Nepeta Volatile Oil In Vitro Intervention Experimental Study Of The Activity Of 5-l0, And The Method Of Evaluation

Herba schizonepetae Briq, the aerial ground part of schizonepeta tenui-folia Briq(family Labiatae), is one of the commonly used Chinese herb that belongs to the category of relieving Exterior syndrome, which has the effect of expelling wind and treart miseal. The volatile oil is one of the major effective components of Herba schizonepetae Briq., and its contents is high.. The results of previous research showed, Schizonepetae Volatile oil (STO) has obvious anti-inflammation effect, which the anti-inflammatory mechanism be related with the pathway of arachidonic acid (AA). Based on the previous studies, in this study we concentrated on the 5-lipoxygenase (5-LO), that is an important enzyme of AA metabolism, to study the effect of STO on the activity of 5-LO in vitro so as to elucidate its mechanisms of anti-inflammatory action.Objective: To investigate the effect of STO on the activity of 5-LO in vitro so as to elucidate its mechanisms of anti-inflammatory action more. Meanwhile, to establish a screening platform for the effective parts of Traditional Chinese Medicine (TCM) that maybe affect the activity of 5-LO in vitro.Methods: First, the ear-swelling induced by xylene were adopted to validate the effect of anti-inflammation of STO in mice. Second, the method of RP-HPLC to extract, separate and detect leukotriene B₄(LTB₄) and 5-hydroxy-6, 8, 11,14-eicosatetraenoic acid (5-HETE) were established and evaluated. At last, the effect of STO and other volatile oil which has anti-inflammatory action on the activity of 5-LO were observed by the established RP-HPLC method in vitro.Results:1. STO exhibited definite anti-inflammation action. STO inhibited the ear-swelling induced by xylene in mice by ig 3 days or ig twice in a day (P< 0.05 or < 0.01) .2. The methods of RP-HPLC to extract, separate and detect LTB4 and 5-HETE were established. Thoracic cavity leukocytes, which from the pleurisy model rat induced by carrageenan, were used as the reaction system in vitro. Exogenous AA, STO in different concentration and calcium ionophore A₂₃₁₈₇ were added into this system to be incubated. The metabolites of 5-LO, LTB4 and 5-HETE, were extracted, separated and detected by means of RP-HPLC.3. Anti-inflammatory mechanism of STO was related to suppress the activity of 5-LO. STO both dissolved in dimethylsulfoxide and contained in serum (blood plasma), inhibited obviously the activity of 5-LO in rat leukocytes in dose dependent way in vitro, which decreased the production of LTB₄ and 5-HETE. Compared with STO, the synthesis of LTB₄ and 5-HETE were not significantly suppressed by main components of STO, menthone and pulegone used alone or used mixing (M+P). However, the suppressed effect of M+P on the activity of 5-LO was stronger than menthone and pulegone used alone. Compared with STO drug-serum or plasma from normal rats, the synthesis of LTB4 and 5-HETE were more significantly inhibited by STO drug-serum or plasma from the inflammatory model rats induced by AA.4. The methods of RP-HPLC to extract, separate and detect LTB₄ and 5-HETE were applied to investigate the effect of six kinds volatile oil of drugs for relieving exterior syndrome on the activity of 5-LO in vitro. Compared with Bohe and Juhua in same dose, the synthesis of LTB₄ and 5-HETE were significantly inhibited by Guizhi, Xinyi, Xiangru and Qianghuo. It is suggested that the inhibition effect of volatile oil on the activity of 5-LO, drugs for relieving the exterior syndrome with pungent in flavor and warm in nature were stronger than the drugs of pungent in flavor and cool in nature.Conclusion: The results showed that STO had the significant anti-inflammation action, which the mechanism were related to the inhibition effect on the activity of 5-LO in dose dependence, which be beneficial to guide clinical treatment. Meanwhile, the established research method was sensitive to reflect the action of drug affecting the activity of 5-LO, which could be a screening platform for the anti-inflammatory and anti-allergic drugs in vitro.