| Artemisia argyi is a famous species with a long history in traditional Chinese medicine,which is the dry leaf of Artemisia argyi Lévl.et Vant.It was first recorded in Tao hongjing's " Mingyi Bielu".Traditional Chinese medicine believes that Artemisia argyi tastes bitter and spicy,warm in nature,distributes to liver,spleen and kidney channels.The major material basis of Artemisia argyi is artemisia argyi essential oil(AAEO).One of the important basic substances for the mechanism of TCM is Chinese medicine volatile oil.It has been reported that AAEO has special effect in the treatment of pain.Thus,this study was based on the compatibility theory of traditional Chinese medicine prescriptions and prepared a kind of high safety,drug loadings appropriate analgesic volatile oil compound microemulsion gel.The study carryed out a systematic study from the optimization of AAEO extraction process,component analysis,pharmacological efficacy,preparation and evaluation of analgesic microemulsion gel preparation,etc.The main research contents and results are as follows:1.Extraction process optimization and component analysis of AAEO.Volatile oil was extracted by steam distillation,and the extraction rates of AAEO in different producing area were used as the evaluation index.The influences of the immersion time,the extraction time and the ratio of material to liquid on the extraction rate of AAEO in different producing area were studied.Firstly,the extraction process parameters were obtained preliminarily by the single factor investigation.Then the optimum extraction parameters were obtained through Box-Behnken response surface method.The best immersion time,extraction time,ratio of material to liquid for AAEO in Qichun were 0.8h,3.29 h,1:9.12(g/mL)respectively.The best predicted oil yield was 1.023%.The best immersion time,extraction time,ratio of material to liquid for AAEO in Tangyin were 1.21 h,3.86 h,1:11.28(g/mL)respectively.The best predicted oil yield was 0.917%.The best predicted oil yield was 1.023%.The best immersion time,extraction time,ratio of material to liquid for AAEO in Sichun were 0.56 h,3.19 h,1:10.60(g/mL)respectively.The best predicted oil yield was 0.961%.The best predicted oil yield was 1.023%.The best immersion time,extraction time,ratio of material to liquid for AAEO in Jiangxi were 1.15 h,2.99 h,1:9.45(g/mL)respectively.The best predicted oil yield was 0.963%.The optimal process parameters were verified by three batches of experiments,and the results showed that the extraction rate of AAEO in different producing area were basically consistent with the predicted values.Among the four producing areas examined,Qichun AAEO yield(1.023%)was the highest,followed by Jiangxi AAEO(0.963%),Sichuan AAEO(0.961%)and Tangyin AAEO(0.917%).The components of AAEO in different producing areas were analyzed by gas chromatography and mass spectrometry(GC-MS)and the results showed that the AAEO components were abundant in four producing areas.The common ingredients were eucalyptol,camphor and caryophyllene oxide.And the highest percentage of eucalyptus oil was Qichun AAEO,about 41.36%.Next was Tangyin AAEO(35.95%),Jiangxi AAEO(29.04%)and Sichuan AAEO(26.88%).The percentage of camphor is the most in Jiangxi AAEO(12.84%),followed by Qichun AAEO(7.52%),Tangyin AAEO(5.28%)and Sichuan AAEO(4.68%)respectively.The highest percentage of Caryophyllene oxide was Jiangxi AAEO(21.19%),followed by Qichun AAEO(3.76%),Sichuan AAEO(2.78%),and Tangyin AAEO(1.48%).2.Preparation and evaluation of analgesic microemulsion gel.Based on the previous studies of our research group,the compound analgesic microemulsion gel was obtained.The physicochemical parameters of analgesic microemulsion gel were measured.The results showed that the particle size of the microemulsion met the requirements,and the physicochemical parameters such as pH values,refractive index and conductivity were relatively stable.At the same time,the determination methods of contents of volatile oil in analgesic microemulsion gel were investigated including specificity,linear relationship,precision,stability,repeatability and sample recovery.The results were as following: the specificity results showed that the blank gel had no effect on the determination of eucalyptol and menthone in analgesic microemulsion gel;eucalyptol and menthone showed good linearity in the concentration range of 50.18~802.88?g/mL and 50.23~251.15?g/mL,respectively;the precision results showed that the RSD(n=6)values of eucalyptol and menthone were 1.192% and 2.04% respectively,both less than 3%,which suggested that the precision of this method was good;the RSD(n=6)values of the stability of eucalyptol and menthone were 0.652% and 1.82% respectively,both less than 3%,indicating that the method was stable;Repetitive experiment showed that the RSD(n=6)values of eucalyptol and menthone were 0.361% and 1.66% respectively,both less than 3%,indicating that the method had good repeatability;the RSD(n=6)values of the average recovery of eucalyptol and menthone were 1.99% and 1.64%,respectively,which met the requirements.According to the linear,we got eucalyptol and menthone contents in the gel and the contents of eucalyptol and menthone were 187.13 ?g/mL and 175.88 ?g/mL,repectively.3.Study on in vitro transdermal permeability and skin irritation of compound analgesic microemulsion gel.The percutaneous permeability of analgesic microemulsion was measured,and the standard curves of eucalyptol and menthone in the receiving solution of analgesic microemulsion gel were established,which showed good linearity in the concentration range of 1.21~300.05 ?g/mL and 1.07~374.91?g/mL,respectively.At the same time,the results of in vitro transdermal permeability study showed that the release of eucalyptol and menthone within 12 hour was close to the Higuchi equation,indicating that the release of analgesic microemulsion gel was sustained.According to skin irritation test,rabbits were used as experimental subjects and compound analgesic microemulsion gel was explored by self control method.The results showed that single or multiple administration of the gel had no skin irritation,such as erythema and edema.4.Study on analgesic pharmacodynamics of analgesic microemulsion gel.The anti-inflammatory and analgesic effects of analgesic microemulsion gel were investigated by acetic acid writhing test and xylene ear swelling test.Acetic acid writhing experiment showed that high,medium and low concentrations of analgesic microemulsion gel could inhibit the writhing times of mice to a certain extent.They could increase the pain threshold of mice,extend the incubation time of mice in response to pain stimuli,and reduce the times of writhing induced by pain stimulus.Among them,the analgesic microemulsion gel in the medium dose group had the most inhibitory effect on the torsion of mice,with a pain inhibitory rate of 40.28%.The experimental results of ear swelling induced by xylene showed that high,medium and low concentrations of analgesic microemulsion gel could inhibit the ear swelling induced by xylene in mice to a certain extent.The inhibition rates of ear swelling induced by high,medium and low concentration of analgesic microemulsion gel were 32.63%,57.89% and 15.79%,respectively.The inhibition rate of mice in the medium dose group was the most obvious in the three concentration groups,which could significantly reduce the degree of swelling in the right ear of mice in the experimental group. |
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