Tannin Assay Research Articles

Antibacterial Activity of Dendrophthoe pentandra (L.) Leaves Extract Against Staphylococcus aureus and Escherichia coli

Sapuran, Wonosobo is a popular area with several local tea plantations, which are infested by mistletoe, such as Dendrophthoe pentandra (L.) Miq. Mistletoe has been widely reported to have antibacterial activity due to its secondary metabolite content. Therefore, this study aimed to identify and explore the antibacterial activity of D. pentandra leaves extract against Staphylococcus aureus and Escherichia coli. The test samples were extracted using the maceration method, followed by phytochemical screening and total assay of flavonoids, phenols, and tannins. Antibacterial activity tests were carried out using disk diffusion, Minimum Inhibitory Concentration (MIC), and Minimum Bactericidal Concentration (MBC). The yield from the maceration method using 96% ethanol solvent was 8.78%, containing secondary metabolite compounds, including alkaloids, saponins, steroids, terpenoids, and flavonoids. The total flavonoid, phenol, and tanin content obtained were 4.09 ± 0.70 mg QE/g, 1.76 ± 0.16 g GAE/g, and 1.37 ± 0.14 mg TAE/g, respectively. In addition, the extract showed medium inhibition of S. aureus at a concentration of 400 and 525 mg/m, with no inhibitory effect against E. coli. The MIC and MBC from the microdilution method for S. aureus were at 4400 and 8800 mg/mL extract, while values of 28000 and 56000 mg/mL were obtained for E. coli. Based on these results, the inhibition activity of ethanol extract of D. pentandra against S. aureus was greater compared to E. coli.

Colour properties and tannin concentrations of polymeric phenolic materials extracted from Pinot Noir wines of a single NZ producer

SummaryPolymeric phenolic material (PPM) was separated from monomeric anthocyanins by removing the latter using isoamyl alcohol in 22 Pinot Noir wines of different vintages and grades. The colour and tannin concentrations of PPM were determined using a modified Somers assay and methylcellulose precipitation (MCP) tannin assay, respectively. With wine age increasing, the ratio of absorbance at 520 nm at pH 1.0 to that at 3.4 decreased from 2.6 to 1.1. This decreasing trend with wine age is reported for the first time in Pinot Noir wines. The presence of bleachable polymeric pigments was also observed in PPM, and the degree of ionisation of these bleachable pigments increased along with wine age. Statistical analysis demonstrated the dominant effect of age on colour properties of PPM, regardless of vintage and grade. In contrast, total phenolics and tannin concentration mainly related to vintage, rather than age.

Optimization of the Extraction of Proanthocyanidins from Grape Seeds Using Ultrasonication-Assisted Aqueous Ethanol and Evaluation of Anti-Steatosis Activity In Vitro.

Conventional extraction methods of proanthocyanidins (PAC) are based on toxic organic solvents, which can raise concerns about the use of extracts in supplemented food and nutraceuticals. Thus, a PAC extraction method was developed for grape seeds (GS) and grape seed powder using food-grade ethanol by optimizing the extraction conditions to generate the maximum yield of PAC. Extraction parameters, % ethanol, solvent: solid (s:s) ratio, sonication time, and temperature were optimized by the central composite design of the response surface method. The yields of PAC under different extraction conditions were quantified by the methylcellulose precipitable tannin assay. The final optimum conditions were 47% ethanol, 10:1 s:s ratio (v:w), 53 min sonication time, and 60 °C extraction temperature. High-performance liquid chromatography analysis revealed the presence of catechin, procyanidin B2, oligomeric and polymeric PAC in the grape seed-proanthocyanidin extracts (GS-PAC). GS-PAC significantly reduced reactive oxygen species and lipid accumulation in the palmitic-acid-induced mouse hepatocytes (AML12) model of steatosis. About 50% of the PAC of the GS was found to be retained in the by-product of wine fermentation. Therefore, the developed ethanol-based extraction method is suitable to produce PAC-rich functional ingredients from grape by-products to be used in supplemented food and nutraceuticals.

