Aqueous extract of large cardamom inhibits vascular damage, oxidative stress, and metabolic changes in fructose-fed hypertensive rats.

PURPOSE: Since metabolic abnormalities such as elevated glucose level and imbalanced lipid profiles increase the risk for hypertension and cause endothelial dysfunction, we evaluated the effect of aqueous extract of large cardamom (AELC) on fructose-induced metabolic hypertension and oxidative stress. METHODS: The male Sprague-Dawley rats were divided into 6 groups with 5 rats in each group, and each group was fed with 10% fructose in drinking water for 8 weeks. Starting from week 5, animals were treated with 50, 100, and 200 mg/kg/day AELC or Losartan (10 mg/kg/day). Systolic, diastolic, and mean arterial blood pressure was measured once in every seven days using the tail-cuff method. Vascular function, plasma nitric oxide (NO), glucose, lipid profiles, serum biochemical, and anti-oxidant parameters were also evaluated. RESULTS: Rats fed with fructose showed higher blood pressure, serum cholesterol, and triglyceride levels, but decreased in the AELC or Losartan treatment group. Treatments with AELC prevented exaggerated plasma glucose and oxidative stress and restored the nitric oxide level in fructose-fed rats. Besides, it also reduced vascular proliferation and improved the relaxation response of acetylcholine in the aorta pre-contracted with phenylephrine. CONCLUSION: In summary, the obtained results suggest that AELC can prevent and reverse the high blood pressure induced by fructose, probably by restoring nitric oxide level and by improving altered metabolic parameters.

Antioxidant and vasorelaxant effects of aqueous extract of large cardamom in L-NAME induced hypertensive rats.

PURPOSE: The present work aimed to study the effect of aqueous extract of large cardamom (AELC) to prevent vascular remodeling and oxidative stress in Nω-Nitro-L-arginine methyl ester (L-NAME)-induced hypertension. METHOD: Male Wistar rats were administered with L-NAME 40 mg/kg/day for 28 days by oral gavage. The treatments included captopril (20 mg/kg/day) or AELC (100, 200 and 400 mg/kg/day) along with L-NAME administration. RESULTS: L-NAME treated rats showed high systolic, diastolic and mean arterial pressure, decreased nitric oxide level, increased level of malondialdehyde in plasma, heart, aorta and kidney, hypertrophy of the vascular wall and reduced vascular response to acetylcholine in phenylephrine-precontracted aorta. Treatment with AELC markedly reduced the blood pressure, restored the nitric oxide level, reduced the malondialdehyde level, alleviated the hypertrophy in L-NAME-induced hypertensive rats. Additionally, it also improved the vascular response to acetylcholine in phenylephrine pre-contracted aorta. CONCLUSION: In conclusion, our results demonstrate the preventive effect of AELC in L-NAME-induced hypertensive model, which is possibly related to antioxidant activities and restoration of nitric oxide level.

Exploring the effects of vasoactive constituents in large cardamom: implications for the anti-hypertensive effect via eNOS coupling pathway - an in-vitro study in rat endothelial cells.

Endothelial dysfunction, linked to reduced eNOS expression and nitric oxide (NO) availability, contributes to cardiovascular diseases (CVDs). Large cardamom exhibits antihypertensive effects by augmenting NO levels and antioxidant activity. To decipher its mechanisms, selected constituents were docked with eNOS-associated target genes such as GTP cyclohydrolase I (GTPCH-1) and (dihydrofolate reductase [DHFR]). Endothelial damage induced by L-NAME and fructose was countered by assessing nitric oxide metabolites (NOx), tetrahydrobipterin (BH4 levels), GCH-I expression and super oxide dismutase (SOD) activity after constituent incubation. Cyanidin-3-O-glucoside and petunidin-3-O-glucoside notably restored impaired vascular markers in both models. These phytoconstituents are likely to activate GCH-BH4-eNOS pathways, upregulating SOD and NO expression, maintaining endothelial integrity. Large cardamom's antihypertensive effects may stem from these components, synergistically enhancing endothelial NO release via the eNOS pathway.

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, ß-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, ß-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.

Large Cardamom Extract Enhances Ramipril's Vasoprotective Action in the Aorta by Modulating Endothelial Redox Biology. An Evaluation based on In-silico and In-vitro Research.

BACKGROUND: The mechanisms that cause a patient's blood pressure to rise are diverse. Controlling blood pressure with monotherapy acting through a single pathway may be unachievable. Combining a clinically used medication with herbal medicine can result in an antihypertensive effect that is two to five times greater than monotherapy. METHOD: This study examined the effects of aqueous extracts of large cardamom and ramipril on the redox biology of nitric oxide and vascular reactivity in the isolated aorta incubated with a nitro-L-arginine methyl ester. Molecular docking study was performed to predict the affinity of constituents of large cardamom extracts with the NOX 2 gene. RESULTS: Nitric oxide (NO) levels, disordered antioxidant enzymes (glutathione and catalase), NADPH oxidase and lipid peroxidation were recovered when aqueous extract of large cardamom and ramipril were combined. A gradual increase in the percentage relaxation of acetylcholine in phenylephrine pre-contracted aorta indicates that the combination therapy prevents endothelial damage. The molecular docking study reveals the important phytoconstituents present in the large cardamom that can effectively bind with the NADPH oxidase for its antioxidant activity. Consculsion: According to our findings, it was evidenced that the large cardamom extract's vasoprotective action was mostly related to its ability to restore endothelial redox biology by suppressing NADPH oxidase activity. Our findings suggest that ramipril's direct impact on the eNOS/NO system, along with the antioxidant properties of AELC, could have a synergetic benefit in the treatment of hypertension, as well as lessen ramipril's existing side effects.

