In vitro induction of tuber formation for the synthesis of secondary metabolites in Chlorophytum borivilianum Sant. et Fernand

Chlorophytum borivilianum Sant. et Fernand an endangered herb is valued for several medicinal properties in its tuberous roots. An efficient and reproducible method for inducing in vitro tubers from stem disc explant has been developed. Stem disc possessing shoot buds were induced to develop multiple shoots in Murashige and Skoog (MS) medium supplemented with vitamins, 3% sucrose, 0.8% agar and 5 mg/L benzylaminopurine (BAP). Healthy regenerated shoots were rooted in MS basal medium containing 3% sucrose (w/v), 0.8% agar supplemented with indole-3-acetic acid (2 mg/L). On further sub culturing, the maximum percentage of tuber formation was obtained in growth hormone free half (½) MS liquid media supplemented with vitamins and 1.5% sucrose after 8 to 9 weeks. The saponin contents of the in vitro and in vivo raised tubers were qualitatively and quantitatively analyzed by highperformance liquid chromatography (HPLC) and liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS). There was a significant similarity in the saponins in both tubers. The in vitro raised tubers showed similar high metabolite content than in vivo grown tubers which is required for medicinal applications. The rooted plantlets were transferred to peat and sand (2:1) with more than 80% success. This is the 1st report of in vitro tuber formation and secondary metabolites screening of C. borivilianum. This work will give a strong impetus to the pharmaceutical and neutraceutical sectors.Keywords: Chlorophytum borivilianum, tuber formation, saponin, liquid medium, high-performance liquid chromatography (HPLC)African Journal of Biotechnology Vol. 12(20), pp. 2900-2907

Development and standardisation of Laghu Sudarshan Churna – An Ayurvedic polyherbal formulation

106-112Laghu Sudarshan Churna, LSC is an Ayurvedic polyherbal formulation employed for different types of jvaras (fevers). The present study was undertaken to prepare its standardised formulation and to standardise the finished product using quality control procedures mentioned in Ayurvedic Pharmacopoeia of India (API). For this, four batches of the finished products were prepared on a laboratory scale and performed the pharmacognostical parameters (macroscopic, microscopic and powder drug analysis); thin layer chromatography; quantitative physicochemical evaluation including loss on drying, total ash, acid-insoluble ash, alcohol & water soluble extractive values, and pH; & measuring the level of aflatoxins, microbial load, heavy metals and pesticide residues of the finished product. This study is the foremost effort to develop the standardised formulation along with the evaluation parameters for LSC. Thus, obtained results would be beneficial and will act as the reference for the standardisation of LSC

MAK-4 and -5 supplemented diet inhibits liver carcinogenesis in mice

<p>Abstract</p> <p>Background</p> <p>Maharishi Amrit Kalash (MAK) is an herbal formulation composed of two herbal mixtures, MAK-4 and MAK-5. These preparations are part of a natural health care system from India, known as Maharishi Ayur-Veda. MAK-4 and MAK-5 are each composed of different herbs and are said to have maximum benefit when used in combination. This investigation evaluated the cancer inhibiting effects of MAK-4 and MAK-5, <it>in vitro </it>and <it>in vivo</it>.</p> <p>Methods</p> <p><it>In vitro </it>assays: Aqueous extracts of MAK-4 and MAK-5 were tested for effects on <it>ras </it>induced cell transformation in the Rat 6 cell line assessed by focus formation assay. <it>In vivo </it>assays: Urethane-treated mice were put on a standard pellet diet or a diet supplemented with MAK-4, MAK-5 or both. At 36 weeks, livers were examined for tumors, sera for oxygen radical absorbance capacity (ORAC), and liver homogenates for enzyme activities of glutathione peroxidase (GPX), glutathione-S-transferase (GST), and NAD(P)H: quinone reductase (QR). Liver fragments of MAK-fed mice were analyzed for connexin (cx) protein expression.</p> <p>Results</p> <p>MAK-5 and a combination of MAK-5 plus MAK-4, inhibited <it>ras</it>-induced cell transformation. In MAK-4, MAK-5 and MAK4+5-treated mice we observed a 35%, 27% and 46% reduction in the development of urethane-induced liver nodules respectively. MAK-4 and MAK4+5-treated mice had a significantly higher ORAC value (<it>P </it>< 0.05) compared to controls (200.2 ± 33.7 and 191.6 ± 32.2 <it>vs. </it>152.2 ± 15.7 ORAC units, respectively). The urethane-treated MAK-4, MAK-5 and MAK4+5-fed mice had significantly higher activities of liver cytosolic enzymes compared to the urethane-treated controls and to untreated mice: GPX(0.23 ± 0.08, 0.21 ± 0.05, 0.25 ± 0.04, 0.20 ± 0.05, 0.21 ± 0.03 U/mg protein, respectively), GST (2.0 ± 0.4, 2.0 ± 0.6, 2.1 ± 0.3, 1.7 ± 0.2, 1.7 ± 0.2 U/mg protein, respectively) and QR (0.13 ± 0.02, 0.12 ± 0.06, 0.15 ± 0.03, 0.1 ± 0.04, 0.11 ± 0.03 U/mg protein, respectively). Livers of MAK-treated mice showed a time-dependent increased expression of cx32.</p> <p>Conclusion</p> <p>Our results show that a MAK-supplemented diet inhibits liver carcinogenesis in urethane-treated mice. The prevention of excessive oxidative damage and the up-regulation of connexin expression are two of the possible effects of these products.</p