Tannin Concentration of Gyrinops Tea Taken Form Different Agarwood Plantation and Different Processing Method

Tannin is one of the essential components that determine the quality of herbal tea products. This research aims to conduct a quantitative tannin assay of Gyrinops tea with different leaf sampling locations and processing methods. Factorial Completely Randomized Design was used as an experimental design with two treatment factors. The first factor is sampling location (Kekait, Lingsar, and Pejaring). The second factor is the processing method (Fresh leaves and Dried leaves). A titrimetric method with KMnO4 and Indigo carmine reagent was used for quantitative tannin measurement. ANOVA and DMRT at a 5% significant level were used for tannin concentration analysis. The result indicated that the first, second, and interaction factors significantly affected tannin concentration. Drying the G. versteegii leaves could increase the tannin concentration of the Gyrinops tea product. G. versteegii leaves taken from Kekait have shown highest tannin concentration among other regions. It could be concluded that dried G. versteegii leaves Kekait could produce Gyrinops tea with high quality based on tannin concentration measurement.

Development and Validation of Dimeric Macrocyclic Tannin Assay Method in Dosage Forms

Introduction. Quantitative assessment of the active substance is necessary and perhaps the most significant part of the drug quality control. Validation of the analytical methods of quantitative assessment ensures their compliance with high requirements. The present study describes the development and validation of a spectrophotometry method for the quantitative evaluation of the active substance in the drug form of the national antitumor and antiangiogenic drug «Dimeric macrocyclic tannin (DMT) lyophilizate for solution for injection, 100 mg».Aim. The development and validation of the assay method for the standardization of «DMT lyophilizate for solution for injection, 100 mg».Materials and metods. The study used «DMT lyophilizate for solution for injection, 100 mg» and the active substance DMT. Method – spectrophotometry.Results and discussion. The methodof the quantitative assessment of the active substance in the DMT lyophilized drug by direct UV spectrophotometry was developed and the validation characteristics of the method were defined as a result of the study.Conclusion. The validation results showed that the assay method of DMT in the drug form has the appropriate accuracy, precision and linearity. The obtained results correspond to the approved criteria that allow the use of the developed methodology for evaluating the quality of the drug.

Antimicrobial and radical scavenging activities of Moroccan Ziziphus lotus L. seeds

Natural plant extracts contain a variety of phenolic contents, which are associated to various biological activities. In this study, we evaluated the antioxidant and antimicrobial activities of organic (ethanol and methanol) and aqueous extracts prepared from Ziziphus lotus L seeds. The total polyphenol content of the extracts was determined using the Folin-Ciocalteu reagent, it was in the range of 50.67 ± 1.44 (Ethanolic Extract), 39.32 ± 1.44 (Methanolic Extract) and 23.54 ± 0,44 (Aqueous Extract) mg gallic acid equivalent/g DW. The content in flavonoids was estimated at 69.19 ± 0.10 (Ethanolic Extract), 53.13 ± 0.55 (Methanolic Extract) and 9.63 ± 0.88 (Aqueous Extract) mg equivalent quercitin/g DW. The condensed tannin assay revealed that the methanol extract was rich on tannin (9.12 ± 1.07 mg/g) relatively to the ethanol and aqueous extracts (4.97 ± 0.95 and 1, 88 ± 0.47 mg/g respectively). The antioxidant activity was evaluated in vitro by DPPH and phosphomolydbate (total antioxidant capacity). The results reveal that the three extracts have a capacity to trap the DPPH radical with IC50 1.33 ± 0.01, 1.32 ± 0.09 and 3.11 ± 0.05 mg/ml for the methanol, ethanol and aqueous extract respectively. This antioxidant activity is confirmed by the phosphomolybdate test. The antimicrobial activity of the studied extracts was evaluated using the broth microdilution, on five microbial strains: Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Enterococcus faecalis and Candida tropicalis. Results revealed an activity on the four bacterial strains tested. While the fungal strain (Candida tropicalis) showed an amount of resistance to the tested extracts. All this results could justify the use of Ziziphus lotus in the treatment of various infections in traditional environments.

