Method for batch-determining content of amino acid by using chimney-top 96-well PCR (Polymerase Chain Reaction) plate

Abstract

The invention belongs to the field of analytical chemistry, and aims at providing a method for batch-determining the content of amino acid by using a chimney-top 96-well PCR (Polymerase Chain Reaction) plate. According to the method provided by the invention, the content of amino acid in a sample solution is determined through a ninhydrin colorimetric method; and during the determination process,all small tubes on the chimney-top 96-well PCR plate made of a polypropylene material are used as containers for filling the sample solution and a ninhydrin reagent, and the ninhydrin reaction is carried out by heating the PCR plate which is supported by a hollowed-out ethylene vinyl acetate plastic pad in boiling water. Compared with the traditional ninhydrin colorimetric method, the method has the advantages that the operation is simpler and more convenient, labor force, time and the use amount of the reagent are saved, therefore the method can be widely applied to batch-determining of the content of amino acid in food, beverage, seasonings and healthcare products. Meanwhile, in the invention, the chimney-top 96-well PCR plate used in molecular biology research and diagnosis for batch-amplifying nucleic acid is used for the determination through the ninhydrin colorimetric method in analytical chemistry, and a new purpose of the chimney-top 96-well PCR plate is provided.

Description

technical field [0001] The invention belongs to the field of analytical chemistry, and in particular relates to a method for batch-determining amino acid content in samples by using a chimney-top 96-hole PCR plate in a ninhydrin colorimetric method. Background technique [0002] The reaction of amino acid and ninhydrin under heating conditions is a quantifiable color reaction. The color of the reaction product varies with amino acid, pH, temperature, reducing agent and metal ion strength, etc. Most amino acids react with ninhydrin to produce a blue-purple compound. The color of the compound is proportional to the content of the amino acid. By measuring the absorbance of the chromogenic compound at its absorption peak (usually at 570nm), the content of the amino acid can be determined. The ninhydrin colorimetric method is simple and reliable, and is widely used in the detection of amino acid content in food, beverages, condiments and health products. [0003] At present, th...

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Problems solved by technology

Generally, heat-resistant polypropylene plastic products have poor transparency and cannot meet the requirements for the absorbance of the solution in the detection hole.

[0004] Recently, manufacturers, such as Eppendorf in Germany, have made highly transparent microwell plates from polypropylene (the current price is about twice that of conventional polystyrene microwell plates), which can be used to detect the absorbance of the solution in the wells. ; but its entire bottom is the same as the conventional microwell plate, when heating with boiling water, the bottom of the peripheral well of the plate and the side of the edge of the plate can be heated at the same time, while only the bottom of the inner hole is heated, resulting in the difference between the periphery and the inner well. The solution has differences in heating, color development rate and evaporation. As a result, the absorbance of the same sample solution is different in the peripheral hole and the inner hole after reaction, resulting in a large detection error

Moreover, boiling water sometimes enters the pores and interferes with colorimetric determination

In addition, the heat-resistant microwell plate can be heated evenly with a preheated oven with precise temperature control, but in order to prevent the solution from evaporating under baking, it is necessary to cover the reaction solution with a layer of transparent oil that does not participate in the reaction (Efewmonkiekie W. Iyamu, Toshio Asakura, Gerald M. Woods. Analytical Biochemistry 2008, 383: 332-334), not easy to operate

[0005] Therefore, there is still a lack of easy, reliable and practical method for batch detection of amino acid content in samples by ninhydrin colorimetry

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