CN111537501A - Kit and method for rapidly detecting total amount of ethylene oxide and 2-chloroethanol in medical instrument - Google Patents
Kit and method for rapidly detecting total amount of ethylene oxide and 2-chloroethanol in medical instrument Download PDFInfo
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Abstract
The invention provides a kit and a method for rapidly detecting the total amount of ethylene oxide and 2-chloroethanol in medical equipment. The color matching material comprises a sodium hydroxide solution, a potassium iodide solution, a mixed solution of periodic acid and hydrochloric acid, a sodium thiosulfate solution, an ethylene diamine tetraacetic acid solution, a 4-amino-3-hydrazine-5-mercapto-1, 2, 4-triazocene solution, a potassium periodate solution and a standard color matching card. The method is simple and convenient to operate, low in cost, high in speed and high in sensitivity, can be used for rapidly and simultaneously detecting the residual quantity of ethylene oxide and 2-chloroethanol in a medical instrument on site, can obtain an experimental result within 40 minutes without using special instruments and equipment, and is suitable for rapid detection on site.
Description
Technical Field
The present invention relates to the field of chemical detection technology, in particular to the field of testing or analyzing materials by means of determining chemical or physical properties of the materials, and more particularly to the rapid detection of the total amount of ethylene oxide and 2-chloroethanol residues.
Background
Ethylene Oxide (EO) is a broad-spectrum low-temperature sterilizing agent, and the sterilization mechanism of the agent is that ethylene oxide reacts with carboxylic acid, amino group, sulfenyl group and hydroxyl group on the protein of the microorganism to generate alkylation reaction, so that the normal function of the microorganism is prevented, and the microorganism is killed. Can kill various microorganisms at normal temperature, and is widely applied to the disinfection of medical instruments. EO penetrability is very strong, can pierce through the micropore and reach the inside corresponding degree of depth of product to improve the sterilization effect greatly. However, when microorganisms are killed, the residual ethylene oxide may cause some damage to the user's body, may cause erosion to the eyes and respiratory tract, may cause vomiting, nausea, diarrhea, headache, central depression, dyspnea, pulmonary edema, etc., and may also cause liver and kidney damage and hemolysis.
2-chloroethanol (ECH) is an organic substance derived from ethylene oxide sterilization processes. It can stimulate the body surface, has acute toxicity, can be rapidly absorbed by the skin, and can cause damage to the lung, kidney, central nervous system and cardiovascular system.
At present, GB/T16886.7-2001 sets forth the permissible limits for ethylene oxide sterilization residues (including ethylene oxide and 2-chloroethanol) on ethylene oxide sterilized medical device unit products. The method for simultaneously detecting the ethylene oxide and the chloroethanol adopts a gas chromatography-mass spectrometry combined method, the two methods are general authoritative methods of domestic professional detection institutions at present, and the detection result is accurate. However, the method has strong equipment dependence, needs large-scale instruments such as a gas chromatograph and the like and professional experiment operators, has high requirements on the experiment environment, and finally causes complex detection procedures, long time consumption and high detection cost.
In a complete medical instrument detection system, a technical method for carrying out corroborative detection based on a large-scale precision instrument is required, and a determination method capable of rapidly meeting the requirements of on-site detection and rapid screening of new outbreak epidemic and major public health events is required.
Through retrieval, the prior art which can simultaneously realize the rapid detection of the ethylene oxide and the 2-chloroethanol in a simpler, faster and more economical way is not found, and only the following related technical schemes exist:
chinese patent application 201710227947.X discloses a method for measuring the residual amount of ethylene oxide in a daily protective mask and an evaluation method thereof, dilute hydrochloric acid is used as an extracting agent, periodic acid is used for oxidizing an extraction liquid in sequence, acetylacetone is used for developing color, and finally an ultraviolet spectrophotometer is used for measuring the absorbance value of a developing solution. Compared with the prior art, the method has the advantages of simple pretreatment, stable test result, low detection limit, high accuracy and the like.
However, firstly, the above-mentioned techniques do not achieve simultaneous detection of ethylene oxide and 2-chloroethanol; secondly, the technology adopts acetylacetone for color development, and the technology adopts 4-amino-3-hydrazine-5-sulfydryl-1, 2, 4-triazene metallocene (AHMT) for color development; furthermore, according to the specification, the detection is carried out according to the method through technical reproduction, and the shortest time is about 140 min. This is clearly not satisfactory for solving the technical problem of rapid detection.
