Method for measuring content of amylose and amylopectin in corn by using full-automatic flow injection analyzer method

Abstract

The invention discloses a method for determining the content of amylose and amylopectin in corn through a full-automatic flow injection analyzer method, and particularly relates to the field of determination of the content of amylose and amylopectin in corn, the method comprises the following steps: taking a corn kernel sample, crushing into powder, and passing through a 80-mesh screen to obtain corn kernel sample powder; the corn kernel sample powder is subjected to reflux extraction degreasing through a degreasing agent, and a degreased corn kernel sample is obtained; the method comprises the following steps: weighing a degreased corn kernel sample, putting the degreased corn kernel sample into a conical flask, adding a 95% ethanol solution, transferring a sodium hydroxide solution into the conical flask, and heating the conical flask in a boiling water bath for 10 minutes to obtain a to-be-detected corn sample with starch dispersed; taking out the to-be-detected corn sample, cooling to room temperature, transferring into a volumetric flask, adding distilled water to a constant volume, shaking uniformly, analyzing by a flow injection analyzer, and calculating to obtain the content of amylose in the corn sample. The method has the advantages of high automation degree, high detection speed and simple sample treatment.

Description

Technical Field

[0001] The present application relates to the field of determination of the content of amylose and amylopectin in corn, and in particular to a method for determining the content of amylose and amylopectin in corn using a fully automatic flow injection analyzer. Background Art

[0002] The amylose contained in corn is broken down into glucose by digestive enzymes in the human body and can be absorbed and utilized by the human body to provide energy for the body. Compared with amylopectin, amylose is digested and absorbed relatively slowly in the human body. It is a carbohydrate that releases energy slowly, has a relatively low glycemic index, and causes a slow increase in blood sugar after consumption. Therefore, in the food industry, amylose can be used to make products that require slow digestion and have certain gel properties, such as some low-sugar, high-satiety functional foods. Therefore, the amylose content is one of the important indicators for measuring corn quality.

[0003] Compared with amylose, although amylopectin is more easily broken down into glucose by amylase in the human digestive system, resulting in a relatively rapid rise in blood sugar, amylopectin can be used by some beneficial bacteria in the intestines, promoting the growth and reproduction of beneficial bacteria and helping to maintain the balance of intestinal microecology. A good intestinal microecology has a positive effect on regulating body metabolism and enhancing immunity. In the long run, it may have a beneficial effect on the overall health of diabetic patients and indirectly help control blood sugar. Therefore, the determination of amylopectin content has important guiding significance for the adjustment of carbohydrate dietary structure.

[0004] For example, Chinese invention patent CN201911374176.2 discloses a method for detecting the amylose content of a single grain of wheat. This method uses an ELISA reader to read the absorbance and calculate the percentage of amylose.

[0005] Another example is the method for determining the content of amylose in sweet potato starch disclosed in CN202211379761.3. This method uses a continuous flow analyzer to detect the absorbance values ​​of the sample dispersion and standard dispersions of different concentrations to calculate the percentage of amylose.

[0006] For the first patent document, the main defects of this method are cumbersome operation, long time consumption, poor accuracy and repeatability. For the second patent document, the main defect of this method is that the oil contained in the sample easily interferes with the sample test results. Therefore, how to design a method suitable for determining the content of amylose in corn samples is an urgent problem to be solved by those skilled in the art. Summary of the invention

[0007] The invention provides a method for determining the contents of amylose and amylopectin in corn by using a fully automatic flow injection analyzer. The method has the advantages of high automation, fast detection speed and simple sample processing.

[0008] The objective of the present invention is achieved through the following technical solutions: The method for determining the content of amylose and amylopectin in corn by fully automatic flow injection analyzer comprises the following steps: S01. Sample preparation: Take a corn kernel sample, crush it into powder, and pass it through an 80-mesh sieve to obtain a corn kernel sample powder; S02. Sample defatting: The corn kernel sample powder is subjected to reflux extraction and defatting by a degreasing agent to obtain a defatted corn kernel sample; S03. Preparation of test solution: weigh defatted corn kernel sample into a conical flask, add 95% ethanol solution, transfer sodium hydroxide solution into the conical flask, and gently shake to obtain a mixture; The obtained mixture was placed in a boiling water bath and heated for 10 min to obtain a corn sample to be tested with starch dispersed; S04. On-machine analysis: Take out the corn sample to be tested, cool it to room temperature, transfer it to a volumetric flask, add distilled water to make up the volume and shake it to mix well. Analyze it with a flow injection analyzer to calculate the content of amylose in the corn sample.

