Stage-specific metabolomics suggests a trade-off between primary and secondary metabolites for nutritional advantage in Lepidium latifolium L.

Lepidium latifolium L. is an established phytofood of the Ladakh Himalayas that contains differential content of important glucosinolates (GLS) in specific stages of sprouts. Therefore, in order to harness its nutraceutical potential, a comprehensive mass spectrometry-based stage-specific untargeted metabolomic analysis was performed. A total of 318 metabolites were detected, out of which 229 were significantly (p ≤ 0.05) changed during different stages. The Principal Component Analysis plot clearly differentiated different growth stages into three clusters. The nutritionally important metabolites, including amino acids, sugars, organic acids, and fatty acids, were found significantly (p ≤ 0.05) higher in the first cluster consisting of 1st, 2nd and 3rd week sprouts. The higher energy requirements during the early growth stages were observed with the higher metabolites of glycolysis and the TCA cycle. Further, the trade-off between primary and secondary sulfur-containing metabolites was observed, which may explain the differential GLS content in different growth stages.

Paclobutrazol improves surface water use efficiency by regulating allometric trait behavior in maize.

Paclobutrazol (PBZ) role in drought management of maize is least understood. In maize, root traits are linked with surface water management. Over three years, early and terminal deficit irrigation (EDI and TDI) with or without PBZ were imposed on DKC-9144 and PG-2475 maize varieties. Several allometric parameters viz. stem height, stem diameter, leaf area and root traits along with physiological processes were measured. Implication of these parameters in the management of soil surface irrigation in terms of water use efficiency (WUE) was demonstrated in maize. Increased number of lateral roots and root number density in DKC-9144 provided more surface area for water absorption for better management of EDI. Root growth rates showed a similar pattern with root length, root surface areas, and root numbers in EDI. Elevated expressions of ZmRTCL, ZmRTCS and ZmARF34 in EDI and EDI plus PBZ were associated with seminal roots and root laterals initiation. Under TDI alone or in combination with PBZ, root lengths (BRL, CRL, SRL) and root surface areas varied in DKC-9144 and PG-2475 over control. Furthermore, correlation analysis showed that decrease in WUE under TDI was significantly associated with a reduction in stem thickness and leaf surface area. For WUE_N in TDI and PBZ plus TDI, structural equation modelling proposed, brace root surface area (BRSA_N) as a positive contributor, while a negative contributor was seminal root surface area (SRSA_N). Present study explained the importance of specific root traits and their association with other allometric parameters for improving WUE in DKC-9144 variety of maize and the crop in general.

Nickel excess affects phenology and reproductive attributes of Asterella wallichiana and Plagiochasma appendiculatum growing in natural habitats.

Bryophytes are potent metal absorbers, thriving well on heavy metal (HM)-polluted soils. Mechanisms controlling uptake, compartmentalization and impacts of HMs on bryophytes life cycle are largely unknown. The current study is an effort to decipher mechanisms of nickel (Ni) excess-induced effects on the phenological events of two bryophytes, Asterella wallichiana and Plagiochasma apendiculatum growing in natural habitats. Observations revealed Ni-excess induced negative impacts on abundance, frequency of occurrence of reproductive organs, population viability and morphological traits, spore viability and physiological attributes of both the liverworts. Results led us conclude that P. appendiculatum survived better with the lowest impact on its life cycle events than A. wallichiana under Ni excess in natural habitats. Our findings collectively provide insights into the previously unknown mechanisms of Ni-induced responses in liverworts with respect to phenological attributes, as well as demonstrate the potential of P. appendiculatum to survive better in Ni excess habitats.

Effect of temperature and insect herbivory on the regulation of glucosinolate-myrosinase system in Lepidium latifolium.

The glucosinolate-myrosinase (GLS-MYR) system is an important component of plant-insect interactions. However, there is no report on its performance in field conditions where the plants are subjected to both abiotic and biotic pressures simultaneously. We investigated the GLS-MYR system in a Himalayan ecotype of Lepidium latifolium that is recognized for its adaptive potential in field conditions. In order to understand the independent contribution of temperature and Pieris brassicae herbivory on the components of the GLS-MYR system, different conditions were simulated in the growth chamber. During field conditions, the final GLS hydrolysis products were found to be regulated by the metabolic GLS levels, the temperature conditions, and the density of insect interactions. These factors influence the expression of the hydrolyzing and specifier proteins, which further affects the GLS hydrolysis products. Our results suggest that the production of hydrolysis products is differentially affected under field conditions. While allyl isothiocyanate is significantly (P ≤ 0.05) affected by temperature but not insect density, 1-cyano-2,3-epithiopropane is not affected by either. The study shows that the outcome of the GLS-MYR system in a plant is a consequence of the combinatorial effect of ecophysiological factors and the insect interactions that eventually decide the performance of a plant in an environment.

