Uptake Of A Mixture Of Amino Acids On Mouse Embryo Development In Vitro

Objective:Preimplantation embryos can produce and consume amino acids in a manner dependent upon the stage of development that may be predictive of subsequent viability. this experiment to study the amino acids from the medium of net consumption and net amino acid production rate,In the study of embryonic development is important, and to provide more follow-up study, high-quality embryos.Methods:1 By a reversed-phase high performance liquid chromatographic method, simultaneous determination of 17 free amino acids in the mouse embryo culture was carried out. The samples were derivatized with PITC. RT-HPLCgradient elution to detect 17amino acids appearance and depletion. 2. Studied in KSOM medium under the conditions added to amino acids, respectively, to preimplantation embryos cultured in different periods. Select a high rate of embryonic development of the cultivation of droplets will be removed embryos, draw lml medium as centrifuge tube pr eserved in-20℃. Through reversed-phase high performance liquid chromatography (HPLC)analysis of mouse preimplantation embryo in vitro culture medium in each period changes in amino acid concentration. The net rate of amino acid depletion or appearance varied according to amino acid (P< 0.001) and, apart from cysteine and glutamine.conclusions:1.Amino acids were linear over the range of 0.125～0.525pmol/μL with the peak areas and the correlated coefficient were above 0.9919. The recoveries of amino acids added were 95.48%-103.64%, except cystine. The detection limit of amino acids ranged from 1.25-8 to 8 pmol/ul. There were% RSD ware 1.21%-4.83%. The method is sensitive, accurate, and reliable for the determination of amino acids in culture.2.The embryonic development at different times in different amino acid content can be roughly divided into 5 types, Type I:amino acids having high rates of net appearance on 2-cell that reached a nadir on 4-cell or 8-16cell but ubsequently increased by blastocyst (glycine,ala-nine,leucine, threonine valine,phenyl lanine,tyrosine); TypeⅡ:those that exhibited a stea-dy fall in net depletion from 2-cell to blastocyst(glutamic acid and methionine); Type III:amino acids which showed a continuous fall in net appearance(Serine and lysine); TypeⅣ:those that exhibited lower rates of net appearance on 2-cell but subsequently increased from 4-cell to blastocyst(Aspartic acid, proline, tryptophan); TypeⅤ:those that exhibited lower rates of net appearance on 2-cell to 8-16 cell but depletion in blastocyst.(isoleucine);3 blastocyst rate of different changes in their amino acid consumption, results show that the higher the rate of blastocysts, the more obvious changes in amino acids.