DNA barcoding for authentication of Orthosiphon Aristatus herbal medicinal products

Abstract

Orthosiphon aristatus (Misai Kucing) was traditionally known for its anti-diuretic properties and currently being studied for modern medicine applications had became part of high-value herbs targeted for production of commercialised herbal medicinal products (HMPs) in Malaysia. However, issues of adulterated HMPs rose as an unintended consequence of the booming herbal industry and current conventional methods including morphological identification and chemical analysis were unable to identify the source of contaminated herbs within adulterated HMPs. In this study, DNA barcoding was applied for authenticating six Orthosiphon aristatus HMPs that consisted of four monoherbal tea products and two polyherbal capsule products using chloroplastic (rbcL) and nuclear (ITS2) barcodes. Genomic DNA (gDNA) was extracted from Orthosiphon aristatus plant (MKP) and five HMPs but only rbcL and ITS2 barcodes of MKP and two HMPs (MKT 1 and MKC 1) were able to be amplified, direct sequenced and verified. Verified rbcL and ITS2 barcodes of MKP were included as reference barcodes for HMPs authentication and only rbcL barcode of MKP was deposited in GenBank with accession ID (MW492548.1). Bioinformatic analysis of HMPs barcodes using BLAST and Neighbour-Joining (NJ) method revealed that the monoherbal MKT 1 was authentic with both rbcL and ITS2 barcodes being identified as Orthosiphon aristatus. As for the polyherbal MKC 1, the rbcL barcode was identified as Astragalus membranaceus whereas the ITS2 barcode was identified as Strobilanthes namkadingensis. MKC 1 was considered contaminated as Strobilanthes namkadingensis, a species belonging to Acanthaceae family was an unlisted active ingredient within the labelled polyherbal composition. Overall, the findings from this study can served as a preliminary guidance for future authentication using DNA barcoding and potentially beneficial towards official regulatory bodies in standardising the quality and safety of HMPs.