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Analysis of Fatty Acid Composition in Overall Lipid Content

            of Human ES Cells                                                                                                                                                                                                                  Subcutaneous Tissue  Mass Imaging of


              Data
                                                                                                                                      Docosahexaenoic acid (DHA) is a fatty acid present in large quantities in the
                                                                                                                                      brain. It accounts for 40% of the fatty acids in the brain and 60% of the fatty
                  In an effort to establish practical applications for regenerative medicine, research is actively being conducted on ES cells and iPS cells.  acids in the retina. Based on experiments with DHA-deficient mice, DHA
                  Due to the undeveloped mitochondria in undifferentiated ES cells, the primary energy source is from the glycolytic system. When   deficiency is reported to affect brain and nerve system development.                       Analysis of
                  mitochondria develop as the ES cells become differentiated, the TCA cycle is said to become the energy source.      Consequently, DHA has attracted attention as a component in dietary                                        Comprehensive
                  This example describes the results from analyzing the composition of fatty acids in human ES cells. After extracting all cellular lipids   supplements, which are claimed to have benefits such as reducing the risk of    Glycerophospholipids
                  from frozen pellets of human ES cells, the fatty acids were methyl-esterified. Analyzing the methylated fatty acids by GC-MS resulted   cardiovascular disease, reducing neutral fats in the blood, improving blood
                  in identification of 19 types of fatty acids.                                                                       pressure, aiding growth and behavioral and visual development in infants, and
                                                                                                                                      reducing the symptoms and frequency of depression. Many research results
                                    (×10,000,000)                                                                                     have been reported on the effectiveness of such functions.
                                 3.0
                                    TIC                                                                                                                                                                                                      Technology
                                 2.5                                                                                                                                                                                                           Supercritical Fluid   Lipid Analysis Using


                                 2.0

                                 1.5


                                 1.0                                                                                                                                                                                                             Analysis of
                                                                                                                                            UFMS Technology                                                                                    Glycerophospholipids
                                 0.5                                                                                                        Gas chromatograph mass spectrometers (GC-MS) are especially
                                                                                                                                            effective at analyzing the many different molecular species of   Smart Metabolites Database
                                                                                                                                            fatty acids. GC-MS analysis requires optimizing analytical
                                 0.0                                                                                                                                                                      Pyruvic acid  Acetyl
                                                                                                                                                                                                            ac
                                                                                                                                                                                                            aci
                                                                                                                                                                                                            a
                                                                                                                                                                                                                Acetyl C-CoA
                                                                                                                                                                                                                Acety
                                                                                                                                                                                                                Acety
                                                                                                                                                                                                            aci
                                                                                                                                                                                                            aci
                                                                                                                                                                                                            a
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                                             45.0      50.0      55.0       60.0      65.0                                                  conditions, such as the method used to derivatize analytical             Citric acid
                                                                                                                                            samples and the analytical column used. To eliminate tedious
                                                                                                                                                                                                                               aci
                                         Fig. 1   TIC of Methyl Ester Derivatives from Human ES Cell Extract Sample                                                                                           Oxalacetic acid  Aconitic acid d  Blood Serum
                                                                                                                                            jobs and streamline these analytical processes, Shimadzu has                                       Mediators in Human   Analysis of Lipid
                                                                                                                                                                                                               a
                                                                                                                                                                                                              ali
                                                                                                                                                                                                              ali
                                                                                                                                                                                                               acid
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                                                                                                                                                                                                             Malic
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                                                                                                                                                                                                             Malic acid
                                                                                                                                                                                                             Malic
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                                                                                                                                                                                                               acid
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                                                                                                                                                                                                             Malic
                                                                                                                                            established a database of various fatty acid and metabolite      Malic  l ic ic c c c  acid i d d  Isocitric acidd
                                                 Table 1   List of Detected Fatty Acid Methyl Esters                                        components and offers columns and analytical conditions         Fuumaricc acid d
                                                                                                                                                                                                             um
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                                                                                                                                                                                                               c acid
                                                                                                                                                                                                            Fuma
                                                                                                                                                                                                                             2-Ketooglutarric
                                                                                                                                                                                                                              -Ket
                                                                                                                                                                                                                             2-Ket o glut
                                                                                                                                            suited to target compounds.                                                        accid
                         Compound Name                    Retention Time (min)  Retention Index  Similarity*  S/N                           Using the Smart Metabolites Database allows GC-MS and            Succinic acic acid d d  Succinyl-CoA A
                                                                                                                                                                                                                c
                                                                                                                                                                                                              ucc
                                                                                                                                                                                                               ic ac
                                                                                                                                                                                                              cc
                                                                                                                                                                                                              ccin
                                                                                                                                                                                                             Succin
                                                                                                                                                                                                              ucc
                                                                                                                                                                                                                              yl
                                                                                                                                                                                                                               C
                                                                                                                                                                                                                            Succin
                         Methyl myristate                     41.925        2204       98       380                                         GC-MS/MS analysis of a wide variety of fatty acids.
                         Methyl pentadecanoate                43.755        2309       99       372
                         Methyl palmitate                     45.505        2413       98      12739                                                                                                                                           of Glycolipids
                         Methyl palmitoleate                  46.760        2490       95       143                                                                                                                                              Structural Analysis
                         Methyl margarate                     47.150        2515       98       606
                         Methyl stearate                      48.735        2619       98      29218
                         Methyl oleate                        49.790        2689       96        96
                         Methyl linoleate                     51.300        2793       99      1101
                         Methyl arachisate                    51.690        2821       98       811
                         Methyl cis -11-icosenoate            52.675        2891       99      1602
                         Methyl linolenate                    53.005        2913       98       122
                         Methyl cis -11,14-Icosadienoate      54.165        2991       98       124
                         Methyl eicosa-8,11,14-trienoate      55.315        3069       99       632                                                                                                                                              Analysis of Fatty Acid
                         Methyl erucate                       55.535        3083       98        93                                                                                                                                          Content of Human ES Cells  Composition in Overall Lipid
                         Methyl cis -11,14,14-Icosatrienoate  55.910        3108       97        24
                         Methyl arachidonate                  56.240        3130       99       457
                         Methyl cis -5,8,11,14,17-Eicosapentaenoate  58.230  3254      99       243
                         Methyl cis -7,10,13,16,19-docosapentaenoate  61.995  3478     98       617
                         Methyl cis -4,7,10,13,16,19-Docosahexaenoate  63.150  3547    99        62
                                                                                                                                                                      Triple Quadrupole Gas Chromatograph Mass Spectrometer
                         Methyl margarate-d3                  47.085        2511       97      19864                                                                            GCMS-TQ8040
                         *Similarity reflects the results obtained in the NIST mass spectrum library search.                                                                                                                                 Esters by GC  Acid Methyl   Analysis of Fatty
                  Reference: Technical Report “Determination of Fatty-acid Composition of Lipids in Human Embryonic Stem Cells Using GC-MS” (C146-E243)  Reference: GCMS-TQ8040 brochure (C146-E251), Smart Metabolites Database flyer (C146-E277)
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