Establishment of the mechanism of purification and levigation of green chemistry-assisted biocomposites of red ochre (Gairika): synthesis, characterization, and antibacterial, prebiotic, antioxidant, and antacid activities of the traditional Ayurvedic medicine Laghu Sutashekhara Rasa

Gairika (red ochre) has a long history of influencing human civilization. Gairika is a rich source of nutrients used for reproductive and brain health. Gairika is mentioned as an antacid drug in Indian Ayurvedic medicine under Laghu Sutashekhara Rasa (LSR). However, a detailed study on LSR has not been reported to date. In the present study, LSR was prepared, and a pharmaceutical SOP (standardization procedure) was reported to obtain batch-to-batch reproducibility. LSR was characterized using FTIR, XRD, SEM-EDX, and TGA analyses. LSR was tested in vitro for its antacid activity. Advanced instrumentation revealed that LSR formation produced symmetrical particles (5–8 µm) with kaolin, kaolinite, quartz, goethite, and hematite, along with the phytoconstituents of Goghrita (clarified cow’s butter), Shunthi, and Nagawalli, as confirmed by GC-MS/MS analysis. The FTIR study revealed the formation of a chelating complex of goethite and hematite along with their phytoconstituents. XRD analysis confirmed the presence of kaolin, kaolinite, quartz, goethite, and hematite. Using in vitro antacid experiments, LSR and Shunthi demonstrated significant antacid activity as compared to antacid drugs and standards in the market, such as CaCO3. The DPPH assay revealed IC50 values of 12.16 ± 1.23 mg/mL, which is 0.0029 of Trolox-equivalent antioxidant activity. The inhibition (18 ± 4 mm) against pathogens (S. aureus, E. coli, P. aeruginosa, and B. subtilis) and the prominent growth of gut microbiota-supported strains (S. boulardii, L. paracasei, and L. plantarum) observed on LSR formulation were indicative of LSR application as a prebiotic. Here, the mechanism of purification and levigation mentioned in the classical literature of LSR was established. Overall, purification of Gairika with cow ghee and levigation with Nagawalli may enhance the solubility, bioavailability, and shelf-life of LSR through hydration and co-crystallization mechanisms. This is the first comprehensive report on the pharmaceutical validation of LSR and its characterization. The results of the present study could contribute to the development and reliable reproduction of LSR and the utility of environmental red ochre as a medicine in combination with Shunthi (Zingiber officinale Roxb.), as prescribed under Indian Ayurvedic medicine

Standardization of lyophilization medium for Streptococcus thermophilus subjected to viability escalation on freeze drying

