Standardization of Euphorbia tithymaloides (L.) Poit. (Root) by Conventional and DNA Barcoding Methods

Adulteration and substitution of medicinal plants havebecome amatter of great concern in recent years. Euphorbiatithymaloides is one such medicinal plant that hasgained importance but is often confused with other plants of the samespecies. In order to address this issue, this study aimed to conducta conventional and molecular pharmacognostic study for the identificationof the root of E. tithymaloides. Theroot of the plant was studied for the macroscopic observations, andthen, the root was ground into coarse powder for microscopic studiesand to determine the physiochemical properties. The powder was subjectedto extraction with solvents such as ethanol, ethanol/water (1:1),hexane, and ethyl acetate. The extracts were then used for qualitativeand quantitative (phenol, alkaloids, and flavonoids) phytochemicalanalysis. The molecular study was performed with the DNA barcodingtechnique. The DNA was extracted from the root of the plant, and itspurity was examined by gel electrophoresis (1% w/v). The DNA was thenamplified using an Applied Biosystems 2720 thermal cycler for therbcL, matK, and ITS primers. The amplified primers were sequencedwith a 3130 Genetic Analyzer, and the generated sequences were searchedfor similarity in the GenBank Database using the nucleotide BLASTanalysis. The micro- and macroscopic studies revealed the morphologicaland organoleptic characters as well as the presence of medullary rays,fiber, cork, sclereids, parenchymal cells, and scalariform vessels.The physiochemical properties were found within the limit. The phytochemicalanalysis revealed the presence of terpenoids, flavonoids, saponins,and alkaloids. In addition, the alkaloidal content was high in theethanol extract (63.04 & PLUSMN; 3.08 mg At E/g), while the phenol contentwas high in the hexane extract (10.26667 & PLUSMN; 1.77 mg At E/g), andthe flavonoid content was high in the ethyl acetate extract (41.458 & PLUSMN; 1.33 mg At E/g). After the BLAST analysis from the GenBankdatabase, the rbcL, ITS, and matK primers showed a similarity percentageof 99.83, 99.84, and 100. The phylogenetic tree for the species closestto each primer was generated using the MEGA 6 software. The matK locihad the highest percentage similar to the rbcL and ITS loci, indicatingthat the matK loci can be used to identify the root of E. tithymaloides as a standalone. The results fromthis study can be used to establish a quality standard for E. tithymaloides that will ensure its quality andpurity.