GC-MS based metabolomic approach to understand nutraceutical potential of Cannabis seeds from two different environments.

Cannabis sativa L. is a valuable plant that has regained its importance for medicinal use. Gas chromatography-mass spectrometry based untargeted metabolomic study was conducted in seeds of two accessions from different environments. A total of 236 metabolites were observed, and 43 metabolites were found differentially significant (p ≤ 0.05) in both the accessions. Based on the qualitative and quantitative accumulation of the nutraceutically important amino acids, cannabinoids, alkaloids, and fatty acids, the high altitude temperate Himalayan accession (CAN2) was found to have an advantage over the low altitude subtropical accession (CAN1). Seed oil from CAN2 showed the exclusive presence of linoleic acid and α- linolenic acid. The reducing power and DNA nicking assay on the methanolic extracts suggested higher antioxidant and nutraceutical potential in CAN2 and corroborated with the metabolic content of phenols and flavonoids. The environmental effect on the antioxidant and nutraceutical value in seeds is further discussed.

Photochemical efficiency is negatively correlated with the Δ9- tetrahydrocannabinol content in Cannabis sativa L.

Cannabis sativa L is an important plant, which is a source of durable fibers, nutritious seeds, and medicinally important phytocannabinoids including Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD). Light has shown to be a key modulator of biomass and cannabinoid yield suggesting responsive photochemical machinery. The present study was envisaged to understand the effect of the increasing levels of metabolic THC on the photochemical efficiency in Cannabis. The chlorophyll a fluorescence kinetics, photosynthetic pigments and immuno-detection of the photosynthetic machinery was analyzed on seven accessions from different environments, in conjunction with the cannabinoid content. All the accessions were clearly divided into three groups based on their relative content of CBD and THC. Group I with (CBD/THC > 1) had a clear advantage in terms of the damage to the D1, RbCL and Lhc1 protein holo-complex. Performance indicators of photochemistry based on the OJIP kinetics suggested a stoichiometrically negative correlation with the THC content. Zeaxanthin-dependent quenching is primarily responsible for lower NPQ in Group III with high THC content (THC > 6%). The THC treatment on Arabidopsis thaliana also suggested dose-dependent decrease in the photochemical efficiency suggesting the exclusivity of THC in causing the response. This resulted in the damage of photosynthetic machinery and the generation of free radicals, thereby compromising the yield. The study also opens a new screening method for Cannabis, based on cannabinoid content.

Physiological, Biochemical and Reproductive Studies on Valeriana wallichii, a Critically Endangered Medicinal Plant of the Himalayan Region Grown under In-Situ and Ex-Situ Conditions.

Valeriana wallichii, a perennial herb belonging to family Valerianaceae, is an important medicinal herb of the Himalayan region. The incessant exploitation of nature for meeting the demands of the pharmaceutical industry has put unbearable pressure on its natural habitats. A study on its physiological, biochemical, growth and reproductive attributes was planned. Physiological study revealed that ex-situ (outside their natural habitat) populations faced severe stress as compared to in-situ (natural habitat) plants. The difference in the performance of these habitat plants was related to superoxide and H2O2 in the leaves. Photosynthetic attributes were increased in in-situ populations. Proline content and its biosynthetic enzymes ornithine aminotransferase, and pyrroline-5-carboxylate reductase showed an increase in ex-situ plants; proline oxidase decreased. Glucose-6-phosphate dehydrogenase, shikimic acid dehydrogenese, phenylalanine lyase, and flavonoids content showed an increment in ex-situ plants. Antioxidants enzyme superoxide dismutase, catalase, ascorbate peroxidase and reduced glutathione showed an increment in ex-situ conditions. Growth and reproductive attributes were more in ex-situ plants. The observations made are suggestive that a comprehensive conservation programme involving in-situ as well as ex-situ strategies will be effective for the conservation and long term survival of the species.

Data set of in-silico analysis and 3D modelling of boiling stable stress-responsive protein from drought tolerant wheat.

