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Determination of amino acids in fodders and raw materials using capillary electrophoresis

Determination of amino acids in fodders and raw materials using capillary electrophoresis. Methods for analysis of AA contents in fodder s. Ion-exchange chromatography (in the amino acids analyzer) RP-HPLC with gradient elution Calculation method

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Determination of amino acids in fodders and raw materials using capillary electrophoresis

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  1. Determination of amino acids in fodders and raw materials using capillary electrophoresis WWW.LUMEX.RU

  2. Methods for analysis of AA contents in fodders • Ion-exchange chromatography (in the amino acids analyzer) • RP-HPLC with gradient elution • Calculation method ________________________________________________________________ • CE - only for free form of AAs (juice, beer) WWW.LUMEX.RU

  3. Express analysis of technologically important amino acids – Lys, Met, Thr, Cys-Cys, Trp. Complete analysis of proteinogenous amino acids (20AAs) 2 schemes of AAs analysis in fodders by CE Pre-capillary derivatization Direct detection (without derivatization) WWW.LUMEX.RU

  4. Direct determination of technologically important AAs • Detection • Optimization of separation, including: • on 190 nm (Lys, Met, Thr, Cys-Cys)acid hydrolysis • on 219 nm (Trp)alkaline hydrolysis • pH and composition of BGE • additives to the BGE (organic solvents, surfactants, macrocycles) WWW.LUMEX.RU

  5. CZE separation of the 19 underivatized amino acids Buffer: 10 mM borate with the addition 10 mM -CD (pH 9.18) Voltage: +20 kV; Detection:190 nm. WWW.LUMEX.RU

  6. Assay characteristics for technologically important amino acids WWW.LUMEX.RU

  7. The data of the underivatized amino acids determination in the real samples using HPLC, CZE and IEC WWW.LUMEX.RU

  8. Direct determination of Trp • Alkaline hydrolysis • Temperature 40°C • Detection on 219 nm: • increase of detection sensitivity • decrease of interference of concomitant AAs WWW.LUMEX.RU

  9. The complete analysis of proteinogenous amino acids • Only for derivative forms • PITC advantages: • simplicity of use • effective separation of the derivatives • sufficient reproducibility of the reaction condition • good solubility of the derivatives in water WWW.LUMEX.RU

  10. NH 2 S NH C pH 10.5 C S N R CH NH COOH CH R PITC AAs COOH PTC - AAs N S O PTH - AAs NH R Derivatization of AAs with PITC + WWW.LUMEX.RU

  11. CE analysis of PTC-AAs Voltage: 25 kV; BGE: phosphate with ß-CD Temperature: 20°C No pressure was applied WWW.LUMEX.RU

  12. CE analysis of PTC-AAs • Controlled pressure during analysis • Highly precise changeable temperature of a capillary cooling Voltage: 25 kV; BGE: phosphate with ß-CD Temperature: 30°C Pressure 30 mbar was applied after 17 min WWW.LUMEX.RU

  13. b-CD as a component of BGE Chiral activity for Trp, Tyr, Phe ! Protein molecules consist of L-AAs, but mixed fodders’ preparation is followed by insertion of synthetic AAs. Technology nonobservance – use racemates or D-AAs. Fodders quality control and falsifications reveal. WWW.LUMEX.RU

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