Ashu Gulati

Seasonal Variations in Quality of Kangra Tea ( Camellia sinensis(L) O Kuntze) in Himachal Pradesh

Periodic seasonal variations in infusion quality of orthodox Kangra tea over various growth flushes are reported. Theaflavins, thearubigins and caffeine recorded maximum content during early flush and gradual decline with progress in season, showing a minimum during main flush and slight improvement through backend flush. Flavour profile analysis also revealed qualitative and quantitative seasonal variations in aroma complex. High proportions of provisionally identified flavour components, linalool, geraniol, β-ionone, methyl salicylate, phenyl acetaldehyde, trans-2-hexenal, and several unidentified components with typical Kangra notes recorded during early flush exhibited comparatively lower contents or even total loss through main flush. Flavour quality also registered slight improvement in backend flush over main flush. Results suggest increasing crop productivity during early and backend flushes could enhance profitability.

Differential display mediated cloning of anthocyanidin reductase gene from tea (Camellia sinensis) and its relationship with the concentration of epicatechins

Tea [Camellia sinensis (L.) O. Kuntze] leaves are a major source of epicatechin (EC) and its gallolyl derivatives epicatechin gallate, epigallocatechin and epigallocatechin gallate, collectively known as epicatechins (ECs). Epicatechins are important factors determining tea quality, and they also possess many medicinal properties. To gain further information about the regulation of the biosynthesis of ECs, we cloned the gene encoding anthocyanidin reductase from tea (CsANR) by first quantifying changes in the concentrations of ECs in response to drought, gibberellic acid (GA(3)), abscisic acid (ABA) and wounding treatments, followed by differential display of mRNAs and analysis of those bands exhibiting a change in expression paralleling the treatment-induced changes observed in the EC data. Analysis of 133 bands yielded a partial cDNA of CsANR that was later cloned to the full length by rapid amplification of the cDNA ends. The full-length CsANR (Accession No. AY641729) comprised 1233 bp with an ORF of 1014 bp (from 79 to 1092 bp) encoding a polypeptide of 337 amino acids. Expression of CsANR in an Escherichia coli expression vector yielded a functional protein that catalyzed the conversion of cyanidin to EC in the presence of NADPH. Analysis of ECs and gene expression in leaves at different developmental stages and across five tea clones exhibiting variable concentrations of ECs revealed a positive correlation between concentration of ECs and CsANR expression. Expression of CsANR was down-regulated in response to drought, ABA and GA(3) treatments and up-regulated in response to wounding.

Catechin and catechin fractions as biochemical markers to study the diversity of Indian tea (Camellia sinensis (L.) O. Kuntze) germplasm.

The heterogeneous Indian tea germplasm includes ‘China’, ‘Assam’, ‘Cambod’, and their hybrids which were evaluated using biochemical markers viz., total catechin and their fractions, for varietal identification and characterization. Principal component analysis (PCA) of biochemical characters showed that the total catechin and trihydroxylated catechin has higher eigenvalues. The first two principal components (PCs) could differentiate more than 90% of the clones studied. This grouping based on first two principal component matrices differentiated ‘China’, and their hybrids with ‘Assam’ and ‘Cambod’ variety. Morphologically indistinct large‐leaved ‘Cambod’ variety and ‘Assam’ varieties could not be differentiated using biochemical markers, since both varietal types taxonomically belong to a single species. Clones of ‘China’ type showed low total catechin content and catechin ratio which are distinctly grouped. The ‘China–Assam’ and ‘China–Cambod’ hybrids formed intermediate groups between ‘China’ PC group and ‘Cambod’/‘Assam’ PC groups, providing evidence for genetic control of catechin ratio variation. Tea clones which are differentially positioned in the PC group could be explained based on the genetic contribution by other varietal type as parents. This biochemical characterization will be a useful tool in the development of quality‐tea clones with different proportion of total catechin and their fractions.