Boiling stable proteins are widespread, evolutionary conserved proteins from several kingdoms including plants, fungi and bacteria. Accumulation evidences in response to dehydration, suggest a wide spread adaptation and an evolutionary role of these protein families to protect cellular structures from water loss effects in a wide range of water potentials. Boiling stable proteins, although represents just 0.1% of total plant proteins, resist coagulation upon boiling and believed to be involved in water stress adaptation in plants. The present data profiles in-silico analysis of cloned boiling stable protein encoding gene wBsSRP from drought tolerant cultivar of wheat. The data presented here was of a gene isolated from total RNA/mRNA samples of wheat variety PBW 175 subjected to drought stress. The gene is available with EMBL data repository with accession number LN832556.

Myrosinase: insights on structural, catalytic, regulatory, and environmental interactions.

Glucosinolate-myrosinase is a substrate-enzyme defense mechanism present in Brassica crops. This binary system provides the plant with an efficient system against herbivores and pathogens. For humans, it is well known for its anti-carcinogenic, anti-inflammatory, immunomodulatory, anti-bacterial, cardio-protective, and central nervous system protective activities. Glucosinolate and myrosinase are spatially present in different cells that upon tissue disruption come together and result in the formation of a variety of hydrolysis products with diverse physicochemical and biological properties. The myrosinase-catalyzed reaction starts with cleavage of the thioglucosidic linkage resulting in release of a D-glucose and an unstable thiohydroximate-O-sulfate. The outcome of this thiohydroximate-O-sulfate has been shown to depend on the structure of the glucosinolate side chain, the presence of supplementary proteins known as specifier proteins and/or on the physiochemical condition. Myrosinase was first reported in mustard seed during 1939 as a protein responsible for release of essential oil. Until this date, myrosinases have been characterized from more than 20 species of Brassica, cabbage aphid, and many bacteria residing in the human intestine. All the plant myrosinases are reported to be activated by ascorbic acid while aphid and bacterial myrosinases are found to be either neutral or inhibited. Myrosinase catalyzes hydrolysis of the S-glycosyl bond, O-ß glycosyl bond, and O-glycosyl bond. This review summarizes information on myrosinase, an essential component of this binary system, including its structural and molecular properties, mechanism of action, and its regulation and will be beneficial for the research going on the understanding and betterment of the glucosinolate-myrosinase system from an ecological and nutraceutical perspective.

Molecular cloning, characterization, heterologous expression and in-silico analysis of disordered boiling soluble stress-responsive wBsSRP protein from drought tolerant wheat cv.PBW 175.

The structural and physico-chemical properties that account for the multi-functionality of dehydrins remain largely unknown. In this study, we identified, sequenced and cloned a stress regulated cDNA encoding a dehydrin-like boiling stable protein (designated as wBsSRP; wheat boiling stable stress responsive protein) from drought stressed seedlings of drought tolerant cultivar of wheat (PBW 175). qRT-PCR analysis documented high transcripts levels of wBsSRP during drought and cold conditions in the tolerant cv. PBW 175 as a part of adaptive response to stress while the levels were significantly lower in the sensitive cv. PBW 343. We also describe in-silico characterization and molecular modeling of wBsSRP through homology search, motif analysis, secondary structure prediction, active site prediction and 3D structure analysis. The physico-chemical properties and theoretical data of wBsSRP depicts that it is a canonical group 2 LEA protein. The recombinant wBsSRP protein when expressed in E. coli detected a specific differential band (∼11 kDa) on SDS- PAGE after IPTG induction. The functional analysis of wBsSRP in E. coli revealed that wBsSRP is essential for the survival of E. coli as well as for maintaining bacterial growth under various stress conditions. In vitro peroxidase protection assay during heat stress (50 and 100 °C) showed that in the presence of wBsSRP, peroxidase activity was significantly retained and/or increased. Based upon the findings, it is suggested that wBsSRP accentuated the effects of stress by acting as a protectant and by the stabilization of membranes, thereby contributing to the improved stress tolerance of the recombinant E. coli under various abiotic stress conditions. We suggest that these findings might provide the rationale for the mechanism of how these proteins obviate the adverse effects of dehydration stress.

Dynamics of glucosinolate-myrosinase system during Plutella xylostella interaction to a novel host Lepidium latifolium L.

