergistic activity of Black tea sample with antibiotics (Chloramphenicol, Tetracycline, Levofloxacin and Gentamycin) showed best response against most of the bacteria (0.1mg/ml).

Comparison of the antioxidant capacity and the antimicrobial activity of black and green tea

Tea is a common beverage. The green tea is preferentially recommended for its strong antioxidant properties and also for its antimicrobial activity. The antioxidant capacities of 30 samples (black and green tea) were determined by the chemiluminescent Trolox equivalent antioxidant capacity (TEAC) determination. The average value of TEAC of the non-fermented (green tea) and semi-fermented tea samples was 1.43 mM and the average value of TEAC of the fermented teas (black tea) samples was 1.43 mM. All samples were stored in freezer (−20 °C) and the TEAC determination was repeated after a year. The average values of TEAC of non-fermented and semi-fermented tea samples were twofold lower in comparison to fermented tea samples and only 20% of average value of TEAC of the fresh tea infusion. The parallel determinations of minimal inhibitory concentration (MIC) on gram-positive and gram-negative bacterial strains (Enterococcus faecalis, Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli) were done. Also the MIC was possible to determine after a year. The assumed prevailing antioxidant and antimicrobial activities of non-fermented tea infusions were not confirmed as well as the dominant antioxidant and antimicrobial properties of specific type of tea infusion.

Extract of black tea (pu‐ehr) inhibits postprandial rise in serum cholesterol in mice, and with long term use reduces serum cholesterol and low density lipoprotein levels and renal fat weight in rats

A water-soluble extract of a traditional Chinese fermented black tea, pu-ehr, decomposes bile acid cholesterol micelles. This black tea extract (BTE) was studied to see if it could decrease the postprandial elevation of blood cholesterol levels after a single administration in ddY mice. It was found that BTE (0.3 g/kg) significantly decreased the postprandial rise in blood cholesterol levels after oral administration of cholesterol (130 mg/kg). A non-fermented tea (i.e. green tea) extract did not prevent the postprandial increase in blood cholesterol. In a subsequent study, 5-week-old Sprague-Dawley (SD) rats were fed BTE for 3 weeks, following which a dose-dependent and significant decrease in serum total cholesterol levels (1.36 mmol/L, 0.1% BTE, p < 0.05) was found and also in renal fat weight (0.3% BTE, p < 0.05). LDL cholesterol levels (0.51 mmol/L, 0.1% BTE, p < 0.05) were also significantly decreased. There were no significant changes in the weights of other organs or in the serum levels of other clinical markers. Thus, BTE has a specific antihypercholesterol effect in rodents, which might potentially aid in the management of hyperlipidaemia in man.

발효정도가 다양한 녹차의 첨가가 유과의 저장 중 품질 특성에 미치는 영향

Yukwa were added with non-fermented tea (Bucho-cha and Okro-cha), semi-fermented tea (Ooreung-cha), and fermented tea (black tea) to investigate the effect of different kinds of tea powder on quality characteristics of Yukwa during storage. Yukwa samples were used for analysis such as crude lipid, moisture, texture, antioxidative property (TBA value), and sensory test. There were no significant differences on crude lipid, moisture, and texture analysis. Green tea powder treatment showed strong antioxidant activity. In particular, antioxidant property of non-fermented tea exhibited a higher antioxidative effect than that of the other samples. Sensory evaluation showed that sensory scores of samples added with non-fermented tea were significantly higher than the other samples, indicating that the addition with 0.1% of non-fermented tea powder could contribute to the improvement of quality and shelf-life of Yukwa during storage.

Effects of non-fermented tea extracts on in vitro digestive hydrolysis of lipids and on cholesterol precipitation

Extracts of non-fermented teas precipitated cholesterol and decreased dialysability of lipids in an in vitro model that simulated digestion in humans. The efficiency of tea extracts was affected by catechins and mainly by EGCG, ECG, GCG and CG. Total amounts and concentrations of individual catechins in extracts depended strongly on the tea brands. There were significant relationships between amounts of dialysable products of triglyceride hydrolysis, amounts of precipitated cholesterol, and concentrations of individual catechin gallates in tea brews. Dialysability of lipids was effectively reduced by tea extracts high in GCG and CG. Dialysability of lipids was strongly correlated with the sum of GCG and CG (R2 = 84.64%) whereas precipitation of cholesterol was highly correlated with total amounts of catechin gallates in the tea extracts (R2 = 89.59%).