In Silico Analysis of the Apoptotic and HPV Inhibitory Roles of Some Selected Phytochemicals Detected from the Rhizomes of Greater Cardamom.

Occurrence of cervical cancer, caused due to persistent human papilloma virus (HPV) infection, is common in women of developing countries. As the conventional treatments are expensive and associated with severe side effects, there is a need to find safer alternatives, which is affordable and less toxic to the healthy human cells. Present study aimed to evaluate the anti-HPV and apoptotic potential of four compounds from the greater cardamom (Amomum subulatum Roxb. var. Golsey), namely rhein, phytosphingosine, n-hexadecenoic acid and coronarin E. Their anti-HPV and apoptotic potential were studied against viral E6, E7 and few anti-apoptotic proteins of host cell (BCL2, XIAP, LIVIN) by in silico docking technique. Phytochemicals from the plant extract were analysed and identified by LC/MS and GC/MS. Involvement of the target proteins in various biological pathways was determined through KEGG. Structural optimization of the three-dimensional structures of the ligands (four phytochemicals and control drug) was done by Avogadro1.1. Receptor protein models were built using ProMod3 and other advanced tools. Pharmacophore modelling of the selected phytochemicals was performed in ZINCPharmer. Swiss ADME studies were undertaken to determine drug likeness. The ligands and proteins were digitally docked in DockThor docking program. Protein flexibility-molecular dynamic simulation helped to study protein-ligand stability in real time. Finally, the correlation of evaluated molecules was studied by the use of principal component analysis (PCA) based on the docking scores. All the ligands were found to possess apoptotic and anti-cancer activities and did not violate Lipinsky criteria. n-Hexadecanoic acid and its analogues showed maximum efficacy against the target proteins. All the protein-ligand interactions were found to be stable. The uncommon phytochemicals identified from rhizomes of greater cardamom have anti-cancer, apoptotic and HPV inhibitory potentials as analysed by docking and other in silico studies, which can be utilized in drug development after proper experimental validation.

A Simplified Multiplex PCR Assay for Simultaneous Detection of Six Viruses Infecting Diverse Chilli Species in India and Its Application in Field Diagnosis.

Chilli is infected by at least 65 viruses globally, with a mixed infection of multiple viruses leading to severe losses being a common occurrence. A simple diagnostic procedure that can identify multiple viruses at once is required to track their spread, initiate management measures and manage them using virus-free planting supplies. The present study, for the first time, reports a simplified and robust multiplex PCR (mPCR) assay for the simultaneous detection of five RNA viruses, capsicum chlorosis orthotospovirus (CaCV), chilli veinal mottle virus (ChiVMV), large cardamom chirke virus (LCCV), cucumber mosaic virus (CMV), and pepper mild mottle virus (PMMoV), and a DNA virus, chilli leaf curl virus (ChiLCV) infecting chilli. The developed mPCR employed six pairs of primer from the conserved coat protein (CP) region of the respective viruses. Different parameters viz., primer concentration (150-450 nM) and annealing temperature (50 °C), were optimized in order to achieve specific and sensitive amplification of the target viruses in a single reaction tube. The detection limit of the mPCR assay was 5.00 pg/µL to simultaneously detect all the target viruses in a single reaction, indicating a sufficient sensitivity of the developed assay. The developed assay showed high specificity and showed no cross-amplification. The multiplex PCR assay was validated using field samples collected across Northeast India. Interestingly, out of 61 samples collected across the northeastern states, only 22 samples (36%) were positive for single virus infection while 33 samples (54%) were positive for three or more viruses tested in mPCR, showing the widespread occurrence of mixed infection under field conditions. To the best of our knowledge, this is the first report on the development and field validation of the mPCR assay for six chilli viruses and will have application in routine virus indexing and virus management.

Data on large cardamom transcriptome associated with Chirke disease.

