Chemical and DNA authentication of taste variants of Gynostemma pentaphyllum herbal tea

Gynostemma pentaphyllum Makino (Gp) was once used as a sweetener in Japan and is now widely consumed as an herbal tea worldwide for lowering cholesterol levels. Two taste variants, bitter and sweet, of Gp exist in the commercial market, but they cannot be differentiated morphologically nor by existing chemical analytical methods. This has been creating a problem in quality control of Gp products. In the present study, using HPLC-DAD and HPLC–ESI-MS analysis, we found that the Gp saponins, not flavonoids, from the sweet and bitter variants have distinctly different profiles. In addition, the two variants share only 69.01% homology in the ribosomal ITS-1 region, suggesting a phylogenic gap between these two variants. The combinations of chemical profiling and phylogenic analysis clearly confirm, for the first time, the distinction between these two taste variants. This information has direct application in the authentication and quality assessment of the various Gynostemma tea products.     

Chemical composition and anti-proliferative and anti-inflammatory effects of the leaf and whole-plant samples of diploid and tetraploid Gynostemma pentaphyllum (Thunb.) Makino

Leaf and whole-plant samples of the diploid and tetraploid Gynostemma pentaphyllum (GP) were investigated and compared for their chemical compositions, and their potential anti-proliferative and anti-inflammatory effects. The highest levels of total flavonoids and phenolics were observed in the diploid leaf botanical (2L3) at 36.84 mg rutin equiv/g and 41.15 mg gallic acid equiv/g, respectively. The diploid leaf sample (2L2) had the highest amount of rutin and quercetin contents of 77.7 μmol quercetin equiv/g. The tetraploid whole-plant botanical (4L3) had the highest total saponin content of 227.1 mg gypenoside equiv/g. Extracts from all tested GP samples showed time- and dose-dependent antiproliferative effects in HT-29 cells, and the diploid leaf samples had the overall highest inhibitory activity. These extracts had different order of antiproliferative properties in the LNCaP cells, suggesting the potential selective inhibition of GP extracts against different types of cancer cells and the effect of the cell model in screening and evaluation of antiproliferative components. In addition, the diploid leaf extracts showed the strongest inhibitory effects on the expression of TNF-α, IL-6 and COX-2 mRNA at final concentrations of 0.2 and 1 mg botanical equiv/ml media. The results from this study will be used to develop new nutraceutical products from G. pentaphyllum.     

Determination of six lignans in Schisandra chinensis (Turcz.) Baill. Fruits and related Chinese multiherb remedies by HPLC

A simple, rapid and specific HPLC method was established for simultaneous determination of six major lignans in Schisandra chinensis and related Chinese multiherb remedies (CMRs) containing this herb. The six lignans were successfully separated on a C₁₈ column by gradient elution using acetonitrile and water as the mobile phase, and detection wavelength was set at 225 nm. The method was validated through the following performance criteria: linearity, precision, repeatability, stability, accuracy, limit of detection (LOD) and quantification (LOQ). This assay was successfully used for determination of six lignans in 10 raw herbs collected from different regions in China and five Chinese multiherb remedies. Significant variations were demonstrated in the contents of six compounds in these samples. This HPLC method established is suitable for routine quantitative analysis of S. chinensis and multiherb remedies containing this herb.     

Chromatographic determination of polysaccharides in Lycium barbarum Linnaeus

Polysaccharides in Lycium barbarum Linnaeus have been shown to be effective in preventing cancer. The objectives of this study were to develop an appropriate method for molecular weight determination of polysaccharides in L. barbarum. The most suitable analytical condition was: a volume-ratio of L. barbarum sample to deionized water at 1:10, followed by shaking in a 100 °C water bath for 30 min, concentrating to 50 mL and adding 250 mL of 95% ethanol for precipitation at −20 °C for 8 h, hydrolysing protein with 2.5 U/mL of proteinase at pH 8 and 60 °C for 4 h and separating polysaccharide into five fractions by high-performance size exclusion chromatography (HPSEC) with the molecular weight of two major fractions being 79,250 and 24,468 Da. Analysis of monosaccharides by gas chromatography (GC) indicated the presence of rhamnose, arabinose, xylose, mannose, glucose and galactose, with the molar ratio at 0.3:2.7:0.3:0.2:2.7:0.9, respectively.     

