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Shoots of Ajuga turkestanica (Rgl.) Brig. (Labiatae, Mint) are a source of highly effective biologically active compounds such as phytoecdysteroids and natural glycosides [1,2].Ecdysterone, cyasterone, ajugalactone, ajugasterone, α-ecdysone, and turkesterone have been isolated previously from A. turkestanica. Of these, the main components are ecdysterone and turkesterone, which have very high anabolic activities [1].The ecdysteroid content, in particular, that of turkesterone, in the raw material was determined by gravimetry [3][4][5] or UV spectrophotometry [6]. The turkesterone content varied from 0.025 to 0.17, which means that A. turkestanica is not an industrial raw material.HPLC is known to determine with high accuracy the content of ecdysteroids in certain plant sources [7-9]. We investigated the contents of turkesterone and ecdysterone in the aerial part of A. turkestanica by HPLC on a Zorbax Eclipse XDB-C18 column, 3×150 mm, 3.5 µm (Agilent Technologies) using an Agilent LC 1100 chromatograph with a four-eluent pump, deaerator, and variable wavelength UV detector. The chromatograph was controlled and the results were processed using Agilent ChemStation software for the liquid chromatograph. UV detection of separate ecdysteroids was carried out at working wavelength 247 nm. The elution gradient is described below: Time, min 0.00 15.00 30.00 35.00 40.00 45.00 Water Acetonitrile 100 0 95 5 15 85 0 100 0 100 100 0The flow rate was 0.5 mL/min. Ecdysterone and turkesterone were determined quantitatively using an external standard and a two-point calibration curve of the form Y = mx + b, where Y is the peak area and x is the amount of compound in %. Constants m and b were determined for turkesterone (m = 229310.00, b = 2.84×10 -12 ) and ecdysterone (m = 275180.00, b = 5.68×10 -12 ).Standard solutions in ethanol (50%) at concentrations 0.1 and 0.01% were prepared for quantitative determination of ecdysteroids. Samples were introduced through a 20-µL loop on an Agilent manual injector.The amount of dry substances in the extract was determined by refractometry. This enabled sample solutions to be prepared in the concentration range required for HPLC analysis. The analyzed sample was prepared at a concentration of 0.2% in ethanol (50%). The sample volume was 20 µL, as mentioned above. The same solvent gradient was used for elution. Ecdysteroids in the extract from A. turkestanica were identified and analyzed quantitatively using the Agilent ChemStation and the calibration table and curve (Fig. 1).