Large cardamom (Amomum subulatum Roxburg), is an ancient spice native to North-Eastern India and Southeast Asia, which belongs to the family Zingiberaceae under the order Scitaminae. Large cardamom is mostly affected by a viral disease termed Chirke caused by Large Cardamom Chirke Virus (LCCV). These disease has spread due to drastic changes in the ecosystem, inadequate rain in dry months and absence of good agricultural practices by the farmers resulting in aphid infestations. In the present study, using HiSeq™ 2000 RNA sequencing technology transcriptome sequencing was performed for both control (disease not expressed) and diseased large cardamom leaf tissues. RNA-seq generated 77260968 (7.72 GB) and 72239708 (7.22 GB) paired raw reads for large cardamom control and diseased samples respectively. The raw data were submitted to the NCBI SRA database under the accession numbers SRX2529373 and SRX2529372 and the assembled transcriptomes were submitted to TSA under the accession numbers GIAV01000000 and GIAW01000000 for the control and diseased samples respectively. The raw reads were quality trimmed and assembled de novo using TRINITY assembler which created 156822 (control) and 148953 (diseased) contigs with N50 values 2107 (control) and 2182 (diseased). The data were used to identify the significantly differentially expressed genes between control and diseased samples.

Field-usable lateral flow immunoassay for the rapid detection of a macluravirus, large cardamom chirke virus.

A simple and rapid lateral flow immunoassay (LFIA) was developed by utilizing gold nanoparticles conjugated to a polyclonal antibody against coat protein of large cardamom chirke virus (LCCV). The LFIA based on the principle of sandwich immunoassay detected LCCV within ∼10 min and the result could be evaluated visually. The colloidal gold (CG) was made using 1% gold chloride solution. The LCCV IgG (1 µg/µl) and Mouse IgG (0.5 µg/µl) were conjugated with CG individually and coated onto a conjugate pad at 1:1 ratio. A sample extraction procedure was optimized in order to get adequate clear leaf sap of large cardamom leaf within few minutes. The sensitivity limit of the detection was 1:40 dilution of LCCV infected leaf sap. The diagnostic performance of LFIA was compared with ELISA using field samples. The LFIA was free from false positive as no visible test line was developed with healthy and potyviruses such as papaya ringspot virus and potato virus Y. The diagnostic specificity and sensitivity of LFIA was 100% and 90%, respectively. The Cohen's kappa coefficient (0.701) suggested a very good agreement between the ELISA and LFIA. Receiver operating characteristic analysis indicated that LFIA was a robust method as the area under the curve (0.950) is significantly (P <0.0001) broader.

Nine novel DNA components associated with the foorkey disease of large cardamom: evidence of a distinct babuvirus species in Nanoviridae.

Foorkey disease is a serious constraint to the production of large cardamom (Amomum subulatum, family Zingiberaceae). The disease is characterized by profuse proliferation of excessive stunted shoots, which makes the clump totally unproductive. The disease has been known in India since 1936 but the complete genome of the virus had not yet been characterized. In a preliminary study, an associated virus tentatively named as Cardamom bushy dwarf virus (CBDV) was identified based on the partial sequence of a single DNA component (DNA-R). In the present study, a high incidence (37.2-39.3%) of foorkey was recorded in certain plantations in the Darjeeling hills located at lower altitudes (300-1380 m) and CBDV was detected in several field samples by PCR. Nine novel DNA components were isolated and characterized from foorkey affected plants. CBDV contained six major DNA components (DNA-R, -S, -M, -C, -N and -U3) similar to the integral genome components known for the members of the genus Babuvirus in the family Nanoviridae. Additional components, satellite Rep (DNA-sRep1) and unknown components (DNA-Uf1 and -Uf2) were also identified. The size of the genome components ranged from 1028 to 1127. The sequence identity and phylogeny based on the individual components as well as overall genome (59.8-62% identity) distinguished CBDV from the two existing babuvirus species, Banana bunchy top virus and Abaca bunchy top virus. CBDV is the first distinct babuvirus species that affects plant species outside family Musaceae. This study shows further diversity in the genus Babuvirus.

Highly efficient immunodiagnosis of Large cardamom chirke virus using the polyclonal antiserum against Escherichia coli expressed recombinant coat protein.

Large cardamom chirke virus (LCCV), genus Macluravirus, family Potyviridae is an important constrain in large cardamom production in India. Purification of LCCV from large cardamom tissues is difficult and therefore immunodiagnostic reagents are not available. In the present study, we have successfully expressed coat protein (CP) gene of LCCV in Escherichia coli. The purification of expressed protein by Ni-NTA affinity chromatography was inefficient due to precipitation of protein during renaturation. We have optimized a simple, inexpensive and efficient method for purification of the expressed CP through gel extraction with 5 % SDS followed by renaturation in Milli-Q water, which resulted in high yield (4.7 mg/ml) and good quality of the protein. A higher titer (1:256,000) polyclonal antibody (PAb) to the recombinant CP was produced, which strongly recognized LCCV in crude leaf extract and showed minimal background reaction with the healthy leaf extract in enzyme linked immunosorbent assay (ELISA) and dot immunobinding assay (DIBA). The sensitivities of the ELISA and DIBA were 5 and 0.1 ng of expressed protein, respectively. Both the ELISA and DIBA were validated with 100 % accuracy in detecting LCCV in field samples. The PAb differentiated Cardamom mosaic virus, another close relative of LCCV. Our study is first to report highly efficient immunodiagnosis with PAb to E. coli expressed recombinant CP of a virus under the genus Macluravirus. The antigen expression construct and PAb developed in the present study will be useful in production of virus free planting materials of large cardamom.