Antidiabetic Activity of leaf extract from three types of Mangrove Originating from Sambera Coastal Region Indonesia

This study designed to determine the antidiabetic profile ethanol extracts of leaf extract from three types of mangroves; Avicennia sp., Rhizopora sp, and Sonneratia sp. Extraction process used maceration with ethanol solvent. The phytochemical assayis carried out qualitatively including assay of alkaloids, flavonoids, steroids, terpenoids, phenolic compounds, tannins, and saponins. The method used for the antidiabetic test is the method of blood glucose tolerance. The results showed that ethanol extract of three types of mangrove leaves Avicennia sp., Rhizopora sp, and Sonneratia sp. containing secondary metabolite group alkaloids, flavonoids, steroids, terpenoids, phenolic compounds, tannins, and saponins. Ethanol extract of leaves of the three types of mangroves plants has antidiabetic activity. The ethanol extract of leaves of mangrove Rhizophora sp. with a dose of 200 mg/kg BB + glucose 2 g / kg (group P-2) showed the most excellent antidiabetic activity and lowering blood glucose levels by 31.27%. This study confirmed that three types of Mangrove from Sambera Coastal Region Indonesia showed the antidiabetic profile.

Spectrophotometric Analysis of Phenolic Compounds in Grapes and Wines.

Phenolic compounds are of crucial importance for red wine color and mouthfeel attributes. A large number of enzymatic and chemical reactions involving phenolic compounds take place during winemaking and aging. Despite the large number of published analytical methods for phenolic analyses, the values obtained may vary considerably. In addition, the existing scientific knowledge needs to be updated, but also critically evaluated and simplified for newcomers and wine industry partners. The most used and widely cited spectrophotometric methods for grape and wine phenolic analysis were identified through a bibliometric search using the Science Citation Index-Expanded (SCIE) database accessed through the Web of Science (WOS) platform from Thompson Reuters. The selection of spectrophotometry was based on its ease of use as a routine analytical technique. On the basis of the number of citations, as well as the advantages and disadvantages reported, the modified Somers assay appears as a multistep, simple, and robust procedure that provides a good estimation of the state of the anthocyanins equilibria. Precipitation methods for total tannin levels have also been identified as preferred protocols for these types of compounds. Good reported correlations between methods (methylcellulose precipitable vs bovine serum albumin) and between these and perceived red wine astringency, in combination with the adaptation to high-throughput format, make them suitable for routine analysis. The bovine serum albumin tannin assay also allows for the estimation of the anthocyanins content with the measurement of small and large polymeric pigments. Finally, the measurement of wine color using the CIELab space approach is also suggested as the protocol of choice as it provides good insight into the wine's color properties.

Spray drying of pomegranate juice using maltodextrin/cyclodextrin blends as the wall material

Microencapsulation protects sensitive nutrients, masks flavors, or enhances delivery. Ratios of maltodextrin and γ‐cyclodextrin (20:0, 19:1, and 17:3% w/w) were dissolved in water and mixed with pomegranate juice for spray drying with inlet temperatures of 120, 140, and 160°C. The effects on physical properties (water activity, % water content, color, pH, soluble solids (Brix), and methyl cellulose precipitable tannin assay (MCPTA) were examined. Based on the principle component analysis, formulation influenced color parameters and pH accounted for 46.8% of the variation in the data. Temperature influenced Chroma and water‐holding capacity with 31.8% of the variation. The pH of the reconstituted spray‐dried powder significantly influenced color. Blending of γ‐cyclodextrins to maltodextrins slightly increased the water‐holding capacity, increased pH, slightly affected color, and preserved the color over time, slightly better. Increased inlet temperature affected color, decreased water‐holding capacity, and decreased astringency index. Small additions of γ‐cyclodextrin affect spray‐dried powders.

Robust Ultraviolet-Visible (UV-Vis) Partial Least-Squares (PLS) Models for Tannin Quantification in Red Wine.

