Abstract: This article discusses the method of determining the content of acid orange â…¡ and acid golden in soybean products by high performance liquid chromatography. The acid golden â…¡ in soybean products is crushed, extracted and filtered. The resulting sample extract is determined by high performance liquid chromatography, mobile phase It is 0.02 mol / L = 75 25 of methanol ammonium acetate. When the detection wavelength is 450 nm, the flow rate is 1.0 ml / min. The column is SymmetryC18.
Acid orange â…¡ and acid golden in soybean products are colorants used in medicine for tissue dyeing, wooden furniture, wool, cloth, etc., but illegal vendors use their bright colors, stable coloring, and low prices to illegally add to soybean products In this case, it has seriously endangered the health of consumers [1]. There is no corresponding standard test method in this country. This article has preliminarily established a method for the determination of acid orange â…¡ and acid golden by high performance liquid chromatography. , Simple detection of acid orange â…¡ and acid golden, can meet the needs of food inspection.
1 Experimental part 1.1 Instruments and reagents 1.1.1 instruments High performance liquid chromatograph includes Waters 1525 type pump, 2487 type ultraviolet detector, Breeze chromatography workstation and manual sampler.
1.1.2 Standard acidic orange II (purity 86%) and acid golden (industrial purity 90%).
1.1.3 Methanol chromatographic purity.
1.1.4 Pure water High purity water provided by AKUPâ…¡I20 pure water machine.
1.1.5 0.02 mol / L ammonium acetate solution, pH = 4.7.
1.1.6 Alkaline extraction solution absolute ethanol: ammonia water: water = 70:20:10.
1.2 Chromatographic conditions The column is SymmetryC18 (250 mm × 4.6 mmid, 5 μm) mobile phase methanol: 0.02 mol / L ammonium acetate solution = 75 25, solution flow rate 1.0 ml / min, column temperature is normal temperature, detection wavelength 450 nm, Sample volume 20 μl.
1.3 Preparation of standard products Pipette the appropriate amount of mixed standard stock solution 1.00 mg / ml into a series of standard concentrations of 0, 0.01 μg / ml, 0.05 μg / ml, 0.10 μg / ml, 0.50 μg / ml, 1.00 μg / ml .
1.4 Sample preparation Weigh and mix the crushed sample 5.00 g ~ 10.00 g in a 100 ml beaker, add 50 ml of water with pH = 6, mix and filter, add 1.00 g of polyamide powder to the filtrate, G3 funnel suction filtration, pH = 6 water rinse 2 ~ 3 times, alkaline extraction solution desorption 2 ~ 3 times, 5 ml each time, collect the desorption solution, dry in a water bath, dissolve the residue in distilled water, dilute to 5 ml, then over 0.45 μm filter membrane. The standard and sample filtrates were injected into the HPLC system and determined according to the above chromatographic conditions. Qualitative by retention time, peak area quantification.
2 Results 2.1 Wavelength selection Dilute the standard solution with the mobile phase and perform an ultraviolet scan. From the scan spectrum, it can be seen that Acid II and Acid Golden have a large absorption at 450 nm, so we selected 450 nm as the detection wavelength.
2.2 Working curve and detection limit The linear regression of the chromatographic peak area Y with the working fluid content X shows that the acid orange Ⅱ and the acid golden color have a good linear range in the range of 0.11 μg / ml ~ 1.00 μg / ml. Regression equation, linear range, correlation coefficient and detection limit (S / N = 3) 2.3 method precision (n = 6)
2.4 Prepare a sample containing acid orange Ⅱ (purity 86%) and acid golden yellow 0.05 μg / ml, and inject it into the HPLC system for six determinations according to the above chromatographic conditions. The results show that the method has good precision (RSD is 2.06%, 1.18%).
3 Discussion This article has made a preliminary discussion on the determination of acid orange â…¡ and acid golden by high performance liquid chromatography, and the understanding is still superficial.
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