Development and Validation of High-Performance Liquid Chromatography for Identification and Quantification of Phytoecdysteroids Ecdysterone and Turkesterone in Dietary Supplements

Abstract

Phytoecdysteroids are a group of naturally occurring ecdysteroid hormones found in certain plant families and used for centuries for their adaptogenic, tonifying, and antioxidant properties. Ecdysterone and turkesterone are the dominant phytoecdysteroids found in the plant species Rhaponticum carthamoides Willd., Cyanotis arachnoidea C.B.Clarke and Ajuga turkestanica (Regel) Briq., and have been recognized for their adaptogenic potential and ability to enhance physical performance. However, over the past few years, supplementation with ecdysterone by professional athletes has raised some concerns about its safety and quality, leading to its inclusion in the World Anti-Doping Agency monitoring program. This study proposes a simple and reliable method for quality control of ecdysterone- and turkesterone-containing products using high-performance liquid chromatography (HPLC) analysis. The HPLC method is based on reversed-phase chromatography and gradient elution to achieve a superior separation of phytoecdysteroids. The method has been shown to qualify both phytoecdysteroids at low concentrations such as 10.98 μg/mL for ecdysterone and 11.43 μg/mL for turkesterone. This method could be successfully used in research programs and routine quality control of dietary supplements to ensure their safety in professional athletes’ nutrition.

Approaches, Strategies and Procedures for Identifying Anti-Inflammatory Drug Lead Molecules from Natural Products

“…HPLC is commonly used for the purification and isolation of SM compounds from NPs due to its high sensitivity and specificity [ 173 ]. Biologically active compounds are often found in NP extracts as a minor constituent.…”

Ecdysterone and Turkesterone—Compounds with Prominent Potential in Sport and Healthy Nutrition

“…Ecdysterone is a structurally characteristic ecdysteroid-2β,3β,14α,20β,22α,25β-Hexahydroxycholest-7-en-6-one (ecdysterone, β-ecdysone, 20-hydroxyecdysone) [5,41]. Important for the biological activity of ecdysteroids are the double bond at C-7, the keto group at C-6, and the hydroxyl groups at positions C-2, C-3, C-14, and C-22, and the hydroxyl group at C-20 correlates with anabolic activity [28].…”

“…In recent decades, significant research has been performed on the potential performanceenhancing benefits and therapeutic applications of ecdysterone [15,23,28]. The World Anti-Doping Agency (WADA) [33] agreed to add ecdysterone to their monitoring program in 2020, and the inquiry has continued ever since.…”

“…Turkesterone is a PE, with 27 carbon atoms and seven OH-groups; it is considered that the OH groups at C-20 and C-11 are responsible for the anabolic effects [28,215]. Turkesterone is considered to possess an anabolic effect, and it is used as DS by athletes, instead of anabolic steroids [28,30]. It is found in some endemic plants, such as A. turkestanica, R. carthamoides, Triticum aestivum L., Vitex scabra, etc.…”

“…The highest content of turkesterone is considered to be in Ajuga turkestanica [19]. Moreover, in DSs, turkesterone is derived from A. turkestanica [28].…”