Food safety testing is an issue that everyone is concerned about, and the pretreatment of food samples is a key step that affects the test results. This article sorts out several sample pretreatment methods in food testing, hoping to help inspectors engaged in food testing.
Sample pretreatment: the process of sample preparation and extraction, purification and concentration of the components to be measured in the sample. In the detection and analysis of the whole food safety, 70% to 80% or even more time is spent on the pretreatment of the sample, and more than 60% of the errors brought to the experiment come from the pretreatment of the sample.
The purpose of sample pretreatment is to concentrate the measured substance, eliminate matrix interference, protect the instrument, and improve the accuracy, precision, selectivity and sensitivity of the method.
Main sample preparation methods
1. Ultrasonic extraction;
2. Microwave extraction;
3. Liquid-solid extraction;
4. Accelerated solvent extraction;
5. Supercritical extraction;
6. Solid phase extraction;
7. Solid-phase microextraction;
8. Matrix dispersive solid phase extraction;
9. Liquid-liquid extraction;
10. Microchemical technology;
11. Column chromatography.
Basic Requirements for Sample Preparation
1. Analysis of food hazard residues, features: complex matrix; detection limits of target compounds are becoming more and more stringent; certain hazardous residues exist in extremely low concentrations in food samples; the properties of each target compound are quite different; multiple substances may exist at the same time kinds of components.
2. To evaluate whether the pretreatment method is reasonable, the factors that should be considered are: whether the operation is simple and time-saving; whether the recovery rate of the tested components is high; whether the cost is low; whether it has an impact on the human body and the environment.
Extraction technology: use organic solvents and other methods to extract the analyte from the sample, and use it for determination after purification. Extraction technology requires solvents to selectively dissolve residual hazardous substances as much as possible, rather than dissolving or dissolving food matrix in a small amount. The key to the extraction effect is the choice of solvent. The extraction recovery rate of residual hazardous substances directly determines the accuracy of the entire analysis step.
What purification technologies include?
1. Liquid-liquid extraction;
2. Solid phase extraction;
3. Solid-phase microextraction;
4. Matrix dispersive solid phase extraction;
5. Column chromatography (gel, ion exchange, affinity, partition, adsorption column chromatography, etc.)
6. Immunological detection technology (molecular imprinting method)
Immunological detection technology is an analytical detection technology based on the specificity and reversible binding reaction of antigen and antibody. On this basis, combined with some biochemical or physical methods as a signal display or amplification system, an immunoassay can be established. That is, the antigen can be specifically combined with the antibody, and the detection can be carried out through the specific recognition reaction of the antigen-antibody.
Antibodies are immunoglobulins (Ig) that can specifically bind to antigens, and those related to immunoassays are mainly IgG and IgM. The reaction between antigen and antibody is combined by local intermolecular forces.
There are four main types: hydrogen bonding, van der Waals forces, electrostatic and hydrophobic interactions. Antigen-antibody binding reactions are highly specific, cross-reactive, and reversible. The physical and chemical properties of antigen-antibody binding are mainly manifested as the conversion from hydrophilic colloids to hydrophobic colloids.
A successful immunological assay must have three elements: an antibody with excellent performance, a sensitive and specific marker, and an efficient separation method. There are many existing immunoassay methods, which are classified according to different methods. Classical immunoassays do not require labels; labeled immunoassays are divided into radiolabeled assays and non-radioactively labeled assays, and are divided into homogeneous and heterogeneous immunoassays according to the reaction medium. Classical immunoassays are seldom used in the analysis of food safety and sanitation, while there are many kinds of highly sensitive heterogeneous and non-radioactive labeling immunoassays, which develop rapidly, have high detection sensitivity, and are widely used.
Sample pretreatment in food testing has become a major research direction in the food industry. In the process of sample testing, the treatment of test samples can ensure the authenticity and reliability of the results.
The first step in food testing is sample pretreatment!