Plutella xylostella L. is a notorious pest of cruciferous crops causing worldwide losses of $4-5 billion per year. Developing classical biological control to this pest include an introduction of host plants that act as natural enemies showing deviation from the preference-performance regimen in the evolutionary ecology of plant-insect interactions. The present study was designed to understand the role of glucosinolate-myrosinase system during P. xylostella interactions with a novel host. Adult moth preference and larval performance study were conducted on a novel host Lepidium latifolium L. (LL) that has high sinigrin content and was compared with its laboratory host Arabidopsis thaliana (AT). The glucosinolate-myrosinase system was studied in a time course experiment during larval feeding in choice and no-choice experiments. Adult moths visit and prefers LL over AT for oviposition. Conversely, LL leaves were not preferred and proved detrimental for P. xylostella larvae. Aliphatic and indolic glucosinolates were found to decrease significantly (p≤0.05) in AT during initial 12h of P. xylostella challenge, whereas, they were not affected in LL. Also, MYB transcription factor expression and myrosinase activity in LL do not suggest a typical host response to a specialist insect. This preference-performance mismatch of P. xylostella on LL mediated by glucosinolate pattern suggests that this novel plant could be utilized in P. xylostella management.

Two-stage phyto-microremediation of tannery effluent by Spirodela polyrrhiza (L.) Schleid. and chromium resistant bacteria.

Two-stage sequential treatment of tannery effluent was conducted employing a wetland plant, Spirodela polyrrhiza (L.) Schleid., and chromium (Cr) resistant bacterial strains. The bacterial strains were isolated from Cr-enriched environmental matrices and rhizosphere of Spirodela polyrrhiza. The phyto-rhizoremediation of tannery effluent by Spirodela and its rhizospheric bacteria (Cellulomonas biazotea APBR1-6, Bacillus safensis APBR2-12, Staphylococcus warneri APBR3-5, Microbacterium oleivorans APBR2-6), followed by microremediation by Cr resistant bacteria (Micrococcus luteus APBS5-1, Bacillus pumilus APBS5-2, Bacillus flexus APBE3-1, Virgibacillus sediminis APBS6-1) resulted in reduction of pollution parameters [COD (81.2%), total Cr (97.3%), Cr(VI) (99.3%), Pb(II) (97.0%), Ni (95.7%)]. The LC-MS analysis showed that many pollutants detected in untreated tannery effluent were diminished after bioremediation or long chains of alcohol polyethoxylates viz. C18EO6 in untreated effluent were broken down into smaller unit of alcohol polyethoxylate ((+)HHO[CH2CH2O]H), indicating that bacteria and Spirodela polyrrhiza, alongwith its rhizospheric associates utilized them as carbon and energy source.

Physiological and biochemical plasticity of Lepidium latifolium as 'sleeper weed' in Western Himalayas.

To understand the spread of native populations of Lepidium latifolium growing in different altitudes in Ladakh region of Western Himalayas, photosynthetic and fluorescence characteristics were evaluated in relation to their micro-environment. Three sites representing sparsely populated (SPS), moderately populated (MPS) and densely populated site (DPS) were selected. Results showed that the DPS had higher photosynthetic accumulation than MPS and SPS. The higher transpiration rate at DPS despite lower vapor pressure deficit and higher relative humidity suggest the regulation of its leaf temperature by evaporative cooling. Intrinsic soil parameters such as water holding capacity and nutrient availability also play crucial role in higher biomass here. The quantum efficiency of PSII photochemistry (Fv/Fm, non-photochemical quenching (NPQ), ΦPSII) and light curve at various PPFDs suggests better light harvesting potential and light compensation point at DPS than the other two sites. Concomitantly, plants at SPS had significantly higher lipid peroxidation, suggesting a stressful environment, and higher induction of antioxidative enzymes. Metabolic content of reduced glutathione also suggests an efficient mechanism in DPS and MPS than SPS. High light intensities at MPS are managed by specialized contrive of carotenoid pigments and PsbS gene product. Large pool of violaxanthin and lutein plays an important role in this response. It is suggested that L. latifolium is present as 'sleeper weed' that has inherent biochemical plasticity involving multiple processes in Western Himalayas. Its potential spread is linked to site-specific micro-environment, whereby, it prefers flat valley bottoms with alluvial fills having high water availability, and has little or no altitudinal effect.

Purification and Characterization of a Novel Redox-Regulated Isoform of Myrosinase (ß-Thioglucoside Glucohydrolase) from Lepidium latifolium L.