柱前衍生化HPLC法分析真菌多糖的单糖组成

通过柱前衍生化HPLC法分析了6种真菌多糖的单糖组成。将6种真菌的胞外粗多糖及其菌体用1mol/L的硫酸水解成单糖,用1-苯基-3-甲基-5-吡唑啉酮(PMP)衍生化,利用高效液相色谱法检测各单糖。结果表明,灵芝、树舌类真菌胞外多糖的产率均较高,胞外多糖产率在0.0921～0.1528g/100mL之间。胞外多糖主要以甘露糖为主,其次含有葡萄糖和半乳糖,并且一般含有少量的阿拉伯糖。胞内多糖中葡萄糖含量最高,其次含有甘露糖和半乳糖,个别菌种还含有少量的阿拉伯糖和葡萄糖醛酸。而虫草胞外多糖的组成依次为半乳糖、葡萄糖、甘露糖和葡萄糖醛酸,胞内多糖同样是主要含有葡萄糖,其次含有甘露糖、半乳糖和葡萄糖醛酸。     

Simultaneous quantification of selected compounds from Salvia herbs by HPLC method and their application

A simple and reliable reversed-phase high performance liquid chromatographic method was developed and validated for simultaneous determination of danshensu, protocatechuic aldehyde, rosemarinic acid, salvianolic acid B, dihydrotanshinone I, tanshinone I, cryptotanshinone and tanshinone IIA in Salvia herbs. The contents of these eight main components were compared between Salvia miltiorrhiza Bunge and Salvia przewalskii Maxim, which were used for treating coronary heart diseases, cerebrovascular disease, bone loss, hepatitis, hepatocirrhosis and chronic renal failure. The samples were successfully separated by the HPLC method. All the standard compounds showed a good linearity (R² > 0.9994) in the relatively wide concentration range. The limit of detection of the eight compounds was in the range of 0.05–0.5 μg/ml and the limit of quantification was in the range of 0.25–1.50 μg/ml. The intra-day variability was in the range of 0.03–8.08% and the inter-day variability was in the range of 0.18–12.36%. The recoveries of the selected compounds were in the range of 96.2–108.1%. This method was accurate, precise and reproducible, and could be successfully applied to the quality control and stable experiment for the preparations consisted of these active components, and the content comparison of the herbs from Salvia species.     

Identification and quantification of phenolics in aromatic bitter and cherry liqueur by HPLC with electrochemical detection

An HPLC-based method with electrochemical detection was optimized for quantitative analysis of phenolic compounds in an aromatic bitter and a cherry liqueur, which are different types of alcoholic beverages with a high phenolic content. The method requires only filtering of samples prior to the HPLC analysis and is recommended for routine analysis. Two slightly different gradients were needed for the analysis of the two beverages. Eleven and eight different phenolic compounds (cinnamic acid derivatives, benzoic acid derivatives, flavan-3-ols, flavanons, and eugenol) were identified and quantified in the bitter and the cherry liqueur, respectively.     

Simultaneous analysis of purine alkaloids and catechins in Camellia sinensis, Camellia ptilophylla and Camellia assamica var. kucha by HPLC

Simultaneous analysis of purine alkaloids and catechins in tea from dry leaves of Camellia sinensis, Camellia ptilophylla and Camellia assamica var. kucha by a reversed-phase high-performance liquid chromatography (RP-HPLC) method. This HPLC method had been proved to be appropriate for the identification and quantification of purine alkaloids and catechins, and exhibited good correlation coefficients, detection levels and recovery rates. Caffeine, theobromine, theacrine, (+)-catechin, (−)-epicatechin, (−)-epigallocatechin, (−)-epigallocatechin gallate, (−)-epicatechin gallate, (−)-gallocatechin gallate and (−)-gallocatechin were identified and quantified in the three species of genus Camellia Sect. Thea. There was 2.72% caffeine and 0.26% theobromine in C. sinensis, 4.85% theobromine in C. ptilophylla, and 1.58% theacrine, 0.94% caffeine and 0.45% theobromine in C. assamica var. kucha. Theacrine in C. sinensis and C. ptilophylla, and caffeine in C. ptilophylla were not detected. These data highlight differences in the relative proportions of purine alkaloids in the three species of Camellia Sect. Thea. In addition, different catechins were identified and quantified. The highest content of catechin in dry leaves was (−)-epigallocatechin gallate (EGCG) 3.51%, (−)-gallocatechin gallate (GCG) 9.88% and (−)-epigallocatechin gallate (EGCG) 6.78% in C. sinensis, C. ptilophylla and C. assamica var. kucha, respectively.     

