Comparative Study On The Active Components And In Vitro Thrombolytic Activities Of Four Medicinal Aloe

Aloe vera,as a traditional medicinal plant,has many pharmacological activities such as anti-inflammatory,anti-allergy,immunomodulation,antibacterial,anti-tumor,anti-virus and so on.The commonly used medicinal aloe in Asia include Aloe barbadensis Miller,A.vera L.var.chinensis(Haw.)Berger,A.ferox Miller and A.arborescens Miller.Anthraquinones and proteins are the two main active components of aloe,but there are no systematic comparative studies on these two components in four medicinal aloe.In this thesis,the anthraquinones in four medicinal aloe was analyzed by HPLC and LC-MS.Then the protein in aloe was analyzed by gel electrophoresis,and the differentially expressed proteins of the four medicinal aloe were identified by mass spectrometry and bioinformatics.Finally,the thrombolytic and fibrinolysis activity of the active extracts and monomers of aloe were measured in vitro.This thesis includes five chapters.In the first chapter,the four medicinal aloe were briefly introduced.Then,the main active ingredients and the pharmacological activities of aloe were summarized.The second chapter was focused on the comparative analysis of anthraquinones in four medicinal aloe.The total contents of anthraquinones in four medicinal aloe were determined and the results showed that the content of anthraquinones was the highest in A.barbadensis.Furthermore,HPLC analysis showed that there was a certain degree of difference in the compositions and contents of anthraquinones in the four medicinal aloe,which may be related to the species and the growth environment of aloe.Meanwhile,the developed HPLC method had good linearity,reproducibility,stability and sensitivity,which was suitable for the routine analysis of anthraquinones in four medicinal aloe.In addition,the LC-ESI-MS/MS analysis results indicated that some anthraquinones such as aloesin,isoaloesin and aloenin A can be used for the differentiation of four medicinal aloe.In the third chapter,the proteins in four medicinal aloe were analyzed by SDS-PAGE,IEF-PAGE,2-DE and MALDI-TOF-MS.Five extraction methods were compared for the preparation of aloe protein.The results of protein content and SDS-PAGE analysis showed that the TCA/acetone-saturated phenol/SDS combined extraction method was the best extraction method for aloe protein.Furthermore,the SDS-PAGE results showed that the number of protein bands and the optical density curves of the four medicinal aloe were slightly different.And 2-DE results showed there were 54 differentially expressed proteins in four medicinal aloe,of which 50 were successfully identified by MALDI-TOF-MS.In addition,some specific proteins may closely related to morphological characteristics of four medicinal aloe.For example,the ran-binding protein 1 homolog c-like,actin,NAD-dependent malate dehydrogenase and cinnamyl alcohol dehydrogenase in A.barbadensis may be one of the components that produce the leaf hypertrophy and lage water content of their leaves;the alpha tubulin subunit and isoflavone reductase-like proteins in A.vera var.chinensis may be one of the components that produce white spots on the surface of their leaves;the auxin-induced protein PCNT115-like isoform 1 in A.arborescens may be one of the components that producing lengthwise growth,which is significantly higher than that of other aloe species.The fourth chapter was focused on the in vitro thrombolytic and fibrinolytic activities tests of the active ingredients(anthraquinones and proteins extracts,and monomeric anthraquinone compounds)in four medicinal aloe.The results showed that the anthraquinone and protein extracts of the four medicinal aloe all exhibited a certain degree of in vitro thrombolysis and fibrinolytic activity.The activity of the protein extracts of A.ferox was significantly higher than that of other three aloe,and the activity of the anthraquinone extracts of A.barbadensis was significantly higher than that of other three aloe.On the other hand,aloin A,aloin B,aloin(A/B)and aloe-emodin also showed a certain degree of in vitro thrombolytic and fibrinolytic activity.These results can provide scientific references for the practical uses of four medicinal aloe.The fifth chapter is conclusion and prospective of the present study.The systematic comparison analysis of anthraquinones and proteins in four medicinal aloe,which not only enriches the understanding of the chemical composition of the two major compounds in aloe,but also provides some reference for the identification of aloe.On the other hand,the evaluation of in vitro thrombolytic and fibrinolytic activity of aloe can provide a scientific basis for the application of aloe in thrombolysis.