Myrosinase (ExPASy entry EC 3.2.1.147) is involved in the hydrolysis of glucosinolates to isothiocyanates, nitriles, and thiocyanates that are responsible for various ecological and health benefits. Myrosinase was purified from the leaves of Lepidium latifolium, a high-altitude plant, to homogeneity in a three-step purification process. Purified enzyme exists as dimer in native form (∼160 kDa) with a subunit size of ∼70 kDa. The enzyme exhibited maximum activity at pH 6.0 and 50 °C. With sinigrin as substrate, the enzyme showed Km and Vmax values of 171 ± 23 µM and 0.302 µmol min(-1) mg(-1), respectively. The enzyme was found to be redox-regulated, with an increase in Vmax and Kcat in the presence of GSH. Reduced forms of the enzyme were found to be more active. This thiol-regulated kinetic behavior of myrosinase signifies enzyme's strategy to fine-tune its activity in different redox environments, thus regulating its biological effects.

Comparative studies for screening of bioactive constituents from various parts of Incarvillea emodi.

A comparative study was performed on various parts (shoots, roots and flowers) of Incarvillea emodi. The alcoholic extracts of different parts were fractionated with solvents of different polarity and studied for the determination of total polyphenol content and total antioxidant potential. Furthermore, we have isolated major iridoid glucosides from the dried flowers of I. emodi followed by the comparative cytotoxicity studies of these iridoids against five different human cancer cell lines. The results have demonstrated that ethyl acetate fraction of all parts have higher phenolic content (167.87-294.31 mg/g as gallic acid equivalent) and higher total antioxidant potential (252.95-384.64 mg/g as trolox equivalent). The results of in vitro cytotoxicity studies have indicated that boschnaloside (2) possesses promising anticancer potential against three human cancer cell lines, THP-1, A-549 and PC-3, which belong to leukaemia, lung and prostate cancers, respectively, while plantarenaloside (1) expressed relevant cytotoxic activity against THP-1 cell lines of leukaemia.

Evaluation of nutritional and antioxidant status of Lepidium latifolium Linn.: a novel phytofood from Ladakh.

Lepidium latifolium Linn. (perennial pepperweed) is one of the preferred phytofoods among cold arid region of Ladakh, India and its leaves contribute significantly to people's diet. This study was conducted to determine its nutritive value and antioxidant activity. Plant samples from three different locations were selected in the present study. Results showed that this plant is an excellent source of glucosinolates, notably sinigrin that is present in very high amount (∼70-90%). Its value ranged from 149 to 199 µg per g fresh weight. Fatty acid composition analysis showed that its leaves were abundant in unsaturated fatty acids, specifically linolenic acid (18∶3) whose percentage is about 50%. Higher glucose and crude protein along with higher nitrogen to sulfur ratio, supplements the nutritive value of this plant. Based on total phenol, flavanoids, free radical scavenging activity and DNA protective activity showed that this ecotype of perennial pepperweed contains high antioxidant properties. The percentage inhibition for O2(-) scavenging activity ranged from 41.3% to 83.9%. Higher content of phenols (26.89 to 50.51 mg gallic acid equivalents per g dry weight) and flavanoids (38.66 to 76.00 mg quercetin equivalents per g dry weight) in leaves could be responsible for the free radical scavenging activity of this plant. Depending upon the location of the plants, variations were observed in different activities. Based on the systematic evaluation in this study, preparations of Lepidium latifolium from Ladakh can be promoted as substitute to dietary requirements.

Antioxidant activity and protective effect against plasmid DNA strand scission of leaf, bark, and heartwood extracts from Acacia catechu.