1. Separate the measured components from complex samples and make them into solutions that are easy to measure.
2. Remove the matrix substances that interfere with the analysis and determination.
3. If the concentration of the component to be measured is low, enrichment is also required.
4. If the component to be measured is difficult to detect by the selected analytical method, it is necessary to quantitatively convert it into another compound that is easy to detect through sample derivatization.
Principles of pretreatment
1. Whether or not the sample needs to be pretreated, how to do it, and the sampling method should be considered according to the characteristics of the sample, the requirements of the test and the performance of the analytical instrument used.
2. Try not to use or use less pretreatment in order to reduce the operation steps, speed up the analysis, and also reduce the adverse effects brought about by the pretreatment process, such as the introduction of pollution, the loss of the analyte, etc.
3. When the decomposition method is used to treat the sample, the decomposition must be complete without loss of the components to be tested, and the recovery rate of the components to be tested should be sufficiently high.
4. The sample cannot be polluted, and the components to be measured and substances that interfere with the determination cannot be introduced.
5. The consumption of reagents should be as little as possible, and the method is simple and easy, fast, and has little pollution to the environment and personnel.
Small method for pretreatment solution preparation
When the measured component in the sample is in a free state – the solution is prepared by the dissolution method.
When the measured components in the sample are in a combined state – the solution is prepared by decomposition.
1. Dissolution method (to dissolve completely)
1) Water-soluble method:
Use water as a solvent, suitable for water-soluble components, such as inorganic salts, water-soluble pigments, etc.
2) Acidic aqueous solution leaching method:
The solvent is an aqueous solution of various acids, suitable for components that have increased solubility and stability in acidic aqueous solutions.
3) Alkaline aqueous solution leaching method:
The solvent is an alkaline aqueous solution, which is suitable for components whose solubility increases and is stable in an alkaline aqueous solution.
4) Organic solvent leaching method:
Suitable for components to be tested that are easily soluble in organic solvents. Commonly used organic solvents are ether, petroleum ether, imitation, acetone, n-hexane, etc. Organic solvents are selected according to the principle of “like dissolves like”.
2. Decomposition method
1) Dry ashing method
Advantage:
①Basically do not add or add a small amount of reagents, so the blank value is low;
②The volume of ash left after most foods are burnt is very small, so a larger amount of samples can be processed, and the sample volume can be increased, and the detection rate can be improved under the same method sensitivity;
③ Thorough decomposition of organic matter;
④The operation is simple, no one needs to keep watch during the ashing process, and the preparations for other experiments can be done at the same time.
Shortcoming:
① It takes a long time to process the sample;
②Due to open ashing and high temperature, it is easy to cause the loss of some volatile elements;
③The crucible containing the sample has a certain occlusion effect on the measured components. Due to the high temperature burning, the structure of the crucible material changes to form tiny holes, which makes it difficult for some measured components to be occluded in the holes, resulting in the measurement results and The recovery rate is low.
Measures to improve recovery rate:
①According to the properties of the components to be tested, adopt an appropriate ashing temperature
Ashing food samples should be carried out at the lowest possible temperature, but too low temperature will prolong the ashing time, usually choose 500-550℃ for 2h or 600℃ for 0.5h. Generally do not exceed 600 ℃.
② Add ashing fixative to prevent the volatilization loss of the measured components and the occlusion of the crucible.
2) Wet digestion method
Advantage:
① Due to the use of strong oxidants, the decomposition rate of organic matter is fast, and the time required for digestion is short;
②Because the heating temperature is lower than that of dry ashing, it can reduce the loss of metal volatilization and dissipation, and at the same time, the occlusion of the container is also less;
③ The measured substance is stored in the digestive juice in an ion state, which is convenient for the determination of various trace elements in it.
Shortcoming:
① During the digestion process, the rapid oxidation of organic matter often produces a large amount of harmful gases, so the operation needs to be carried out in a fume hood;
② In the initial stage of digestion, a large amount of foam overflow is likely to occur, so the operator needs to take care of it at any time;
③A large amount of various oxidants are used in the digestion process, the amount of reagents is large, and the blank value is high.