Rosmarinic acid from beach waste: Isolation and HPLC quantification in Zostera detritus from Arcachon lagoon

Detritus of Zostera noltii and Zostera marina collected from the beaches of Arcachon lagoon (France) over a 30-month period were screened as a new source of rosmarinic acid (RA), a phenolic acid and an economically important metabolite. The seasonal variation of the RA content was quantified in methanolic crude extracts using high-performance liquid chromatography. Concentrations of RA ranged from 2.2 to 18.0 mg g⁻¹ (dw) for Z. noltii and 1.3 to 11.2 mg g⁻¹ (dw) for Z. marina. This is the first time RA has been isolated from Z. noltii; detrital leaves of Zostera have never before been screened for their bioactive substances. The high RA content of Zostera flotsam is of interest for both the cosmetic and herbal industries. These results show that there is a real potential for harvesting beachcast Zostera.     

Determination of the change of flavonoid components as the defence materials of Citrus unshiu Marc. fruit peel against Penicillium digitatum by liquid chromatography coupled with tandem mass spectrometry

A healthy fruit peel of Citrus unshiu Marc. and one infected by Penicillium digitatum were analysed for flavonoids via high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC–MS/MS) in the positive mode with selected ion monitoring (SIM). Among 16 flavonoid components characterised in C. unshiu Marc., four flavanones and nine flavones were identified for the first time. The identified compounds were quantified by HPLC–UV. To investigate the function of the flavonoids as defence materials, the flavonoid content change of the fruit peel inoculated with P. digitatum was monitored by HPLC. The flavonoid concentration in the infected fruit peel decreased initially after the infection and then gradually increased before finally progressively decreasing.     

Determination of aloe emodin in Aloe vera extracts and commercial formulations by HPLC with tandem UV absorption and fluorescence detection

Aloe emodin is an anthraquinone compound found in Aloe vera and other species of the Asphodelaceae and the Polygonaceae families, which has recently attracted much attention as a prospective antineoplastic agent. A HPLC method with tandem UV absorption and fluorimetric detection, was developed and validated for the analysis of aloe emodin in products obtained from Aloe leaves, such as capsules, tablets, dried extracts and mother tinctures. The stationary phase was a C18 reversed-phase column and the mobile phase was composed of water and methanol (30/70, v/v). Satisfactory linearity was obtained over the 10.0–1000.0 ng/mL range for photodiode array detection (limit of detection: 3 ng/mL) and over the 2.5–1000.0 ng/mL range for fluorimetric detection (limit of detection: 0.8 ng/mL). Aloe emodin levels were determined in A. vera extracts and commercial formulations by both detection means, with good precision (R.S.D. < 9%) and accuracy (recovery > 85%) and consistent results. Thus, the method seems to be suitable for the analysis of aloe emodin in different herbal and commercial products; it could also be useful for the identity confirmation of formulations and extracts.     