The validation of ultraviolet-visible (UV-vis) spectroscopy combined with partial least-squares (PLS) regression to quantify red wine tannins is reported. The methylcellulose precipitable (MCP) tannin assay and the bovine serum albumin (BSA) tannin assay were used as reference methods. To take the high variability of wine tannins into account when the calibration models were built, a diverse data set was collected from samples of South African red wines that consisted of 18 different cultivars, from regions spanning the wine grape-growing areas of South Africa with their various sites, climates, and soils, ranging in vintage from 2000 to 2012. A total of 240 wine samples were analyzed, and these were divided into a calibration set (n = 120) and a validation set (n = 120) to evaluate the predictive ability of the models. To test the robustness of the PLS calibration models, the predictive ability of the classifying variables cultivar, vintage year, and experimental versus commercial wines was also tested. In general, the statistics obtained when BSA was used as a reference method were slightly better than those obtained with MCP. Despite this, the MCP tannin assay should also be considered as a valid reference method for developing PLS calibrations. The best calibration statistics for the prediction of new samples were coefficient of correlation (R2val) = 0.89, root mean standard error of prediction (RMSEP) = 0.16, and residual predictive deviation (RPD) = 3.49 for MCP and R2val = 0.93, RMSEP = 0.08, and RPD = 4.07 for BSA, when only the UV region (260-310 nm) was selected, which also led to a faster analysis time. In addition, a difference in the results obtained when the predictive ability of the classifying variables vintage, cultivar, or commercial versus experimental wines was studied suggests that tannin composition is highly affected by many factors. This study also discusses the correlations in tannin values between the methylcellulose and protein precipitation methods.

Evaluation of Antioxidant, Brine Shrimp Lethality and Antimicrobial Activities of Galphimia gracilis Bartl. Leaf Extracts Using in vitro Assay Models

Aims: Galphimia gracilis Bartl. (FamilyMalpighiaceae) is known as gold shower or shower of gold. The current study was designed to evaluate in vitro antioxidant, brine shrimp lethality and antimicrobial activities of methanol, ethyl-acetate and petroleum ether extracts of G. gracilis leaf. Study Design: Phytochemical screening, in vitro antioxidant, brine shrimp lethality and antimicrobial activities. Place and Duration of Study: Department of Pharmacy, Jahangirnagar University, Savar, Dhaka1342. The studies were carried out from November 2013 to February 2014. Original Research Article Hasan et al.; EJMP, 5(1): 66-76, 2015; Article no.EJMP.2015.005 67 Methodology: In vitro antioxidant activity of the extracts were studied using DPPH radical scavenging assay, Nitric oxide (NO) scavenging assay, total phenol content, total flavonoid content, total antioxidant content, total tannin content and lipid peroxidation in human erythrocyte cell assays. Lethality bioassay was performed on Artemia salina Leach nauplii. Antimicrobial activity was investigated by disc diffusion technique. Results: Presence of alkaloids, carbohydrates, flavonoids, steroids, terpenoids, tannins, saponins and glycosides were identified in the extracts. Ethyl-acetate extract (GLEA) showed highest activities in DPPH (IC50 of 21.70±0.51 μg/ml), NO (IC50 of 35.50±0.30 μg/ml), lipid peroxidation in human erythrocyte cells (IC50 of 10.38±0.34 μg/ml), total phenol (934.04±3.21 mg/g Gallic Acid Equivalent), total flavonoid (236.88±2.66 mg/g Quercetin Equivalent) and total antioxidant capacity assays (978.58±1.66 mg/g Ascorbic Acid Equivalent) among three different extracts. Methanol extract (GLM) showed promising reducing capacity than other extracts in cupric reducing (correlation coefficient, r= 0.95 and P<0.05) and reducing power capacity assays (r= 0.993 and P<0.001). Besides, it showed dose dependent activity in both assays. Methanol extract showed maximum content in total tannin assay (89.34±1.37 mg/g Tannic Acid Equivalent). In brine shrimp lethality bioassay, methanol extract was found to be more potent than other extracts (LC50=64.46 μg/ml, χ2=39.87, P<0.0001). In disk diffusion technique, all the extracts showed poor activity. Conclusion: The present findings suggest that leaf extracts of G. gracilis have promising antioxidant and cytotoxic activities. Moreover, the extracts can be used as potential candidates for lead compounds.

