| Objective:Metabolomics is a new science and has been widely used in the medical field. it observes the overall performance of all biological samples metabolites can be tested qualitatively or quantitatively through the use of Mass Spectrometry (MS), Chromatography, Nuclear Magnetic Resonance (NMR) analysis and other modern means of detection equipment. It is to investigate the pathological and physiological changes response caused by organism response to exogenous organisms and genetic variation. These changes response is linked with biological related events in metabolic information to detect chemical change and biological significance in living cells. AIDS is a result of infection with human immunodeficiency virus (human immunodeficiency virus, HIV) which causes chronic fatal diseases. It has shown clear upward trend in the world, and its impact on society has become a major global public health attention and social problems. This study has selected sample with Lung and Spleen Qi Deficiency and Damp and Hot of AIDS general syndrome, using metabolomics approach, looking for their specific metabolites, and the different material between different syndromes, to create a new tool for the TCM syndrome differentiation and quantification of TCM diagnosis, to provide advanced technical support for the effect of progress in the TCM diagnosis and prevention to AIDS.Methods:This study has selected sample with Lung and Spleen Qi Deficiency and Damp and Hot of AIDS general syndrome, Cases are from the patients with Lung and Spleen Qi Deficiency and Damp and Hot of AIDS syndrome in Henan Province, with diagnostic criteria of AIDS treatment guidelines recommended by the Ministry of Health in2005. There is no national standards or professional standards of TCM diagnostic criteria currently. Diagnostic criteria of previous project results HIV/AIDS Diagnostic Scale of common syndromes are used in the project.2ml urine in the morning of selected staff are collected as samples for test, LC-MS technique (HPLC-MS) is used to analyze composition of urine, SPSS13.0and SIMCA-P are used for data processing and analysis. Experimental method:Use EP tube collection met inclusion criteria of normal controls, lung spleen-deficiency syndrome group and hot and humid intrinsic card group morning urine2ml, put in-20℃save a week transferred to80℃refrigerator. Set aside.Urine early treatment:Urine treatment at room temperature, before1ml thaw, remove the urine,4000r/min centrifugal10min, take supernatant fluid. Precision remove the supernatant fluid300mu L, buy1.5ml centrifugal tube, join600muon L methanol, using vortex mixer mixers, then high-speed centrifuge about10min10min,14000r/min,4℃temperature control, take supernatant fluid with800mu L, compact muon m film filtration, asking the product solution.Performance liquid chromatographic conditions:Chromatographic column for Zorbax Agilent C18(150×2.1mm,3.1muon m); Mobile phase:0.2%glacial acetic acid water methanol gradient elution (see table2); Velocity:0.2ml/min; Feeding sample plate temperature:4℃; Enter the sample weight:20mu L; Column temperature:30℃; Chromatograms acquisition time not less than60min.Mass spectrometry conditions:Scanning mode:level1whole scanning, sheath gas velocity:40auxiliary air velocity:bar, bar, spray voltage:104kv, capillary voltage:-4v, capillary temperature:300℃, qualitative load ratios range:80-1000, the anion scanning mode.Analysis:using liquid mass coupled techniques (HPLC-MS) analysis by mass spectrometry, urine composition analysis instruments own software export raw data (quality than the peak intensity corresponding Hollywood for data files,) Excel screening, screening for:each card type the principle of quality same load ratios (data fluctuate in the definition of plus or minus1for the same kind material). Adopt SIMCA SPSS13.0software and data are software-P T test analysis, cluster analysis, and PCA pattern recognition analysis.Results:Export raw data (corresponding to cytoplasmic intensity ratio) was tested and analyzed by T test of SPSS13.0. P value <0.05has statistically significant meaning. The principal component score plot of metabolites showed there were significant differences in the main components between normal group and patient group. The load diagram showed there was significantly different substance between normal group and patient group and between AIDS patients with Lung and Spleen Qi Deficiency group and Damp and Hot group, Its characterization can not be determined. Comprehensive t-test and corresponding plot chart can be seen in AIDS patients in the normal group and load ratios of794, has for412,505,213,368,118,618,388,159,101,109,210,192,124material notable differences; In normal group and AIDS lienal lung qi difficiency card group than for412, Hollywood has 794,505,118,329,352,388,618,562,502,56,159,384,109,101,117,125,339material notable differences; In normal group and AIDS hot and humid intrinsic group than for412, Hollywood has794,368,157,125,109,210,124,192,182material notable differences; In AIDS lienal lung deficiency syndrome group and AIDS hot and humid intrinsic card group than for521, Hollywood has indeed,376,121,338,151notable differences in the matter.Conclusion:1. There is good distinction in urine metabolic profiling between AIDS patients and normal.2. Some potential markers that may exist were found, such as the material with mass to charge ratio of the794,412,388,118, which can help AIDS clinical diagnosis of micro-indicators, There is a certain role for its quantification,3. It is expected to be the new means of AIDS TCM diagnosis and syndromes differentiation. |
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