A high-performance liquid chromatography method for simultaneous detection of melanin and inosinic acid content in black-bone chicken
A technology of high performance liquid chromatography and melanin, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of complex and time-consuming operation, low recovery rate, and low separation degree of thin-layer chromatography, and achieve shortened detection time, The effect of less sample volume and sufficient sample separation
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Embodiment 1
[0029] Materials and Methods
[0030] 1. Reagent and solution preparation
[0031] Methanol (chromatographically pure); 30%H 2 o 2 (excellent grade); ethanol, ether, triethylamine, phosphoric acid, hydrochloric acid (analytical grade); 5,6-dihydroxy-2-indolecarboxylic acid (CAS: 352195-40-5, Sigma); inosine acid (CAS: 1953-54-4, Sigma Company); squid melanin (CAS: 8049-97-6, Sigma Company); trypsin, papain (Invitrogen Company).
[0032] 1mol / L potassium carbonate (K2 CO 3 ) solution: 138g K 2 CO 3 , dissolved in water, and adjusted to 1000mL.
[0033] 10% sodium sulfite (Na 2 SO 3 ) solution: 10g Na 2 SO 3 , dissolved in water, and adjusted to 100mL.
[0034] Phosphoric acid / triethylamine buffer solution: dissolve 3.5mL phosphoric acid and 7.2mL triethylamine in 800mL water, adjust the pH to 6.5, and dilute to 1000mL.
[0035] Phosphate (PBS) buffer: 8g NaCl, 0.2g KCl, 1.44g Na 2 HPO 4 and 0.24g KH 2 PO 4 Dissolve in 800mL of water, adjust the pH value of the s...
Embodiment 2
[0056] Embodiment 2 precision test
[0057] Draw 20 μL of inosinic acid / PTCA mixed standard solution with a concentration of 50 / 5 μg / mL, repeat the injection 6 times, and measure the RSDs of the peak areas of PTCA and inosinic acid to be 0.13% and 0.11% respectively (n=6), indicating that The method has good precision.
[0058]
Embodiment 3
[0059] Embodiment 3 stability test
[0060] At 0, 2, 4, 6, 8, and 16 hours, 20 μL of the same test solution was drawn and injected, and the RSDs of the peak areas of PTCA and inosinic acid were 0.97% and 1.72% (n=6) respectively within 16 hours. ), indicating that the detection value of the test solution is stable within 16 hours.
[0061]
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