Efficacy of Monitoring the Sensory Taste Characteristics in Pomegranate Juice with Electronic Tongue and Chemical Measurements

Taste is imperative in pomegranate juice and is influenced by many production and processing factors. Measuring sweetness, sourness, bitterness, astringency and toothetch without a sensory panel makes economic sense. This investigation compares descriptive sensory evaluation (DSE) with physical methods. Brix degree (Brix), titratable acidity (TA), pH, methyl cellulose precipitable tannin assay (MCPTA) and electronic tongue (e-tongue) techniques were measured in six commercial juices. The MCPTA correlated with astringent (r = 0.57, Pr = 0.01) and bitter (r = 0.58, Pr = 0.01), while pH negatively correlated with sour (r = −0.53, Pr = 0.02). DSE and physical measurements sufficiently differentiated brands. DSE could differentiate between five of the six brands. TA, Brix, pH and MCPTA tests differentiated between all six juice brands. The e-tongue resulted in large differences within four of the brand as well as between brands. Evaluation of these methods by diluting one juice brand facilitated distinguishing how the methods work on the same compounds at declining concentrations. Practical Applications This study compares pomegranate juice chemical measures (Brix, pH, TA, MCPTA) with actual sensory taste and mouth feeling characteristics as well as introduces the e-tongue as an application to monitor quality in processed pomegranate juice. The MCPTA assay reasonably correlated with astringency and can be utilized for estimating pomegranate juice astringency in place of sensory evaluation. The information reported is useful for testing juices in research settings and in monitoring juice production for quality assurance purposes.

Assessment of Tannin Variation in Tamarisk Foliage Across a Latitudinal Gradient

Certain phenotypic traits of plants vary with latitude of origin. To understand if tannin concentration varies among populations of tamarisk (Tamarix spp.) according to a latitudinal gradient, an analytical method was adapted from an enological tannin assay. The tannin content (wet basis) of tamarisk foliage collected from 160 plants grown in a common garden ranged from 8.26 to 62.36 mg/g and was not correlated with the latitude of the original North American plant collection site. Tannins do not contribute to observed differences in herbivory observed among these tamarisk populations.

Relationship between Red Wine Grades and Phenolics. 1. Tannin and Total Phenolics Concentrations

Measuring chemical composition is a common approach to support decisions about allocating foods and beverages to grades related to market value. Red wine is a particularly complex beverage, and multiple compositional attributes are needed to account for its sensory properties, including measurement of key phenolic components such as anthocyanins, total phenolics, and tannin, which are related to color and astringency. Color has been shown to relate positively to red wine grade; however, little research has been presented that explores the relationship between astringency-related components such as total phenolic or tannin concentration and wine grade. The aim of this research has been to investigate the relationship between the wine grade allocations of commercial wineries and total phenolic and tannin concentrations, respectively, in Australian Shiraz and Cabernet Sauvignon wines. Total phenolic and tannin concentrations were determined using the methyl cellulose precipitable (MCP) tannin assay and then compared to wine grade allocations made by winemaker panels during the companies' postvintage allocation process. Data were collected from wines produced by one Australian wine company over the 2005, 2006, and 2007 vintages and by a further two companies in 2007 (total wines = 1643). Statistical analysis revealed a positive trend toward higher wine grade allocation and wines that had higher concentrations of both total phenolics and tannin, respectively. This research demonstrates that for these companies, in general, Cabernet Sauvignon and Shiraz wines allocated to higher market value grades have higher total phenolics and higher tannin concentrations and suggests that these compositional parameters should be considered in the development of future multiparameter decision support systems for relevant commercial red wine grading processes. In addition, both tannin and total phenolics would ideally be included because although, in general, a positive relationship exists between the two parameters, this relationship does not hold for all wine styles.