3) Commonly used digestion methods
In actual work, in addition to the digestion method of using sulfuric acid alone, several different oxidizing acids are often used in combination to make use of the characteristics of various acids and learn from each other to achieve the purpose of safely, quickly and completely destroying organic matter.
Several commonly used digestion methods are as follows:
① Digestion method used alone In this method, only sulfuric acid is added when the sample is digested, and in the case of heating, it relies on dehydration and carbonization to destroy organic matter.
② Nitric acid-high acid digestion method
③ nitric acid digestion method
What are the digestive manipulation techniques?
(1) Open digestion method
(2) Reflux digestion method
(3) Cold digestion method
(4) Sealed tank digestion method
Digestion operation should be noted:
(1) The reagents used for digestion should be high purity acids and oxidizing agents, containing few impurities, and at the same time do the blank test according to the same operation as the product to deduct the effect of digestion reagents on the determination data. If the blank value is high, the purity of reagents should be improved and better quality glassware should be selected for digestion.
(2) Glass beads or porcelain tiles can be added to the digestion flask to prevent flash boiling; the mouth of the Kjeldahl flask should be tilted so that it should not be facing itself or others. When heating, the fire should be concentrated at the bottom, and the neck part of the flask should be kept at a lower temperature to condense the acid mist and reduce the loss of volatilization of the measured components. If a large amount of foam is generated, in addition to quickly reducing the fire, a small amount of defoamer that does not affect the determination can be added, such as octanol, silicone oil, etc.; the sample and digestion solution can also be soaked overnight at room temperature and then heated and digested the next day.
(3) When it is necessary to add acid or oxidant during the heating process, stop the heating first, and then add it slowly along the bottle wall after the digestive juice is slightly cooled, so as to avoid violent reaction and splashing. In addition, adding acid at high temperature will cause the acid to volatilize rapidly, which is wasteful and pollutes the environment.
3. Common separation and enrichment methods
The main method is extraction: rapid operation, good separation effect, and wide application. However, the extraction reagent is usually flammable, volatile and toxic.
During extraction, especially when the solution is alkaline, emulsification often occurs and affects the separation. The methods to destroy emulsification are:
1) Standing for a longer period of time .
2) Gently rotate the funnel to accelerate the stratification.
(3) If the emulsification occurs due to partial mutual solubility of two solvents (water and organic solvents), you can add a small amount of electrolytes such as sodium chloride), the use of salt precipitation to destroy I If the emulsification occurs due to the small density difference between the two phases, you can also add electrolytes to increase the density of the aqueous phase.
(4) If emulsification occurs because the solution is alkaline, it can often be eliminated by adding a small amount of dilute hydrochloric acid or by filtration. Depending on the situation, emulsification can also be eliminated by adding etc. .
Solid phase extraction: divided into four steps: activation of sorbent, sample, washing and elution.
Other methods:
Solid-phase microextraction method
Supercritical fluid extraction method
Distillation and volatilization method
Membrane separation
The sample pretreatment is one of the important steps in the process of pesticide residue detection, which has an important impact on ensuring the accuracy and reliability of the determination results, reducing the contamination of the column and the detection instrument, and improving the detection efficiency. The residues of pesticides in food are generally in the range of 10-6~10-9 (W/W) or lower, which requires a fairly high sensitivity and selectivity of the detection method, but also puts forward higher requirements for the sample pretreatment technology.
The development direction of pretreatment
Different pretreatment techniques have their own advantages and disadvantages and the scope of application, in practice, according to the type of sample to be measured and the matrix, the determination of the results required and the different detection instruments, and combined with the actual conditions of the choice of appropriate sample pretreatment methods. The future direction of the development of sample pretreatment technology for pesticide residue detection should be as fast, accurate, environmentally friendly and highly automated, in order to avoid the loss of sample transfer as much as possible, and reduce the accidental errors of various human factors.