Caffeine Biosynthesis and Degradation in Tea [Camellia sinensis (L.) O. Kuntze] is under Developmental and Seasonal Regulation

To study caffeine biosynthesis and degradation, here we monitored caffeine synthase gene expression and caffeine and allantoin content in various tissues of four Camellia sinensis (L.) O. Kuntze cultivars during non-dormant (ND) and dormant (D) growth phases. Caffeine synthase expression as well as caffeine content was found to be higher in commercially utilized tissues like apical bud, 1st leaf, 2nd leaf, young stem, and was lower in old leaf during ND compared to D growth phase. Among fruit parts, fruit coats have higher caffeine synthase expression, caffeine content, and allantoin content. On contrary, allantoin content was found lower in the commercially utilized tissues and higher in old leaf. Results suggested that caffeine synthesis and degradation in tea appears to be under developmental and seasonal regulation.

Analysis of the essential oil of large cardamom (Amomum subulatum Roxb.) growing in different agro-climatic zones of Himachal Pradesh, India.

BACKGROUND The aim of the present study was to investigate variations in the chemical composition of the essential oil from seeds of large cardamom grown at different altitudes in Himachal Pradesh, India. The composition of the essential oil was determined by gas chromatography (GC), gas chromatography-mass spectrometry (GC-MS) and gas chromatography-olfactometry (GC-O). RESULTS The oil components showed qualitative and quantitative variations in the composition. GC and GC-MS analysis led to the identification of 55 compounds representing 98% of total oil. Major components in the oil were 1,8-cineole, α-terpineol, DL-limonene, nerolidol, 4-terpineol, δ-terpineol, δ-3-carene, β-myrcene, germacrene D, α-terpinene and longifolenaldehyde. The oil yields obtained were 9.8-19.5 g kg(-1). Cardamom oil from Himachal Pradesh was found to contain new compounds, viz. 4-terpineol, δ-3-carene, trans-sabinene hydrate, 1-phellandrene, α-terpinene, bicyclo-germacrene, isopinocarveol and ledenoxid-II. α-Terpenyl acetate, the major constituent of small cardamom, was also detected in the oil of large cardamom grown in Himachal Pradesh. Application of aroma extract dilution analysis revealed 35 compounds having aroma impact with the flavour dilution factor ranging from 2 to 1024, and 34 of these compounds were identified. The five most intense aromatic components are dl-limonene, 1,8-cineole, β-myrcene, α-pinene, α-basabolol. This is the first time that the characterisation of odour-active compounds has been carried out on large cardamom. CONCLUSION The presence of 4-terpineol, δ-3-carene, trans-sabinene hydrate, 1-phellandrene, α-terpinene, 1-terpineol, bicyclogermacrene, isopinocarveol, ledenoxid-II, longifolenaldehyde and α-terpenyl acetate make the aroma of the oil different from large cardamom oil of Sikkim and could offer potential as a new food flavour.

Performance Evaluation of a Novel iTongue for Indian Black Tea Discrimination

In this work, the multi-electrode-single-frequency (MESF), multi-frequency-single-electrode (MFSE), and multi-frequency-multi-electrode (MFME) impedance responses of an impedance-Tongue reported previously, are evaluated for their discriminability of Indian Black Teas. Principal component analysis (PCA) in conjunction with a cluster validity measure, Davis-Bouldin Index (DBI), has been used for discriminability evaluation. The discriminabilities of electrode specific frequency segments chosen by optimization algorithms, namely, Genetic Algorithm (GA) and Particle Swarm Optimization (PSO) have also been evaluated. The results show that the MFSE impedance response of Gold electrode gives the best discriminability without compromising the system complexity as against MESF, MFSE, MFME, and GA/PSO-optimized response. The results also suggest that the cross-sensitivity of electrodes may be enhanced by selecting optimum frequencies and/or electrodes, paralleling the practice of modifying the electrodes. This opens up a new approach towards qualitative and quantitative analysis of complex liquids.

Plant growth promoting potential of the fungus Discosia sp. FIHB 571 from tea rhizosphere tested on chickpea, maize and pea

The ITS region sequence of a phosphate-solubilizing fungus isolated from the rhizosphere of tea growing in Kangra valley of Himachal Pradesh showed 96% identity with Discosia sp. strain HKUCC 6626 ITS 1, 5.8S rRNA gene and ITS 2 complete sequence, and 28S rRNA gene partial sequence. The fungus exhibited the multiple plant growth promoting attributes of solubilization of inorganic phosphate substrates, production of phytase and siderophores, and biosynthesis of indole acetic acid (IAA)-like auxins. The fungal inoculum significantly increased the root length, shoot length and dry matter in the test plants of maize, pea and chickpea over the uninoculated control under the controlled environment. The plant growth promoting attributes have not been previously studied for the fungus. The fungal strain with its multiple plant growth promoting activities appears attractive towards the development of microbial inoculants.

