DEVELOPMENT OF FLAVONOIDS QUANTITATIVE DETERMINATION METHOD IN GLYCYRRHIZA PALLIDIFLORA HERB (GLYCYRRHIZA PALLIDIFLORA MAXIM.)

The licorice genus (Glycyrrhiza L.) plants are widely used in conventional and traditional medicine in different countries around the world. The genus includes about 15 species, but only three of them - G. glabra, G. uralensis and G. inflata are well studied and included in the Europe and Asia pharmacopoeias. Pale-flowered licorice (Glycyrrhiza pallidiflora Maxim.) is a perennial herbaceous plant from the false (not sweet) licorice section (Pseudoglycyrrhiza Krug.), which do not accumulate glycyrrhizin derivatives. This East Asian species grows in the Russian Far East. The distribution area of G. pallidiflora is limited. Only a few of its geographically isolated habitats are known: within the Khabarovsk city, near the Sarapulskoe village (Lower Amur) and in the Primorsky Kray on the Ryabokon Peninsula on the Lake Hanka southeastern coast. The G. pallidiflora successful introduction on the Kemerovo region-Kuzbass territory has been carried out over the past 5 years in the “Apothecary garden” territory of the Kuzbass Botanical Garden of the Institute of Human Ecology, FRC UUH SB RAS, Kemerovo. Currently, the scientific literature contains limited information on the composition and quantitative content of biologically active compounds (BAC) in the aerial part of G. pallidiflora. Therefore, the main task is to conduct phytochemical studies of the main classes of BAC, which have a pronounced pharmacological activity. We have proposed the amount of flavonoids quantitative determination method in the pale-flowered licorice herb, introduced in Kuzbass, by the differential spectrophotometry method, based on aluminum (III) chloride complexation reaction. The flavonoids extraction conditions from raw materials have been studied. They depend on the ethyl alcohol concentration, the plant raw materials grinding degree, the extraction time, and the raw materials and extractants ratio. The optimal extraction conditions were selected and a quantitative determination method was developed. On the base of it was established that the amount of flavonoids content in the herb of pale-flowered licorice (Glycyrrhiza pallidiflora Maxim.), in terms of rutin, is from 2.25 ± 0.38% to 2.44 ± 0, 04%. The relative error of the proposed method is ± 3.21%.

DEVELOPMENT AND VALIDATION OF QUANTITATIVE DETERMINATION METHOD OF DIHYDROSTREPTOMYCIN SULPHATE AND PROCAINE BENZYLPENICILLIN IN INJECTION SUSPENSIONS

The aim of the work was to develop and validate a method for the simultaneous identification and quantification of dihydrostreptomycin and benzylpenicillin in injectable suspensions. The method was validated by testing two preparations in the form of injectable suspensions containing benzylpenicillin 108–144 mg/ml and dihydrostreptomycin 180–220 mg/ml. Test samples were dissolved in purified water P, and standard samples: benzylpenicillin - in methanol (up to a concentration of 126 μg/ml), dihydrostreptomycin - in purified water P (up to a concentration of 200 μg/ml). The maximum allowable total uncertainty of the analysis was 1.64%, which is within the recommendations of SFU 2.0. The samples were separated on a Dionex Ultimate 3000 chromatograph equipped with a Luna C18 (2) 250 × 4.6 mm, 5 μm chromatographic column. The mobile phase was a mixture of acetonitrile and a solution of 0.01 M sodium heptanesulfonate with 0.05 M sodium phosphate trisubstituted, acidified with 0.1 M phosphoric acid to pH 6.0, in a volume ratio of 2: 8. Under mentioned conditions, dihydrostreptomycin and benzylpenicillin were completely separated. The established parameters of the chromatographic system did not exceed the limits specified in the FDA recommendations. The calibration curves were linear in the recommended SFU 2.0 range (80–120% of the nominal concentration of the corresponding active substance). The ratio of the amount of standard sample added to the test samples with its subsequent detection in the preparation was 99.35–100.79% for benzylpenicillin and 99.49–100.12% for dihydrostreptomycin, which does not exceed the limits recommended in SFU 2.0. The precision criterion was 0.07 for dihydrostreptomycin and -0.08 for benzylpenicillin, which is within the limits recommended in SFU 2.0. At the same time, the results of the study by different analysts at different times differed by 1.3% for dihydrostreptomycin and 0.98% for benzylpenicillin, which is well within the limits adopted in the recommendations of the FDA and SFU 2.0. Therefore, the method developed and validated by us for the simultaneous determination of dihydrostreptomycin sulfate and benzylpenicillin procaine in injectable suspensions can be considered suitable for routine analysis.

New Vortex-Synchronized Matrix Solid-Phase Dispersion Method for Simultaneous Determination of Four Anthraquinones in Cassiae Semen

In this study, a green ionic-liquid based vortex-synchronized matrix solid-phase dispersion (VS-MSPD) combined with high performance liquid chromatography (HPLC) method was developed as a quantitative determination method for four anthraquinones in Cassiae Semen. Two conventional adsorbents, C18 and silica gel were investigated. The strategy included two steps: Extraction and determination. Wasted crab shells were used as an alternative adsorbent and ionic liquid was used as an alternative solvent in the first step. Factors affecting extraction efficiency were optimized: A sample/adsorbent ratio of 2:1, a grinding time of 3 min, a vortex time of 3 min, and ionic liquid ([Domim]HSO4, 250 mM) was used as eluent in the VS-MSPD procedure. As a result, the established method provided satisfactory linearity (R > 0.999), good accuracy and high reproducibility (RSD < 4.60%), and it exhibited the advantages of smaller sample amounts, shorter extraction time, less volume of elution solvent, and was much more environmental-friendly when compared with other conventional methods.