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SEPARATION OF THE STANDARD SAMPLE LIPID LAYER IN A HIGH CONCENTRATION SALT SOLUTION BY THIN-LAYER-CHROMATOGRAPHY AND SOLID-PHASE-EXTRACTION
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Abstract
The published TLC, SPE processes can well separate lipid subclasses from biological samples in nature such as (lipids in food, lipids in the human serum...). We have applied lipid analysis in solutions with high alkaline salt concentration (DDM: biological tissue preservation solution), by these TLC, SPE processes; however, the SPE procedure according to Kaluzny and Agren was not repeatable for lipid samples extracted from DDM, the results were not obtained as announced by the author; The obtained TLC results are in agreement with a number of widely accepted publications, but it is difficult to identify cholesterol with DG in the mixed sample analysis results. This is clearly a limitation if the target ingredients are DG and cholesterol. Through the survey, we improved TLC (Institute 69) one step with solvent mixture B (C6-DEE-methanol-acetic acid; 90:20:3:2) due to its simplicity and good separation of standard lipid layers. single as well as mixed. In case it is necessary to separate more MG components, DG can go one step further with a mixture containing polar solvents, we have obtained improved results when expanding one step further with C6-DEE- acetone (60:40:5) to 12 cm. Objective: To evaluate the efficiency of separating lipid components in alkaline salt solution by solid-phase extraction and thin layer chromatography, to select and optimize the application process of lipid analysis in saline solution. Subjects and methods: Conducted on 6 standard lipid samples including FFA (C12:0 - C22:0; Sigma); glycerol acyl with 1 to 3 fatty acids (C14:0 - C18:0) (Sigma); a mixture of 37 FAMEs (Supelco), a mixture of PL phospholipids including PC and PE; 6 standard lipid samples, 6 adipose tissue lipid samples preserved in saline solution (DDM) were repeated n=6 for each sample.Revised TLC Process at Institute 69. Experimental descriptive research method. Results: With the method of Institute 69 for stable results, high repeatability and good separation of lipid layers: Phospholipid; monoglycerides; diglycerides; cholesterol; free fatty acids; triglycerides; cholesterol esther has Rf respectively: 0; (0.0 - 0.03); (0.11 - 0.28); (0.12 - 0.20); (0.33 - 0.52); (0.66 - 0.75); (0.80 - 0.90). The lipid layers from the TLC plate were also obtained quite simply for further analysis. Allows good application for separation of lipids in DDM at Institute 69. Conclusion: Application of the modified TLC method at Institute 69 can effectively separate the soluble lipid layers in DDM, which is an important step in sample preparation for intensive lipid analyzes by gas chromatography, high-performance liquid chromatography
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Keywords
thin layer chromatography (TLC), solid -phase extraction (SPE), lipid, saline solution (DDM)
References
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