High Level of Genetic Diversity Among the Selected Accessions of Tea (Camellia sinensis) from Abandoned Tea Gardens in Western Himalaya

To revive cultivation of the tea unique to the western Himalayan region, it is important to evaluate the seed-derived bushes available in the area’s abandoned gardens. This study used quantitative leaf characters, catechin content, and AFLP markers to assess these China cultivar type bushes. Compared with other China cultivar germplasm, these accessions showed a higher level of diversity among themselves. Among the quantitative morphological characters, leaf length is important in distinguishing the accessions studied, with a high loading value in the principal component analysis. The catechins and AFLP markers displayed the genetic makeup of the accessions. Other than total catechins, the trihydroxylated catechins showed a high loading value in differentiating the accessions. The genetic control of the ratio of dihydroxylated and trihydroxylated catechins is found to be based on a correlation with AFLP markers. The genetic similarity between Kangra Asha and Kangra Jat suggests that Kangra Jat must be descended from Kangra Asha. Kangra Jat is well adapted to local environmental conditions, as is evident from its high catechin content.

Increase in flavan-3-ols by silencing flavonol synthase mRNA affects the transcript expression and activity levels of antioxidant enzymes in tobacco.

Flavonoids are plant secondary metabolites widespread throughout the plant kingdom involved in many physiological and biochemical functions. Amongst the flavonoids, flavan-3-ols (catechin and epicatechin) are known for their direct free radical scavenging activity in vitro, but studies on their antioxidant potential and interaction with antioxidant enzymes in vivo are lacking. Here, the flavonoid pathway was engineered by silencing a gene encoding flavonol synthase (FLS) in tobacco to direct the flow of metabolites towards production of flavan-3-ols. FLS silencing reduced flavonol content 17-53%, while it increased catechin and epicatechin content 51-93% and 18-27%, respectively. The silenced lines showed a significant increase in expression of genes for dihydroflavonol reductase and anthocyanidin synthase, a downstream gene towards epicatechin production, with no significant change in expression of other genes of the flavonoid pathway. Effects of accumulation of flavan-3-ols in FLS silenced lines on transcript level and activities of antioxidant enzymes were studied. Transcripts of the antioxidant enzymes glutathione reductase (GR), ascorbate peroxidase (APx), and catalase (CAT) increased, while glutathione-S-transferase (GST), decreased in FLS silenced lines. Enhanced activity of all the antioxidant enzymes was observed in silenced tobacco lines. To validate the affect of flavan-3-ols on the antioxidant system, in vitro experiments were conducted with tobacco seedlings exposed to two concentrations of catechin (10  and 50 μm) for 2 days. In vitro exposed seedlings produced similar levels of transcripts and activity of antioxidant enzymes as FLS silenced seedlings. Results suggest that flavan-3-ols (catechin) might be increasing activity of GR, Apx and CAT by elevating their mRNAs levels. Since these enzymes are involved in scavenging of reactive oxygen species, this strategy would help in tailoring crops for enhanced catechin production as well as making them tolerant to oxidative stresses.

Variation in chemical composition and quality of tea (Camellia sinensis) with increasing blister blight (Exobasidium vexans) severity

Analysis of quality-related parameters at the green leaf-stage and in orthodox-made tea revealed marked quality deterioration with increasing blister blight severity. Total phenols, catechin(s), total nitrogen, amino acids, and chlorophylls, as well as polyphenol oxidase activity, declined in tea shoots with increasing disease severity. Depending on disease intensity, orthodox tea made from infected shoots showed loss in leaf-style. Likewise, theaflavins, thearubigins, caffeine, and high polymerized substances as well as total liquor colour, brightness and briskness also exhibited marked progressive decline in tea made from shoots with increasing disease severity. Aroma components, particularly hexene-I-ol, phenyl acetaldehyde, linalool, methyl salicylate, geraniol, indole, β-ionone, nerolidol, and several unassigned components were also lower in disease affected teas. The magnitude of decline in various aroma constituents corresponded to disease intensity.