UNLABELLED: The antioxidant activity of methanol extract/fractions of leaf, bark, and heartwood of Acacia catechu was evaluated by various antioxidant assays, including free radical, superoxide and hydroxyl radical, reducing power, metal ion chelation, as well as hydroxyl radical induced DNA strand scission. The leaf, bark, and heartwood powder was extracted in methanol and the lyophilized methanol extract was fractionated with different solvents in the order of increasing polarity. The results indicate that ethyl acetate fraction of heartwood has the highest antioxidant capacities, presenting lower EC(50) values particularly in free radical scavenging activity, including DPPH radicals (4.76 ± 0.14 µg/mL), superoxide anions (26.21 ± 0.79 µg/mL), and hydroxyl radicals (33.69 ± 1.42 µg/mL), in direct assay systems. Reducing power was also highest in ethyl acetate fraction of heartwood (EC(50) of 79.05 ± 1.02 µg/mL). As for the chelating power on ferrous ions, leaf extract was more effective than bark and heartwood extracts. Furthermore, the ethyl acetate and acetone fractions of heartwood significantly protected pBR322 supercoiled plasmid DNA against strand scission induced by hydroxyl radicals in a Fenton's reaction mixture. PRACTICAL APPLICATION: The present investigation suggests that the three organs of A. catechu differ significantly in their antioxidant potential as seen in the DPPH radical scavenging assay, reducing power assay, metal ion chelating assay, superoxide radical scavenging assay and hydroxyl radical scavenging assay. Further, our results showed that crude methanol extract and ethyl acetate fraction of heartwood of A. catechu might have a good potential as a source for natural health products due to its antioxidant and DNA protective activities.

Tea (Camellia sinensis) clones with shorter periods of winter dormancy exhibit lower accumulation of reactive oxygen species.

Tea (Camellia sinensis (L.) O. Kuntze) is a perennial crop grown throughout the world. During winter, tea undergoes a dormancy period when growth of apical buds almost ceases, severely reducing the commercial yield of tea. Low temperatures prevail during the period of winter dormancy, which alone or in combination with high solar irradiance have the potential to induce oxidative stress in plants. We studied six tea clones under field conditions to test whether a relationship exists between oxidative stress and winter dormancy. Data on the behavior of the enzymatic antioxidative system was collected for all clones during different phases of winter dormancy. There was a strong positive correlation among clones between accumulation of reactive oxygen species (ROS) and the length of the dormancy period. Clones having shorter dormancy periods exhibited higher induction of antioxidative enzymes. Results suggest that efficient scavenging of ROS is a desirable feature in tea because it leads to lower accumulations of ROS during winter months and is associated with reduced winter dormancy.

Plants at high altitude exhibit higher component of alternative respiration.

Total respiration, capacities of cytochrome (CytR) and alternative respiration (AR) were studied in two varieties of barley (Horedum vulgare) and wheat (Triticum aestivum) each and one variety of pea (Pisum sativum) at low (Palampur; 1300 m) and high altitudes (Kibber; 4200 m). Similar studies were carried out in naturally growing Rumex nepalensis and Trifoilum repenses at Palampur, Palchan (2250 m) and Marhi (3250 m). All the plants species exhibited lower CytR but significantly higher AR capacity at high altitude (HA) (72-1117% higher) as compared to those at low altitude (LA). Glycolytic product, pyruvate and tricarboxylic acid cycle intermediate, citrate increased with increase in altitude. While the role of these metabolites in relation to HA biology is discussed, significantly higher AR at HA is proposed to be an adaptive mechanism against the metabolic perturbations wherein it might act to lower reactive oxygen species and also provides metabolic homeostasis to plants under the environment of HA.

Tea (Camellia sinensis (L.) O. Kuntze) clone with lower period of winter dormancy exhibits lesser cellular damage in response to low temperature.

There is no literature available on the response of tea plant to low temperature. We studied the effect of low temperature on two clones of tea with contrasting periods of winter dormancy, a phenomenon in which the growth of apical shoots of tea is diminished during winter months. Clone 'Teenali 17/154' (TNL) showed shorter periods of winter dormancy than clone 'Kangra Jat' (KNJ). Low temperature (5 degrees C) resulted in increase of metabolic superoxide (O2*-) content and cellular damage (as measured by tetrazolium chloride reduction test) in both the clones, however, the increase was lesser in the case of TNL compared to KNJ. Activities of superoxide dismutase (SOD; EC 1.15.1.1), ascorbate peroxidase (APX; EC 1.11.1.11) and glutathione reductase (GR; EC 1.6.4.2) increased in both the clones in response to low temperature however, GR activity exhibited significant differences (P < 0.05) between the two clones. Low temperature caused increase in the intensity of various isozymes of SOD, APX and GR. A new isozyme of SOD (Cu/Zn type) was induced in both the clones at low temperature. Significantly higher GR activity in both the clones suggested a role of this enzyme in imparting better protection to tea at low temperature. Also, clonal variation for GR isozyme was observed between the clones. Based on these results it appears that TNL, a clone with relatively lesser period of winter dormancy experiences lesser oxidative stress in response to low temperature compared to KNJ, a clone with relatively higher period of winter dormancy.