Comparative phytochemical Screening and Biological activities of in vitro and in vivo Vanda tessellata (Roxb.) Lodd. ex G. Don and V. testacea Rchb. f.

Abstract

V. tessellata and V. testacea are most commonly used medicinal orchid. This study aims to develop a sustainable alternative through plant tissue culture technique, by comparing the phytochemicals and biological activities of in vitro raised samples with their wild counterparts. Tissue culture of V. tessellata and V. testacea was carried out through asymbiotic seed germination on Murashige and Skoog (MS) medium. The methanolic extracts of wild (root and leaf) and in vitro (protocorm and plantlets) samples were qualitatively screened for alkaloids, phenols, flavonoids, saponins, tannins, phliobatannins, terpenoids, steroids and anthocyanins, and quantitatively screened for Total Phenolic Content (TPC) and Total Flavonoid Content (TFC) using FC-reagent and AlCl3 colorimetric test. Also, the extracts were examined for antioxidant, antibacterial and cytotoxic activity by using DPPH, agar well diffusion and MTT assay respectively. The best results for seed germination and protocorm formation of V. tessellata and V. testacea were observed in full strength MS medium with 10% coconut water (FMSCW) and quarter strength MS medium with 10% coconut water (QMSCW) respectively. The highest number of shoots (10.5±0.65) of V. tessellata was observed in FMSCW fortified with 1.5 mg/L BAP and 0.5 mg/L NAA and longest shoot length (2.98±0.18cm) was recorded in FMSCW fortified with 1 mg/L BAP and 0.5 mg/L NAA. In V. testacea, highest number of shoots (10.75±0.48) was found in FMSCW and 0.5 mg/L BAP and the longest shoot length (3.1±0.24 cm) was obtained in FMSCW. Phytochemical screening of V. tessellata and V. testacea revealed the presence of alkaloids, phenols, flavonoids, saponins, tannins, phliobatannins, terpenoids, and steroids in both wild and in vitro raised samples. The highest amount of TPC (223.69±0.33 mg GAE/gm) and TFC (87.93±0.43 mg QE/gm), lowest IC50

value (109.79±0.57 µg/mL) for antioxidant and the highest zone of inhibition was obtained in wild leaf extract of V. testacea followed by V. tessellata leaf extract. All samples showed very high IC50 values against HeLa and MCF7 cell lines indicating inactive or very weak cytotoxic activity. In case of normal cell, instead of inhibition cell were proliferated. The wild leaf extract of V. testacea showed lower AC50 value (484.94±5.31 µg/mL) for normal cell line than in vitro raised samples. In conclusion, an efficient protocol for in vitro propagation for phytochemical extraction of V. tessellata and V. testacea was established. The V. testacea sample showed better phytochemicals and bioactivity than the V. tessellata sample. The wild V. tessellata and vi

V. testacea plant extracts had higher medicinal potential due to higher TPC, TFC, antioxidant, and antibacterial activity, while in vitro raised plantlets can still be used as an alternative source for bioactive compounds for medicinal and other applications as well as for conservation and sustainable utilization of the species.