Analysis and generalization of key extraction methods, isolation and characteristics of biologically active substances obtained from plant extracts

Introduction. Development of authentic methods for analysis complex herbal medicines, especially poly-herbal formulations, to help reliably formulate phytochemical contents, including quantitative analysis of marker biologically active substances and other essential components, is a serious and pressing issue for many scientists. Materials and methods. The following methods of analysis of compounds obtained from plant extracts were used for this study: spectrophotometry, thin-layer and high-performance liquid chromatography, enzyme-linked immunosorbent assay based on monoclonal antibodies, phytochemical screening analysis, Fourier transform infrared spectroscopy. Spectrophotometric method of analysis and mathematical modeling using Excel and Mathcad software packages were used. Literary sources of domestic and foreign authors who studied the methods of extraction, isolation and characteristics of biologically active substances obtained from plant extracts were also analyzed. Results and discussion. Polar solvents such as methanol, ethanol or ethyl acetate are used to extract the hydrophilic compounds. To remove lipophilic compounds use dichloromethane or a mixture of dichloromethane / methanol in a ratio of 1:1. In some cases, hexane extraction is used to remove chlorophyll.  For Soxhlet extraction the usual solvents are used – methanol, ethanol or a mixture of alcohol and water, room temperature, pressure is not used, the procedure time is 3-4 days, the required amount of solvent depends on the sample size. For the extraction of Soxhlet in the case of sonication also use common solvents such as methanol, ethanol or a mixture of alcohol and water, the temperature depends on the variant of the solvent, the pressure is not applied, the procedure time – 3-18 h. For the maceration procedure during Soxhlet extraction, methanol, ethanol or a mixture of alcohol and water is used as a solvent, this mixture can be heated, the pressure is not applied, the procedure time is 1 h, the required solvent volume is 50-100 ml. Other modern extraction methods include solid-phase microextraction, supercritical liquid extraction, pressurized liquid extraction, microwave extraction, solid-phase extraction, and surface active agents mediated techniques. The following separation methods are also used: thin layer chromatography, column chromatography, flash chromatography, Sephadex column chromatography and high performance liquid chromatography, enzyme-linked immunosorbent assay, phytochemical screening analysis, infrared spectroscopy and facilitating the identification of biologically active compounds. High performance liquid chromatography is a universal, reliable and widely used technique for isolating biologically active substances. This technique is gaining popularity among various analytical techniques as the main choice for plant quality control. Natural products are often isolated after evaluation of the relatively crude extract in biological analysis in order to fully characterize the active substance. The biologically active substance is often present only as a secondary component in the extract, and high performance liquid chromatography resolution is ideal for rapid processing of such multicomponent samples on both analytical and preparative scales. Immunoassay, which uses monoclonal antibodies against drugs and low molecular weight natural bioactive compounds, is becoming an important tool in the analysis of bioactive compounds. This method demonstrates high specificity and sensitivity for receptor binding assays, enzymatic and qualitative assays, and quantitative assays. Phytochemical screening analysis is a simple, fast and inexpensive procedure that gives the researcher a quick answer about different types of phytochemicals in the mixture and is an important tool for the analysis of bioactive compounds. After obtaining the crude extract or active fraction of plant material, phytochemical screening can be performed using appropriate tests to get an idea of the type of phytochemicals contained in the mixture or fraction of the extract. Conclusion. Because the bioactive compounds contained in plant material consist of multicomponent mixtures, their separation and determination still have many open questions. In practice, most of them must be purified by a combination of several chromatographic methods and other purification methods to isolate biologically active compounds.