Trích:
Nguyên văn bởi angiangchem
Cho hỏi tí nhẹ
khi phân tích vitamin A (retinol) bằng đầu dò UV
Ta lấy retinol làm chuẩn. sau khi xử lý mẫu và đo thì các loại khác như vitamin A acetat, palmetat có được định lượng chung với retinol hay khong?
Nếu lấy vitamin A acetat làm chuẩn thì có được không? có cần xử lý gì không?
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theo mình thì khi phân tích vitamin A (retinol) bằng HPLC với đầu dò UV.
lấy Retinol làm chuẩn để định lượng các vitamin A dưới dạng retinol acetate, retinol propionate, retinol palmitate) đều được, nhưng bạn cần chú ý lúc tính toán phải đưa KLPT(khối lượng phân tử) vào CT tính kết quả định lượng. như theo phương pháp sau
Assay for vitamin A
Method 1
— [ N OTE — Where the use of a vitamin A ester (retinyl acetate
or retinyl palmitate) is specified in the following procedure, use the chemical form present in the formulation. USP Vitamin A RS is all- trans retinyl acetate. It is to be used where USP Vitamin A RS is specified.
Mobile phase— Use n-hexane.
Standard preparation— Dissolve an accurately weighed quantity of USP Vitamin A RS in n-hexane, and dilute quantitatively, and stepwise if necessary, to obtain a solution having a known concentration of about 15 µg of retinyl acetate per mL.
Assay preparation- Dissolve an accurately weighed quantity retinol (retinyl acetate
or retinyl palmitate) with n-hexane to obtain a solution having a final concentration of
about 15 µg of vitamin A per mL.
Chromatographic system — The liquid chromatograph is equipped with a 325-nm detector and a 4.6-mm × 15-cm column that contains 3-µm packing L8 (An essentially monomolecular layer of aminopropylsilane chemically bonded to totally porous silica gel support, 10 mm in diameter, column NH2). The flow rate is about 1 mL per minute.
method 2
Carry out the assay as rapidly as possible, avoiding exposure to actinic light and air, oxidising agents, oxidation catalysts (e.g. copper , iron), acids and prolonged heat.
Test solution (a) Introduce 0.200 g of the preparation to be examined into a 100 ml volumetric flask. Add 20-30 mg of bromelains R, 5.0 ml of water R and 0.15 ml of 2-propanol R. Heat gently in a water-bath at 60 °C for about 5 min, swirling occasionally. Add 40 ml of 0.1 M tetrabutylammonium hydroxide in 2-propanol . Swirl gently and let the mixture react for 10 minutes at 60-65 °C for hydrolysis, swirling occasionally. Ensure that all sample material is wetted. Allow to cool to room temperature, dilute to 100.0 ml with 2-propanol R containing 1 g/l butylhydroxytoluene R, and homogenise carefully to avoid air-bubbles. The solution may be turbid.
Test solution (b) Dilute test solution (a) with 2-propanol R to a final concentration of 100 IU/ml. Filter before injection.
Reference solution (a) Introduce about 0.100 g of retinyl acetate CRS into a 100 ml volumetric flask and dissolve immediately in 5 ml of pentane R. Add 40 ml of 0.1 M tetrabutylammonium hydroxide in 2-propanol . Swirl gently and let the mixture react for 10 minutes at 60-65 °C for hydrolysis, swirling occasionally. Cool to ambient temperature, dilute to 100.0 ml with 2-propanol R containing 1 g/l butylhydroxytoluene R, and homogenise carefully to avoid air-bubbles.
Reference solution (b) Introduce into a 50 ml volumetric flask 5.0 ml of reference solution (a) and dilute to 50.0 ml with 2-propanol R. Homogenise carefully to avoid air-bubbles.
Column:
—size: l = 0.125 m, Ø = 4 mm,
—stationary phase: octadecylsilyl silica gel for chromatography R (5 µm).
Mobile phase water R, methanol R (5:95 V/V).
Flow rate 1 ml/min.
Detection Spectrophotometer at 325 nm.
Injection 10 µl of test solution (b) and reference solution (b).
Run time 1.5 times the retention time of retinol.
Retention time Retinol = about 3 min.
Calculate the content of vitamin A from peak of test solution (b) and reference solution (b).
còn nếu bạn lấy vitamin A acetat làm chuẩn thì cũng chẳng có ảnh hưởng gì hết, miễn là khi tính toán bạn cần để ý đến khối lương phân tử.
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Linear Mode