Tannins determined by various methods as predictors of methane production reduction potential of plants by an in vitro rumen fermentation system

Relationships between chemical constituents, including values obtained with tannin assays (i.e., total phenols, total tannins, condensed tannins and tannin activity using a tannin bioassay) for plant materials (n = 17), and methane production parameters at 24 h of incubation in the in vitro Hohenheim gas method were established. The methane production reduction potential (MRP) was calculated by assuming net methane concentration for the control hay as 100%. The MRP of Bergenia crassifolia leaves and roots, and Peltiphyllum peltatum leaves, was >40%. Amongst the chemical constituents, neutral detergent fibre had a high correlation (r = 0.86) with methane concentration. There was negative relationship between total phenol, total tannins or tannin activity and methane concentration. However, a positive relationship existed between these tannin assays and the MRP, with r-values ranging from 0.54 to 0.79 (P<0.05). A very weak relationship (r = 0.09) occurred between condensed tannins and MRP. Similar results to those with MRP were obtained with the percent increase in methane on addition of polyethylene glycol. The highest correlations, 0.79 and 0.92 (P<0.001), were between tannin activity determined using the tannin bioassay and the MRP, or the percent increase in methane on addition of polyethylene glycol, respectively, suggesting that this tannin assay could be used to identify plants possessing antimethanogenic properties. Leaves of

Adams-Harbertson Protein Precipitation-Based Wine Tannin Method Found Invalid

The poor precision of the Adams-Harbertson wine tannin assay which was proposed for commercial winemaking, thereby creating the real possibility of quality control problems, is documented. The method is a version of the Hagerman and Butler protein precipitation-based tannin method. An extensive invalidation of the assay results with luxury wine data shows that the assay cannot distinguish bottled wine with reasonable accuracy. Five laboratories used Adams-Harbertson to assay 9 replicates each, of 3 bottled wines (n = 135) found in California supermarkets, with tannin concentrations of nominally 500 and 1000 ppm by high-performance liquid chromatography (HPLC). Reliability exceeded the +/-5% industry requirement by nominally 5 times (z-score based on 5% distribution). Coefficient of variation was +/-27%, making the standard deviation range 54% for Pinot Noir, 34% for Merlot, and 44% for Cabernet Sauvignon. Validity exceeded the 100% requirement. Intralaboratory validity recovery was 55-63%. Interwinery validity was 71-178% of the mean for Pinot Noir, 81-144% for Merlot, and 83-164% for Cabernet Sauvignon. Range as a function of the mean was 89% for Pinot Noir, 55% for Merlot, and 67% for Cabernet Sauvignon. Expect intermethod validity to be nominally 50%, i.e., percent recovery to HPLC. These statistically significant errors were predicted by the literature. First-order error is related to the tannin-protein equilibrium constant (Ka), as suggested by the original author, Hagerman, and the protein equivalence point error as suggested by Silber. This does not obviate second-order errors for tannin-protein analytical chemistry. Winemakers using the measurements risk making wines that are relatively more tannic than the measurements report.

Tannin Quantification in Red Grapes and Wine: Comparison of Polysaccharide- and Protein-Based Tannin Precipitation Techniques and Their Ability to Model Wine Astringency

Quantification of red grape tannin and red wine tannin using the methyl cellulose precipitable (MCP) tannin assay and the Adams-Harbertson (A-H) tannin assay were investigated. The study allowed for direct comparison between the repeatability of the assays and for the assessment of other practical considerations such as time efficiency, ease of practice, and throughput, and assessed the relationships between tannin quantification by both analytical techniques. A strong correlation between the two analytical techniques was observed when quantifying grape tannin (r(2) = 0.96), and a good correlation was observed for wine tannins (r(2) = 0.80). However, significant differences in the reported tannin values for the analytical techniques were observed (approximately 3-fold). To explore potential reasons for the difference, investigations were undertaken to determine how several variables influenced the final tannin quantification for both assays. These variables included differences in the amount of tannin precipitated (monitored by HPLC), assay matrix variables, and the monomers used to report the final values. The relationship between tannin quantification and wine astringency was assessed for the MCP and A-H tannin assays, and both showed strong correlations with perceived wine astringency (r(2) = 0.83 and r(2) = 0.90, respectively). The work described here gives guidance to those wanting to understand how the values between the two assays relate; however, a conclusive explanation for the differences in values between the MCP and A-H tannin assays remains unclear, and further work in this area is required.