Characterisation of catechins in green and black teas using square-wave voltammetry and RP-HPLC-ECD

Eight green and three black teas have been analysed using square-wave voltammetry at a glassy-carbon electrode, at pH 2 and square-wave frequency 100 Hz. The SWV response of teas consisted of two partially resolved anodic peaks: peak 1 at around 0.372 V, and peak 2 around 0.491 V. First anodic peak was due to oxidation of pyrogallol catechins, i.e. epigallocatechin gallate (EGCG) and epigallocatechin (EGC), and peak 2 can be associated with oxidation of ortho-hydroquinone and galloyl groups in catechins, gallic acid, theaflavins, and other catechol-type phenolics present in tea. Peak 1 has been chosen for quantification purposes, and concentrations of catechins were calculated from the calibration plot constructed for EGC used as external standard. Amounts of catechins in green teas obtained by SWV were in good agreement with those obtained using reverse-phase high-performance liquid chromatography with electrochemical detection (RP-HPLC-ECD). For black tea samples the detector response of peak 1 was diminished due to interferences of coexisting polyphenols, and SWV obtained results were something lower than those obtained by HPLC-ECD.     

Antioxidation and antiglycation of polysaccharides from Misgurnus anguillicaudatus

Misgurnus anguillicaudatus is a fish widely inhabiting rivers, lakes, ponds, swamps and rice fields in China, Japan, Korea and other Asian countries. It is edible and can be used to treat xiao ke (namely diabetic) in traditional Chinese medicine. Polysaccharides from M. anguillicaudatus (MAPs) were extracted and separated into four parts which were named MAP I, MAP II MAP III and MAP IV, respectively. The percentage of polysaccharide in dry weight is about 3.06%. Hydroxyl radical and superoxide radical anion scavenging activities together with glycation inhibition of these polysaccharides were investigated. The results showed that all the MAPs had an inhibitory effect on oxidation and glycation and exhibited a dose-dependent response. Especially MAP IV displayed a stronger antioxidation effect than glucosamine and better antiglycation activities than aminoguanidine. As oxidation and glycation are relevant to diabetic complications, the result of this work suggests that MAPs may be effective in preventing and treating diabetic complications.     

Characterization and in vitro antioxidation of papain hydrolysate from black-bone silky fowl (Gallus gallus domesticus Brisson) muscle and its fractions

Black-bone silky fowl (Gallus gallus domesticus Brisson) (BSF) muscle was hydrolyzed by papain, and the hydrolysate was separated by preparative high performance liquid chromatography (HPLC). The amino acid composition of the BSF hydrolysate (BSFH) and its fractions was determined by HPLC precolumn derivation with 2,4-dinitrofluorobenzene. The molecular weight (MW) distribution of the BSFH and its fractions was measured by a peptide column on an HPLC system. Antioxidant activities of the BSFH and its fractions were studied by testing the reducing power and four radical scavenging systems: superoxide anion (O₂⁻), hydroxyl (·OH), 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS⁺) radicals. The results demonstrated that the BSFH had strong antioxidant capacity to scavenge O₂⁻, DPPH and ABTS⁺, and displayed strong reducing power, but revealed less powerful ability to scavenge ·OH. Fraction II of the BSFH exhibited the highest activity in scavenging O₂⁻ and DPPH, and reducing power, whereas fraction I displayed the strongest ·OH scavenging ability. Besides Glu, Asp and Gly, the rich amino acids of Ala and Leu played an important role in antioxidant activity. The small-size peptides with MW ranging from approximately 200-6000 Da probably contributed to higher antioxidant activity. Results from this study indicated that BSFH and its fractions could be used as food additives and diet nutrients.     

Bioactive dammarane-type triterpenoids derived from the acid hydrolysate of Gynostemma pentaphyllum saponins

From an acid hydrolysate of the crude saponins of Gynostemma pentaphyllum, three triterpenes, including a new compound (23S)-3β-hydroxydammar-20,24-dien-21-oic acid 21,23-lactone (1) and two known aglycons (20S, 23R)-3β,20β-dihydroxydamma-24-dien-21-oic acid 21,23-lactone (2) and (20S, 24S)-20,24-epoxydammarane-3β,12β,25-triol (3), were isolated. Their structures were established on the basis of extensive spectral evidence (HR-ESI-MS, IR, 1D and 2D-NMR experiments). In bioactive assays in vitro, compound 1 was found to have potent cytotoxicity against the human breast cancer cells MDA-MB-435, whereas compounds 2 and 3 exhibited modest inhibitory activity toward porcine pancreatic lipase. The results indicated that acid treatment of G. pentaphyllum extract could produce diverse structures with interesting bioactivity.     