[0009] Preferably, the analysis conditions of the flow injection analyzer are: injection rate: 30 samples / h; injection: cleaning ratio = 2:1; heating distillation device: temperature stable at 145°C; flow system: no leakage, regular bubbles, smooth flow of reagents; data processing system: straight baseline.

[0010] Preferably, the degreasing agent is selected from any one of methanol / petroleum ether.

[0011] Preferably, the blank solution is prepared by using the same operating steps and reagents as those used for measuring the sample, but using 5.0 mL of 0.09 mol / L sodium hydroxide solution instead of the sample to prepare the blank solution.

[0012] Preferably, the preparation steps of the standard curve are: replace corn flour with potato amylose and amylopectin standards, prepare according to steps S02-S03, obtain potato amylose and 1 mg / mL amylopectin standard solutions, respectively, and mix them as follows to prepare 10 mL of amylose standard solution.

[0013] Preferably, in step S03, the concentration of sodium hydroxide is 1.0 mol / L.

[0014] Compared with the prior art, the advantages or beneficial effects of the technical solution of the present application include: 1. High degree of automation: This method uses peristaltic pump technology for automatic sampling, which can be continuously sampled and tested without being limited by the number of samples, reducing manual operation errors and improving the accuracy and repeatability of the test. The detection process is controlled by software, which automatically completes the mixing, reaction, detection and result output of samples and reagents without manual intervention, greatly improving the detection efficiency and reducing the workload of operators.

[0015] 2. Fast detection speed: FIA technology based on non-steady-state chemical reactions can quickly complete sample detection. It is more efficient than traditional methods such as iodine colorimetry and can process a large number of samples in a short time to meet the needs of large-scale sample detection.

[0016] 3. Simple sample pretreatment: The sample pretreatment method is simple and in line with international standards, which simplifies the degreasing operation, reduces the pretreatment time and workload, and also reduces the errors caused by the complexity of the pretreatment process.

[0017] 4. Good safety: The reaction is carried out in a closed pipeline, which prevents operators from contacting toxic and harmful reagents, reduces safety risks, and reduces the pollution of the environment caused by the volatilization of reagents.

[0018] 5. Convenient data management: Equipped with a data management platform, the test data can be stored, queried, counted and analyzed, realizing visual management of data and long-term and short-term analysis of food safety, which is convenient for quality traceability and management. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 It is a curve chart of amylose content in three corn samples; Figure 2 Prepare a table for the standard solution preparation for the standard curve series. DETAILED DESCRIPTION

[0020] The following will describe the implementation methods of the present application in detail with reference to the accompanying drawings and embodiments, so that the implementation process of how the present application applies technical means to solve technical problems and achieve corresponding technical effects can be fully understood and implemented accordingly. The embodiments of the present application and the various features in the embodiments can be combined with each other without conflict, and the technical solutions formed are all within the protection scope of the present application.

[0021] It should be clear that the embodiments described below are only part of the embodiments of the present application, rather than all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by technicians in the relevant field without making creative work are within the scope of protection of the present application. Example

[0022] A method for determining the content of amylose in corn by flow injection instrumentation comprises the following steps: (1) Sample preparation: Take at least 10 g of corn kernels, grind them into powder using a cyclone mill, and pass them through an 80-mesh sieve.

[0023] (2) Sample degreasing: Weigh 5-7 g of sieved sample and degrease it by reflux extraction with methanol / ethanol solution.

[0024] (3) Preparation of test solution: Weigh 100 mg ± 0.5 mg of defatted sample into a 10 mL conical flask, carefully add 1 mL of 95% ethanol solution to the sample, and wash off the sample adhering to the flask wall. Pipette 9 mL of 1.0 mol / L sodium hydroxide solution into the conical flask and shake gently, then heat the mixture in a boiling water bath for 10 min to disperse the starch. Remove and cool to room temperature, then transfer to a 100 mL volumetric flask. Add distilled water to make up to volume and shake vigorously to mix.

[0025] (4) Preparation of blank solution: Use the same operating procedures and reagents as those for the sample determination, but use 5.0 mL of 0.09 mol / L sodium hydroxide solution instead of the sample to prepare the blank solution.