High Throughput Analysis of Red Wine and Grape PhenolicsAdaptation and Validation of Methyl Cellulose Precipitable Tannin Assay and Modified Somers Color Assay to a Rapid 96 Well Plate Format

The methyl cellulose precipitable (MCP) tannin assay and a modified version of the Somers and Evans color assay were adapted to high-throughput (HTP) analysis. To improve efficiency of the MCP tannin assay, a miniaturized 1 mL format and a HTP format using 96 well plates were developed. The Somers color assay was modified to allow the standardization of pH and ethanol concentrations of wine samples in a simple one-step dilution with a buffer solution, thus removing inconsistencies between wine matrices prior to analysis and allowing for its adaptation to a HTP format. Validation studies showed that all new formats were efficient, and results were reproducible and analogous to the original formats.

Quantification of condensed tannins by precipitation with methyl cellulose: development and validation of an optimised tool for grape and wine analysis

A precipitation assay is presented that enables tannin measurement in matrices of red wine, 50% ethanol grape extract and aqueous tannin solutions. By exploiting the polysaccharide polymer methyl cellulose to precipitate tannins, the absorbance of phenolics at 280 nm before and after tannin precipitation (subtractive approach) can be obtained, thus enabling selective measurement of tannin only. This methyl cellulose precipitable (MCP) tannin assay allows complete precipitation of tannin from red wine and from grape homogenate extracts. The subtractive assay is both simple and robust, selective for condensed tannins and does not suffer interference from other 280 nm-absorbing phenolics such as anthocyanins or catechins. Matrix effects have only minimal impact on the assay performance and validation parameters indicate a robust performance. There was good correlation between tannin measured by reverse-phase HPLC and the MCP tannin assay for 121 Australian red wines (r= 0.74) and also 54 grape extracts (r= 0.79). We envisage that the technical simplicity of this tannin assay will enable widespread research and field applications. In addition, an alternative format that requires re-solubilisation of the tannin-polymer pellet in acetonitrile is reported, which is particularly suitable for measurement of smaller tannin concentrations. Notwithstanding that option, technical requirements of the re-solubilisation step lead us to suggest that the subtractive format would be simple for adoption by wine industry practitioners.

Determination of hydrolyzable tannins in the fruit of Terminalia chebula Retz. by high-performance liquid chromatography and capillary electrophoresis.

A RP-HPLC method for determining fourteen components (gallic acid, chebulic acid, 1,6-di-O-galloyl-D-glucose, punicalagin, 3,4,6-tri-O-galloyl-D-glucose, casuarinin, chebulanin, corilagin, neochebulinic acid, terchebulin, ellagic acid, chebulagic acid, chebulinic acid, and 1,2,3,4,6-penta-O-galloyl-D-glucose) in the fruit of Terminalia chebula Retz. is described. The separation was achieved within 80 min using a binary gradient with mobile phases consisting of a pH 2.7 phosphoric acid solution and an 80% CH3CN solution. Capillary electrophoretic analyses were also attempted, and it was found that CZE (25 mM Na2B4O7, 5 mM NaH2PO4, pH 7.0) was an efficient method for the separation of gallotannins, while an MEKC method (25 mM Na2B4O7, 5 mM NaH2PO4, 20 mM SDS, pH 7.0, and 10% acetonitrile) provided a better separation for most of the tannins examined. The HPLC and CE methods developed were both successfully applied to the assay of tannins in commercial samples of Chebulae Fructus.