Amit Kar1, Pulok K Mukherjee2, Sankarshan Saha3, Subhadip Banerjee4, Debayan Goswami5, Motlalepula G Matsabisa6, Rawiwan Charoensub7, Thidarat Duangyod8. 1. School of Natural Product Studies, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, 700 032, India. Electronic address: amit.kar2@gmail.com. 2. School of Natural Product Studies, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, 700 032, India; Institute of Bioresources and Sustainable Development, An Autonomous Institute Under Dept. of Biotechnology, Govt. of India, Imphal, 795001, India. Electronic address: naturalproductm@gmail.com. 3. School of Natural Product Studies, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, 700 032, India. Electronic address: snkrshan@gmail.com. 4. School of Natural Product Studies, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, 700 032, India. Electronic address: ami.subhadipbanerjee@gmail.com. 5. School of Natural Product Studies, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, 700 032, India. Electronic address: debayansmusic@gmail.com. 6. Department of Pharmacology, Faculty of Health Sciences, University of the Free State, PO Box 339, Bloemfontein, 9300, South Africa. Electronic address: MatsabisaMG@ufs.ac.za. 7. School of Health Science, Dept. of Applied Thai Traditional Medicine & Center of Excellence in Medicinal Plants and Thai Tradition Medicine, Mae Fah Luang University, Chiang Rai, 57100, Thailand. Electronic address: rawiwan.cha@mfu.ac.th. 8. School of Health Science, Dept. of Applied Thai Traditional Medicine & Center of Excellence in Medicinal Plants and Thai Tradition Medicine, Mae Fah Luang University, Chiang Rai, 57100, Thailand. Electronic address: thidarat.dua@mfu.ac.th.
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE: Trimada is well-known polyherbal Ayurvedic formulation used in Indian Traditional medicine since ancient times. It consisted of three inebriant herbs including "Chitraka" (Plumbago zeylanica Linn. Family- Plumabaginaceae), "Musta" (Cyperus rotundus Linn. Family- Cyperaceae) and Vidanga (Embelia ribes Burm. F. Family- Myrsinaceae) in equal ratios as mentioned in Ayurveda. Trimada is traditionally used to increase the functioning of the digestive system and metabolism. Along with these, it also assists in the reduction of cholesterol as well as reduces stomach aches and chest pain. AIM OF THE STUDY: This study is aimed to identify the metabolites present in this polyherbal formulation. Further, the cytotoxicity and interaction potential of the formulation and individual herbs with Cytochrome P450 isozymes (CYP3A4, 2D6, 2C9, 1A2) was evaluated by MTT assay and CYP450 enzyme inhibition. The concentration of heavy metals was also determined. MATERIAL AND METHODS: Ultra-performance liquid chromatography-quadrupole/time-of-flight mass spectrometry (UPLC-QTOF-MS) analysis was performed to detect and identify the phytoconstituents in the formulation. Cytotoxicity of the formulation was evaluated by MTT assay. CYP450 enzyme interaction potential of the individual herbs and the Trimada formulation was carried out through CYP-CO assay and fluorometric high throughput screening (HTS) assay for individual isozymes. The content of heavy metal in the formulation was quantified by Atomic Absorption Spectroscopy. RESULTS: Trimada formulation exhibited lower cytotoxicity to human liver carcinoma cell line (HepG2). CYP-CO assay revealed that the interaction potential of individual herbs and Trimada on the liver microsomes was found to be lesser than the standard inhibitor ketoconazole. Individual herbs and Trimada formulation displayed higher IC50 values than the respective standard inhibitors in the fluorimetric assay. UPLC-QTOF-MS analysis showed the presence of a number of active phytoconstituents including sesquiterpenes, phenolic acids, benzoquinones, triterpenes and flavonoids. The heavy metal concentration in the traditional medicinal herbal formulation was found within the approved limit. CONCLUSIONS: This study suggested that the individual herbs and Trimada formulation exhibited low cytotoxicity and contributes insignificant interaction with CYP450 isozymes. So, the formulation is considered to be safe for its therapeutic management without any potential drug interaction involving CYP 450 isozymes.
ETHNOPHARMACOLOGICAL RELEVANCE: Trimada is well-known polyherbal Ayurvedic formulation used in Indian Traditional medicine since ancient times. It consisted of three inebriant herbs including "Chitraka" (Plumbago zeylanica Linn. Family- Plumabaginaceae), "Musta" (Cyperus rotundus Linn. Family- Cyperaceae) and Vidanga (Embelia ribes Burm. F. Family- Myrsinaceae) in equal ratios as mentioned in Ayurveda. Trimada is traditionally used to increase the functioning of the digestive system and metabolism. Along with these, it also assists in the reduction of cholesterol as well as reduces stomach aches and chest pain. AIM OF THE STUDY: This study is aimed to identify the metabolites present in this polyherbal formulation. Further, the cytotoxicity and interaction potential of the formulation and individual herbs with Cytochrome P450 isozymes (CYP3A4, 2D6, 2C9, 1A2) was evaluated by MTT assay and CYP450 enzyme inhibition. The concentration of heavy metals was also determined. MATERIAL AND METHODS: Ultra-performance liquid chromatography-quadrupole/time-of-flight mass spectrometry (UPLC-QTOF-MS) analysis was performed to detect and identify the phytoconstituents in the formulation. Cytotoxicity of the formulation was evaluated by MTT assay. CYP450 enzyme interaction potential of the individual herbs and the Trimada formulation was carried out through CYP-CO assay and fluorometric high throughput screening (HTS) assay for individual isozymes. The content of heavy metal in the formulation was quantified by Atomic Absorption Spectroscopy. RESULTS:Trimada formulation exhibited lower cytotoxicity to humanliver carcinoma cell line (HepG2). CYP-CO assay revealed that the interaction potential of individual herbs and Trimada on the liver microsomes was found to be lesser than the standard inhibitor ketoconazole. Individual herbs and Trimada formulation displayed higher IC50 values than the respective standard inhibitors in the fluorimetric assay. UPLC-QTOF-MS analysis showed the presence of a number of active phytoconstituents including sesquiterpenes, phenolic acids, benzoquinones, triterpenes and flavonoids. The heavy metal concentration in the traditional medicinal herbal formulation was found within the approved limit. CONCLUSIONS: This study suggested that the individual herbs and Trimada formulation exhibited low cytotoxicity and contributes insignificant interaction with CYP450 isozymes. So, the formulation is considered to be safe for its therapeutic management without any potential drug interaction involving CYP 450 isozymes.