Chemical Constituents Of Vitex Negundo Var. Cannabifolia Fruits And Pre-clinical Pharmacokinetics Of Casticin In Rat Plasma

Mu-Jing-Zi is the ripe fruits of Vitex negundo var. cannabifolia （Verbenaceae） and isused to dispel the moisture, relieve cough and asthma, regulate Qi and relieve pains inTraditional Chinese Medicine. Modern pharmacological studies show that Mu-Jing-Zi hasthe effects of analgesic, antiasthmatic, anti-inflammatory, antibacterial and antioxidant.Vitex negundo var. cannabifolia is a shrub growing in the most area of China. It is mainlydistributed in Hebei, Hubei, Hunan, Guangdong, Guangxi, Sichuan and Guizhou province.Recently, the research results show that the plant has been proved to be a rich sources oflignans, as well as flavonoids and diterpenoid, which exhibit a wide variety of biologicalactivities including antioxidant, anti-inflammatory, antisepticise and analgesic effects.Mu-Jing-Zi is commonly used in clinical practice for curing cough and anti-inflammatory infolk medicine. However, the insufficient research of Mu-Jing-Zi resulted in unknownactive chemical constituents. Therefore, figuring out the constituents of Mu-Jing-Zi isbeneficial to its deeper development. The main contents of our study are as following twoaspects:1Study on chemical components of Vitex negundo var. cannabifolia fruits.The air-dried and powdered seeds of Vitex negundo var. cannabifolia were extracted with80%EtOH （×3）, each extraction period lasting2h. After removal of the solvent underreduced pressure, the residue was suspended in H2O and partitioned sequentially withpetroleum ether, CH₂Cl₂, EtOAc and n-butanol, respectively. The CH2Cl2-soluble part wassubjected to column chromatography, TLC, and semi-preparative high performance liquid.A total of30with known compounds were isolated from dichlormethane fraction and23structures were elucidated by detailed spectroscopic analyses on the basis of NMR, IR, andMS data. The substance are vitexilactone, previtexilactone, betulinic acid, casticin,3’,4’,5-hydroxy-7-methoxy–flavanone, p-hydroxy benzaldehyde, β-sitosterol, vanillin,vitedoin A,6-hydroxy-4β-（4-hydroxy-3-methoxyphenyl）-3α-hydroxymethyl-7-methoxy-3,4-dihydro-2-naphthaldehyde, paulownin, vitedoin B, isolariciresinol, sesamin,cis-p-coumaric acid, ursolic acid, acacetin,2,5-Dihydroxy-l-methoxyxanthone,15-Hydroxydehydroabietic acid,5-hydroxy-6,7,3’,4’-tetramethoxyflavone,8-hydroxy-5,7,3’,4’-tetramethoxyflavone, p-hydoxybenzoic acid and 4-Methoxybenzaldehyde respectively. By comparing with the chemical compositions ofVitex negundo L., Vitex negundo var. cannabifolia, Vitex trifolia L., Vitex trifolia L. var.simplicifolia Cham. and Vitex agnus-castus L., we found that casticin is the characteristiccomponents of the Vitex species. In the process of earlier study, casticin is considered asthe major active chemical constituent for the treatment of premenstrual syndrome （PMS） inthe medicinal plants of Vitex species. However, due to its pharmacokinetic properties is notclear, casticin is hard to further study. So, to clarify its pharmacokinetic characteristics isparticularly important to develop casticin into new drug.2Study on pharmacokinetics of casticin in Rat Plasma.Pharmacokinetics is the study of the time course of a drug within the body andincorporates the processes of absorption, distribution, metabolism and excretion. To betterinterpret the effect and toxicity of the drug, an analytical method should be established todetermine the drug and its metabolites quantitatively, which help us to know about theirmechanisms of action and their pharmacokinetic properties during the initial state of drugdevelopment and in clinical therapy. With the improvement of combinatorial chemistry andseparation and purification of natural products, a number of candidate compoundsappeared and required pharmacokinetic analysis. Investigators have shown increasinginterest in developing and optimizing high-throughput analytical methods with moresensitivity，accuracy and lower consumption for quantitating small molecule drugs,metabolites, and other xenobiotic biomolecules in biological matrices. Casticin（3’,5-dihydroxy-3,4’,6,7-tetramethoxyflavone）, also named Vitexicarpin, as a major flavonecomponent has been isolated from a number of medicinal plants of Vitex species, such asVitex trifolia L., Vitex agnus-castus L. and Vitex rotundifolia L. It has been reported thatcasticin has antitumor, anti-inflammation, anti-hyperprolactinemia, antioxidant, andanti-nociceptive properties. In recent years, this compound is considered as the majoractive chemical constituent for the treatment of premenstrual syndrome （PMS） in themedicinal plants of Vitex species. In comparison with conventional drugs such asantiinflammatory agents （ibuprofen）, psychotropic drugs （selective serotonin reuptakeinhibitors）, estrogen and progesterone, naturally occurring products containing casticinhave been extensively used to treat PMS in European for little adverse effects. Consideringthe anti-inflammation, anti-nociceptive, anti-hyperprolactinemia and estrogenic activities of casticin, it has a good potential to be developed as a new therapeutic reagent for PMS.For the development of casticin as a new drug, it is necessary to establish a selective andsensitive analytical method for the quantitative estimation of casticin in preclinicalbiopharmaceutical and clinical pharmacology studies. Several methods for the separationand quantification of casticin have been described using high performance liquidchromatography （HPLC） with ultra-violet detection. However, these methods mainlyfocused on the quantification of casticin in raw materials or pharmaceutical preparations.Few studies can be found in biological specimen. The purpose of this study was to developa simple, rapid and reliable LC-MS assay for the determination of casticin in rat plasmaand applied the assay to the pharmacokinetic study in rats after oral and intravenousadministrations of casticin.