[0026] (5) Standard curve drawing: Substitute potato amylose and amylopectin standard products for corn flour and prepare according to (3) to obtain 1 mg / mL potato amylose and 1 mg / mL amylopectin standard solutions, respectively. Figure 2 Mix to prepare 10 mL of amylose standard solution. Figure 2 shown.

[0027] (6) Determination of maximum absorption wavelength: The UV-visible spectrophotometer was used to scan the standard solution of amylose and the standard solution of amylopectin. The maximum absorption wavelength of amylose was 623 nm, and the maximum absorption wavelength of amylopectin was 536 nm.

[0028] (7) Flow injection analysis: Turn on the flow injection analyzer, preheat and stabilize, adjust the instrument to normal working state according to the manual, and set the measurement wavelength to 620 nm. Adjust the pump speed according to the instrument specifications and clean the pipeline for 10 min~20 min. Connect the pipelines to the sodium hydroxide working solution (0.09 mol / L), iodine reagent (2g potassium iodide + 0.2g iodine, fixed to 100mL volumetric flask) and acetic acid (1 mol / L) solution according to the labels. Prepare a sample table according to the instrument requirements, and place the amylose standard solution and sample solution in the injection rack in sequence. Measure after the baseline is stable.

[0029] (8) Calculation of results: According to the absorbance of the sample solution, the corresponding amylose content is found on the standard curve, and the amylose content in the corn sample is calculated. Figure 1 . Figure 1 The linear regression equation is y=0.00926242x+0.140424, where the correlation coefficient r2=0.99881.

[0030] Optimization of flow injection conditions: including optimization of parameters such as flow rate, temperature, reaction time, etc., to improve the accuracy and repeatability of the measurement.

Claims

1. A method for determining the content of amylose and amylopectin in corn by using a fully automatic flow injection analyzer, characterized in that: The steps include: S01. Sample preparation: Take a corn kernel sample, crush it into powder, and pass it through an 80-mesh sieve to obtain a corn kernel sample powder; S02. Sample defatting: The corn kernel sample powder is subjected to reflux extraction and defatting by a degreasing agent to obtain a defatted corn kernel sample; S03. Preparation of test solution: weigh defatted corn kernel sample into a conical flask, add 95% ethanol solution, transfer sodium hydroxide solution into the conical flask, and gently shake to obtain a mixture; The obtained mixture was placed in a boiling water bath and heated for 10 min to obtain a corn sample to be tested with starch dispersed; S04. On-machine analysis: Take out the corn sample to be tested, cool it to room temperature, transfer it to a volumetric flask, add distilled water to make up the volume and shake it to mix well. Analyze it with a flow injection analyzer to calculate the content of amylose in the corn sample.

2. The method for determining the content of amylose and amylopectin in corn by the fully automatic flow injection analyzer method according to claim 1, characterized in that: The analysis conditions of the flow injection analyzer are as follows: injection rate: 30 samples / h; injection: cleaning ratio = 2:1; heating distillation device: temperature stable at 145°C; flow system: no leakage, regular bubbles, smooth flow of reagents; data processing system: straight baseline.

3. The method for determining the content of amylose and amylopectin in corn by the fully automatic flow injection analyzer method according to claim 1, characterized in that: The degreasing agent is selected from any one of methanol or petroleum ether.

4. The method for determining the content of amylose and amylopectin in corn by the fully automatic flow injection analyzer according to claim 1, characterized in that: The preparation steps of the blank solution are as follows: the same operation steps and reagents as those used for the sample determination are used, but 5.0 mL of 0.09 mol / L sodium hydroxide solution is used instead of the sample to prepare the blank solution.

5. The method for determining the content of amylose and amylopectin in corn by the fully automatic flow injection analyzer according to claim 1, characterized in that: The preparation steps of the standard curve are as follows: replace corn flour with potato amylose and amylopectin standards, prepare according to steps S02-S03, obtain 1 mg / mL potato amylose and 1 mg / mL amylopectin standard solutions, respectively, and mix them as follows to prepare 10 mL of amylose and amylopectin standard solutions.

6. The method for determining the content of amylose and amylopectin in corn by the fully automatic flow injection analyzer method according to claim 1, characterized in that: In step S03, the concentration of sodium hydroxide is 1.0 mol / L.

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