Фітохімічне вивчення сировини та екстрактів на основі видів роду Підмаренник

Abstract

Дисертацію присвячено комплексному фітохімічному дослідженню трави та екстрактів підмаренника справжнього (Galium verum) та трави та екстрактів підмаренника чіпкого (Galium aparine), розробки параметрів їх стандартизації, встановлення груп БАР, які зумовлюють їх імуномодулювальну дію, визначення доцільності комплексної переробки сировини. Встановлено технологічні параметри сировини, динаміку екстрагування БАР з неї, розроблено технологію одержання екстрактів та субстанцій, досліджено їх хімічний склад та виявлено імуномодулювальну активність, встановлено параметри стандартизації. Розроблено проєкти МКЯ «Трава підмаренника чіпкого», «Підмаренника чіпкого екстракт рідкий» та «Підмаренника справжнього екстракт рідкий».

Диссертационная работа посвящена комплексному фитохимическому изучению травы и экстрактов подмаренника настоящего (Galium verum) и травы и экстрактов подмаренника цепкого (Galium aparine), разработке параметров их стандартизации, выявлению групп БАВ, обусловливающих их иммуномодулирующее действие, установлению целесообразности комплексной переработки сырья. Определены технологические параметры травы G. verum и травы G. aparine, динамика экстрагирования БАВ, разработана технология получения экстрактов и субстанций из сырья, исследован их химический состав, и установлена иммуномодулирующая активность, определены параметры стандартизации. Разработаны проэкты МКК «Трава подмаренника цепкого», «Подмаренника цепкого экстракт жидкий» и «Подмаренника настоящего экстракт жидкий».

The present dissertation is aimed at the phytochemical study of the raw materials and extracts from the herb of Galium verum (Lady’s Bedstraw, yellow Bedstraw) and Galium aparine (Cleavers, Goosegrass, Catchweed) of Rubiaceae Juss. family, as well as the development of the parameters of their standardization, research in the the dependence of their immunomodulatory activity on the chemical composition, and the feasibility of complex processing of the raw material. The study in the relationship between the morphological characteristics of the species Galium genus and the presence of iridoids, hydroxycinnamic acids, and individual flavonoids, such as isoroifoline, luteoline-7-О arabinosylglucoside, diosmetin-7-O-glycopyranoside, palustroside, diosmetin-7-O xyloglucoside has been performed. The technological parameters of the herb G. verum and G. aparine, namely a bulk volume, bulk density and tapped density of the medicinal plant raw material (MPRM), a pore volume, porosity, free volume of the layer, absorption coefficients of the solvent, have been determined. The regularities of extracting the main groups of BAS from the herb of G. verum and G. aparine have been revealed have first been established. It has been found that the optimal conditions for BAS extraction are as follows: the MPRM particle size of 3-4 mm, the raw material extractant ratio of 1 : 10; the extraction ratio of 3, with the duration of each extraction of 30 min. Based on the technological parameters of the MPRM determined and the regularities of BAS extraction, the flowchart for obtaining 8 liquid extracts from the herb of G. verum and G. aparine has been developed. The content of the main BAS groups. i.e. extractives, soluble polysaccharides (WSPS), hydroxycinnamic acids, flavonoids and polyphenolic compounds, has been quantified in the extracts obtained in compliance with the flowchart specifications. The study in the WSPS from the herb G. verum and G. aparine has established that their monomeric composition is mainly presented by D-galactose, L-arabinose, D-glucose, D-mannose and L-rhamnose. The content of renewable sugars in WSPS from extract GV-V has been quantified. The amino acid and elemental composition of WSPS in extracts GV-V and GA-V has been studied and the total ash in WSPS has been quantified. By means of the HPLC method, 11 compounds of phenolic nature, including 5 hydroxycinnamic acids (caffeic, сhlorogenic, neoсhlorogenic, 3,5-dicaffeoylquinic and 4,5-dicaffeoylquinic acids) and 6 flavonoids (rutin, hyperoside, quercetin, quercitrin, isoquercitrin and catechin) have been identified in extracts GV-60 and GA-60. It has been determined that the prevaling compounds in GV-60 and GA-60 are rutin and chlorogenic acid. Compounds in the extracts from the herb G. aparine have been studied by the HPLC-MS method with 14 compounds found, among them 13 identified as 3 iridoids, 3 flavonoids, 1 phenolcarboxylic acid and 6 hydroxycinnamic acids. In G. aparine herb, the presence of p-hydroxybenzoic, 3-O and 4-O-caffeylquinic acids, 3,4-, 3,5- and 4,5-O-dicaffeylquinic acids, isorhamnetin-3-O-glucorhamnoside and quercetin-3-O rhamnoglucoside-7-O-glucoside has been first determined. With the use of HPLC method the content of saponins i. e., ursolic, oleanolic, euscaphic, tormentic acids, uvaol, betulin and lupeol has been identified and quantified in extracts GV-96 and GA-96. Prevaling in extract GA-96 is euscaphic acid, whereas in extract GV-96 lupeol. In extract GV-96, prevailing are lupane derivatives, in extract GA-96 prevailing are ursane derivatives. With the view to a coplex processing of raw material from the meal remaining after the production of water extracts from MPRM, pectin substances were obtained. The composition of functional groups in the pectin substances obtained (free carboxylic, methoxylated carboxylic), the total amount of carboxylic, methoxylic substances, as well as the esterification degree were determined. The elemental composition of the pectin substances indicated the prevailing presence of K, Na, Ca and Mg. The total ash in pectin substances has been quantified. With the view to a possible variation on the processing of the raw material, the WSPS-with-pectin complexes were obtained. It was found that for G. verum it contained 46.9 % of WSPS and 53.1 % of pectin substances; and for G. aparine the content of WSPS in the complex was measured at 40.67 % of WSPS; and the content of pectin substances came to 59.33 %. The project of the Drug Quality Control Methods (DQCM) for the herb of G. aparine was developed, and 5 batches of the herb were tested for compliance with the standardization parameters. The standardization parameters for extract GA-V were developed, with the 5 batches of the extract GA-V were studied for compliance with the specifications developed, and the project of DQCM “Galium aparine liquid extract” has been developed. The parameters of standardization of extract GV-96 have been developed, with the 5 batches studied for the compliance with the specifications, and the project of DQCM “Galium verum liquid extract” developed. The antibacterial and antifungal effect of liquid extracts from the herb G. verum and G. aparine has been studied. It has been established that all liquid extracts under study exhibit an antimicrobial effect, with the most active being the one obtained with the use of 96 % ethanol and Bacillus subtilis being the most sensitive to all extracts in research. It has been found that the all extracts, WSPS, pectin substances and the WSPS-with-pectin complexes from the herb of G. verum and G. aparine exhibit an immunomodulatory effect. The synergistic effect of the WSPS and phenolic compounds in the composition of water extracts from the herb G. verum and G. aparine has been studied, leading to the conclusion that the removal WSPS from water extracts when obtaining substances with an immunomodulatory effect is impractical. It has been found that the WSPS-with-pectin complexes from the herb G. verum and G. aparine exhibit a higher level of the immunomodulatory effect than separate WSPS and pectins. The data obtained indicate the feasibility of complex processing of MPRM with the view to producing immunomodulatory agents.