Effect of brewing temperature and duration on green tea catechin solubilization: Basis for production of EGC and EGCG-enriched fractions

In an industrial context of producing catechin-enriched fractions by electromigration, a new technology demonstrated to be effective for concentration of the two main catechins (EGC and EGCG) of green tea, the recovery yield from tea leaves during brewing would be the most important parameter of the whole process rentability. However, the majority of the kinetic studies were carried-out on black tea, a fermented tea. Consequently, the objective of this study was to investigate the effects of temperature (50, 60, 70, 80 and 90 °C) and brewing duration (0, 5, 10, 20, 40 and 80 min) on the catechin solubilization from green tea, a non-fermented tea. The use of mathematical models revealed that there was a variable interdependence between the brewing duration and the brewing temperature on catechin and caffeine concentrations. It was possible to divide catechins in two groups, the time dependent compounds (EGC and EC) and the time/temperature dependent compounds (C, EGCG, GCG and ECG). Furthermore, the 3D models calculated to represent the evolution of the catechins and caffeine concentrations allowed to determine the best combination of time and temperature for their extraction: 50 °C during 20–40 min for time-dependent compounds, 90 °C during 80 min for the time/temperature-dependent compounds, and 70–80 °C during 20–40 min for caffeine. Furthermore, this research pointed out a very simple two-step procedure to fractionate the EGC and EGCG by modifying brewing temperature and time parameters.

Study of catechin and xanthine tea profiles as geographical tracers.

The contents of gallic acid, epigallocatechin gallate, epigallocatechin, epicatechin, epicatechin gallate, catechin, caffeine, theophylline, and theobromine were determined in a set of 45 tea samples, including fermented (black and red) and nonfermented (green) teas of different geographical origins (i.e., China, Japan, Kenya, Sri Lanka, and India). A reversed-phase high performance liquid chromatographic method with gradient elution and photometric detection at 275 nm was used to carry out the analysis. Before the HPLC determination, an extraction step was developed using a mixture of acetonitrile and water (60:40, v/v). Pattern recognition techniques involving principal component analysis (PCA) and linear discriminant analysis (LDA) were applied to differentiate the tea samples according to their geographical origins. Catechins, gallic acid, and tea alkaloids are adequate chemical descriptors to distinguish between fermented and nonfermented tea samples cultivated in different geographical areas.

Catechins are not major components responsible for anti-genotoxic effects of tea extracts against nitroarenes

The anti-genotoxic properties of tea leaf extracts were examined in a Salmonella umu-test. Seven non-fermented teas (green tea), one semi-fermented tea (oolong tea), two fermented teas (black tea and Chinese pu er tea) and two other teas were examined for their anti-genotoxic abilities and for their catechins contents. This was to study the relationship between catechins contents and anti-genotoxic activity of various tea leaf extracts. All types of tea extracts showed more potent suppressive effects against umu gene expression of the SOS response in Salmonella typhimurium TA1535/pSK 1002 induced by four nitroarenes (1-nitropyrene, 2-nitrofluorene, 3-nitrofluoranthene and a mixture of 1,6- and 1,8-dinitropyrene) rather than 4-NQO, AF-2 and MNNG. The anti-genotoxic effect of 12 tea leaf extracts on 1-NP, 2-NF, 3-NF and DNP decreased in the order: oolong tea (semi-fermented tea)>black tea (fermented tea)>sencha (non-fermented tea, an ordinary grade green tea)>tocyucya (other tea)>Chinese pu er tea (fermented tea). The amount of catechins (EGC, C, EGCG, EC and ECG) in various teas in decreasing order was non-fermented tea> semi-fermented tea > fermented tea > other tea. A remarkable feature was the effectiveness of black tea and Chinese pu er tea in suppressing the genotoxicity induced by nitroarenes, in spite of the fact that these fermented teas do not have high catechins contents. Statistical analysis showed that no significant ( P<0.01) correlation was found between the anti-genotoxicity of tea extracts against nitroarenes and the catechins contents in tea leaf extracts. In further experiment, fractionation of sencha extract by HPLC revealed that anti-genotoxicity of the peak fraction corresponding to catechins accounted for <10% of the total anti-genotoxic activity of sencha extract against for 1-nitropyrene. These results suggest that catechins are not major components responsible for the anti-genotoxic effects of tea leaf extracts against direct-acting nitroarenes.

The behaviour of leached aluminium in tea infusions

The behavior of leached aluminium (Al) was studied using GF-AAS with different sample pretreatments and RP-HPLC with lumogallion as the precolumn reagent. The results showed that Al species in tea infusion could be categorized into three groups: large organic compounds, small stable organic compounds and free form Al. The compositions of Al species in a tea infusion would vary with the method of the tea production. For non-fermented teas such as Long-Jin and Moli-Hua, most of the leached Al in the first tea infusion was in large organic compounds and small stable organic compounds, and free form Al consisting of less than 10%. In the case of semi-fermented and fully fermented teas such as Shuihsien and Pu-Erh, the main compositions of Al species in the first infusion were free form Al and small stable organic compounds, while Al in large organic compounds made up about 14% and 4%, respectively. When 1-g tea leaves were infused with 100 ml boiling water, about 21% Al in the tea leaves was leached into the water in the first infusion and a total of about 27% of the Al in the tea leaves was leached into the water in four infusions.