The objective of the present study is to develop a lyophilization medium for <em>Streptococcus thermophilus</em> (NCIM 2904) as the industrial exploitation of this bacterium totally depends upon preservation and lyophilization protocols. Protective effect of 18 compounds were observed individually and in combinations with different sugars, sugar alcohols, polymers, protein concentrates and buffers. Among all the protectants tested, ammonium citrate (1% w/w), K2HPO4 (1% w/w) and KH2PO4 (1% w/w) provided lowest protection to these bacterial cells while 10% (w/w) sodium caseinate, whey protein concentrate, sweet whey powder, and skim milk showed significant results in viability escalation. Survival in carbon sources like lactose, sucrose and maltodextrine was also favored maximally. Combination of sodium caseinate 10%, skim milk 5%, sucrose 5%, lactose 5% and mono sodium glutamate 1% in distilled water in ratio of 1:5 with <em>S. thermophilus</em> showed survival percentage of 96%

Development of quality control parameters for classical ayurvedic formulation: Vyoshadi Churna

Introduction: Ayurvedic formulations have a long history of use and have increased amazingly to follow the global interest in natural products. However, for many classical ayurvedic formulations, no quality parameters of standardization are available. Vyoshadi Churna, a classical Ayurvedic polyherbal formulation that cures indigestion and diarrhea, is being used since ancient times. However, there is no standard method available for its formulation and evaluation. Hence, the current study aims to develop the quality standards for Vyoshadi churna. Methods: The standard procedures for preparation and standardization of Churna described in Ayurvedic Pharmacopoeia of India (API) were used in the present study. Standardization includes pharmacognostical, chromatographic, and physico-chemical studies. The other essential studies were also done in accordance to API and World Health Organization guidelines to find the limits of microbial load, aflatoxin, heavy metals, and pesticide residue using modern analytical techniques such as gas chromatography-mass spectroscopy (MS) and inductively coupled plasma-MS/MS. Results: Vyoshadi Churna is of brown color and bitter in taste. Pharmacognostical studies have shown the characteristic features of almost each contributory crude drug. A unique chemical profile was also developed using toluene: ethyl acetate: methanol: formic acid (4:4:1:0.5) as the mobile phase to resolve maximum components. Furthermore, the standardized limits for the physicochemical studies, microbial load, heavy metals, aflatoxins, and pesticide residues were established and found in limits. Conclusion: This is for the first time when the standard operating procedure of Vyoshadi Churna was developed. Hence, the current technical work would be practically helpful for the future preparation and standardization study of Vyoshadi Churna

Effect of an Ayurvedic intervention (Ayush-64) in mild to moderate COVID-19: An exploratory prospective single arm clinical trial

Background: Ayush-64 is an Ayurvedic formulation, developed and patented by Central Council of Research in Ayurvedic Sciences (CCRAS). In the present study, we repurposed it for use in mild to moderate COVID-19 cases based on a pilot study against Influenza like illness (ILI) and molecular docking study which revealed that several compounds isolated from Ayush-64 demostrated antiviral activity. Purpose: To evaluate the role of Ayush-64 in clinical recovery of mild to moderate COVID-19 pattients Study Design: A single arm, pilot study in mild to moderate COVID-19 patients Methods: The study was conducted from 20th June, 2020 to 11th August, 2020 at Chaudhary Brahm Prakash Ayurved Charak Sansthan (CBPACS), New Delhi, India involving 37 confirmed COVID-19 participants. Ayush 64 tablets in the dose of two tablets (500 mg each) thrice daily was given to the participants for a duration of either 8 or 14 days. Number of participants showing ‘clinical recovery’ was set as primary outcome. Percentage of participants with negative SARS-CoV-2 on nasal or throat swab in a 2-day consecutive real time RT-PCR test was evaluated as secondary outcome. Result: In the study 86.1% participants have shown clinical recovery after 14 days intervention of Ayush-64, out of which 75% clinically recovered within 7 days. Finding of RT-PCR test has shown that 69.4% participants turned negative till 15th day, out of which 50% became negative on 8th day. No AE/ ADR was observed during the course of the study. Conclusion: Ayush-64 is a safe treatment option in mild to moderate COVID-19 cases and is likely to significantly facilitate clinical improvement in terms of duration for clinical recovery and attaining negative conversion, without any ADR/AE