Flow injection spectrophotometry using natural reagent from Morinda citrifolia root for determination of aluminium in tea

A flow injection (FI) spectrophotometric method with using natural reagent extracted from Morinda citrifolia root has been developed for determination of aluminium. The extract contained anthraquinone compounds which could react with Al³⁺ to form reddish complexes which had maximum absorption wavelength at 499.0 nm. The extract could be used as a reagent in FI system without further purification to obtain pure compound. A sensitive method for determination of aluminium in concentration range of 0.1-1.0 mg L⁻¹, with detection limit of 0.05 mg L⁻¹ was achieved. Relative standard deviations of 1.2% and 1.7% were obtained for the determination of 0.1 and 0.6 mg L⁻¹ Al³⁺ (n = 11). Sample throughput of 35 h⁻¹ was achieved with the consumption of 3 mL each of carrier and reagent solutions per injection. The developed method was successfully applied to tea samples, validated by the FAAS standard method. The method is simple, fast, economical and could be classified as a greener analytical method.     

Antioxidant activity of Camellia sinensis leaves and tea from a lowland plantation in Malaysia

Methanol extracts of fresh tea leaves from a lowland plantation in Malaysia were screened for total phenolic content (TPC) and antioxidant activity (AOA). AOA evaluation included 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical-scavenging ability, ferric-reducing antioxidant power (FRAP), and ferrous-ion chelating (FIC) ability. Ranking, based on TPC and AOA, was as follows: shoots > young leaves > mature leaves. TPC and AOA of lowland leaves were comparable to those of highland plants. A green tea produced by drying young leaves in a household microwave oven for 4 min showed significantly higher TPC and AOA than did four commercial brands of green and black tea.     

Effects of physicochemical parameters on the production of phenolic acids from palm oil mill effluent under liquid-state fermentation by Aspergillus niger IBS-103ZA

The present investigation is an effort to develop an environmentally friendly and cost-effective liquid-state fermentation process by introducing a new locally isolated fungal strain of Aspergillus niger (IBS-103ZA) for the production of phenolics from a new source, palm oil mill effluent (POME). Sucrose, manganese sulphate (MnSO₄) and temperature were identified as the most significant variables in improving phenolics production. Optimisation increased the total phenolic content from 856 ± 2.22 to 941 ± 3.72 GAE mg/l at 35.0 °C, 6.0% (w/v) sucrose, 2.7% (w/v) MnSO₄, and with other parameters fixed. The fermented extract (FE) with IC₅₀ value of 0.45 mg/ml showed the strongest antioxidant potency, compared to unfermented extract (UFE), with IC₅₀ of 1.13 mg/ml, and the synthetic antioxidant, BHT, with IC₅₀ of 0.63 mg/ml. The phenolic compounds were identified and quantified by HPLC.     

Isolation,chemical composition and antioxidant activities of a water-soluble polysaccharide from Cyclocarya paliurus (Batal.) Iljinskaja

A water-soluble polysaccharide was isolated from the water extract of Cyclocarya paliurus (Batal.) Iljinskaja, which is a well-known native health tea in China. This polysaccharide was named as CPP-1. The molecular weight of CPP-1 was determined by high-performance gel permeation chromatography, with an average molecular weight of about 1167 kDa. The analysis of monosaccharide composition in the polysaccharide by gas chromatography revealed that it was a heteropolysaccharide and consisted of d-xylose, l-arabinose, d-glucose, d-galactose, l-rhamnose and d-mannose in a molar ratio of 1.00:9.67:9.65:4.96:3.29:2.70. Furthermore, CPP-1 contains 8.44% of protein and 17 general amino acids, and it is rich in glutamic acid, asparagic acid, leucine, glycine, arginine, tyrosine and alanine. The antioxidant activity of CPP-1 was also evaluated. It was found that CPP-1 exerted significant scavenging effects on DPPH radicals with a value of around 91.4%, compared to the reference controls of BHT (91.2%) and ascorbic acid (98.9%) at a concentration of 400 μg/ml, and with EC₅₀ values of 52.3 μg/ml.