Antioxidant Activity of Various Tea Extracts in Relation to Their Antimutagenicity

The relationship between antioxidant activity and antimutagenicity of various tea extracts (green tea, pouchong tea, oolong tea, and black tea) was investigated. All tea extracts exhibited markedly antioxidant activity and reducing power, especially oolong tea, which inhibited 73.6% peroxidation of linoleic acid. Tea extracts exhibited a 65-75% scavenging effect on superoxide at a dose of 1 mg and 30 - 60% scavenging effect on hydrogen peroxide at a dose of 400 microgram. They scavenged 100% hydroxyl radical at a dosage of 4 mg except the black tea. Tea extracts also showed 50 - 70% scavenging effect on alpha, alpha-diphenyl-beta-picrylhydrazyl radical. The antioxidant activity and the scavenging effects on active oxygen decreased in the order semifermented tea > nonfermented tea > fermented tea. Tea extracts showed strong antimutagenic action against five indirect mutagens, i.e., AFB1, Trp-P-1, Glu-P-1, B[a]P, and IQ, especially oolong and pouchong teas. The antioxidant effect of tea extracts was well correlated to their antimutagenicity in some cases but varied with the mutagen and antioxidative properties.

Comparison of Antimutagenic Effect of Various Tea Extracts (Green, Oolong, Pouchong, and Black Tea)

The antimutagenic effects of various tea extracts prepared from nonfermented tea (green tea), semifermented tea (oolong tea and pouchong tea), and fermented tea (black tea) were investigated by Salmonella/microsome assay. No mutagenicity or toxicity in Salmonella typhimurium TA98 and TA100 was observed with any tea extract. The tea extracts markedly inhibited the mutagenicity of 2-amino-3-methylimidazo(4,5-f)quinoline, 3-amino-1,4-dimethyl-5H-pyridol(4,3-b)indole,2-amino-6-methyldipyrido(l,2-a:3′,2′-d)imidazole, benzo[a]pyrene, and aflatoxin B1 toward S. typhimurium TA98 and TA100 in the presence of S9 mixture, especially those of oolong and pouchong teas inhibited over 90% mutagenicity of these five mutagens at the dosage of 1 mg per plate. Among four tea extracts, black tea exhibited the weakest inhibitory effect on mutagenicity of these five mutagens. The mutagenicity of 4-nitroquinoline-N-oxide, a direct mutagen, was not inhibited by black and oolong tea extracts to S. typhimurium TA98 in the absence of S9 mixture but was increased by the tea extracts at the dose of 1 mg per plate to S. typhimurium TA100. As the antimutagenic effect of semifermented tea was stronger than nonfermented and fermented teas, some antimutagenic substances might be formed during manufacturing processes of tea.

Enhancement of mutagenicity of N-nitrosodimethylamine (NDMA) by a modified microsomal/Salmonella pre-incubation assay using acetone and fasting-inducible S9 from rat liver

The anti-genotoxic properties of tea leaf extracts were examined in a Salmonella umu-test. Seven non-fermented teas (green tea), one semi-fermented tea (oolong tea), two fermented teas (black tea and Chinese pu er tea) and two other teas were examined for their anti-genotoxic abilities and for their catechins contents. This was to study the relationship between catechins contents and anti-genotoxic activity of various tea leaf extracts. All types of tea extracts showed more potent suppressive effects against umu gene expression of the SOS response in Salmonella typhimurium TA1535/pSK 1002 induced by four nitroarenes (1-nitropyrene, 2-nitrofluorene, 3-nitrofluoranthene and a mixture of 1,6- and 1,8-dinitropyrene) rather than 4-NQO, AF-2 and MNNG. The anti-genotoxic effect of 12 tea leaf extracts on 1-NP, 2-NF, 3-NF and DNP decreased in the order: oolong tea (semi-fermented tea)>black tea (fermented tea)>sencha (non-fermented tea, an ordinary grade green tea)>tocyucya (other tea)>Chinese pu er tea (fermented tea). The amount of catechins (EGC, C, EGCG, EC and ECG) in various teas in decreasing order was non-fermented tea>semi-fermentedtea>fermentedtea>other tea. A remarkable feature was the effectiveness of black tea and Chinese pu er tea in suppressing the genotoxicity induced by nitroarenes, in spite of the fact that these fermented teas do not have high catechins contents. Statistical analysis showed that no significant (P<0.01) correlation was found between the anti-genotoxicity of tea extracts against nitroarenes and the catechins contents in tea leaf extracts. In further experiment, fractionation of sencha extract by HPLC revealed that anti-genotoxicity of the peak fraction corresponding to catechins accounted for <10% of the total anti-genotoxic activity of sencha extract against for 1-nitropyrene. These results suggest that catechins are not major components responsible for the anti-genotoxic effects of tea leaf extracts against direct-acting nitroarenes.