The method has come forward under the technical background, and aims to solve the problem that the total amount of 1, ethylene oxide and 2-chloroethanol is detected simultaneously; 2. really realizing the technical problem including quick detection.
Disclosure of Invention
The invention aims to develop a rapid detection kit for the total amount of ethylene oxide and 2-chloroethanol in medical instruments and a detection method thereof.
The present invention utilizes the nucleophilic substitution reaction of ethylene oxide and 2-chloroethanol in sample under the action of iodide ion to produce 2-chloroethanol so as to produce ethylene glycol, the ring-opening reaction of ethylene oxide can produce ethylene glycol, and the ethylene glycol can be oxidized into formaldehyde by means of periodic acid, and the excess periodic acid can be removed by means of sodium thiosulfate titration, and the formaldehyde can be reacted with colour-developing agent to produce complex, and its colour shade can be positively correlated with formaldehyde content in a certain range, and the visual colorimetry can be implemented by means of colour-gradation solution or colour-gradation card so as to implement semi-quantitative determination of total quantity of ethylene oxide and 2-chloroethanol in the sample. The experiment quantification mainly comprises that 2moL of formaldehyde can be obtained through the reaction of 1moL of ethylene oxide or 2-chloroethanol, so that the total amount (in molar concentration) of the ethylene oxide and the 2-chloroethanol in a sample can be calculated according to the molar amount of the formaldehyde in a test solution.
In order to complete the experimental purpose, the invention provides a rapid detection kit for ethylene oxide and 2-chloroethanol residues in medical instruments and a detection method thereof.
On the one hand, the detection kit comprises a reagent A, a reagent B, a reagent C, a reagent D, a reagent E, a reagent F, a reagent G, a colorimetric tube, a measuring cylinder and a standard colorimetric card.
Further, the reagent A is sodium hydroxide solution, and the concentration is 0.1mol/L-0.5 mol/L.
Further, the reagent B is a potassium iodide solution, and the concentration is 0.05mol/L-0.2 mol/L.
Further, the reagent C is a mixed solution of periodic acid and hydrochloric acid, the concentration of the periodic acid is 10g/L-30g/L, and the concentration of the hydrochloric acid is 4mol/L-6 mol/L.
Further, the reagent D is a sodium thiosulfate solution, and the concentration is 40g/L-80 g/L.
Further, the reagent E is an ethylene diamine tetraacetic acid disodium solution, and the concentration is 100g/L-200 g/L.
Further, the reagent F is a 4-amino-3-hydrazine-5-sulfydryl-1, 2, 4-triazene solution, and the concentration is 2g/L-8 g/L.
Further, the reagent G is potassium periodate solution, and the concentration is 10G/L-20G/L.
On the other hand, the detection method is implemented according to the following steps:
step 1, preparation of sample liquid:
adding a reagent A and a reagent B into a sample to be detected, and taking supernatant after water bath; adding the reagent C into the supernatant, shaking up, carrying out water bath at 60 ℃, and then dropwise adding the reagent D until the yellow color just disappears; then adding the reagent E and the reagent F, mixing uniformly, standing, adding the reagent G, mixing uniformly, and standing to obtain a sample solution;
step 2, colorimetric determination:
carrying out colorimetric determination on the sample solution and a standard color scale solution or a standard colorimetric card, determining the total content of ethylene oxide and 2-chloroethanol in the sample to be detected, and judging the result within 10 min;
the reagent A is sodium hydroxide solution; the reagent B is a potassium iodide solution; the reagent C is a mixed solution of periodic acid and hydrochloric acid; the reagent D is a sodium thiosulfate solution; the reagent E is an ethylene diamine tetraacetic acid disodium solution; the reagent F is a 4-amino-3-hydrazine-5-sulfydryl-1, 2, 4-triazocene solution; the reagent G is potassium periodate solution.
As some exemplary schemes, the detection method is specifically implemented according to the following steps:
step 1, preparing a sample solution;
accurately weighing 1.0G +/-0.01G, placing the weighed mass in a 50mL centrifuge tube, adding a proper amount of reagent A, adding a proper amount of reagent B, placing the weighed mass in a water bath for 20min, taking 2mL of supernatant in a 15mL centrifuge tube, adding reagent C, covering the centrifuge tube with a cover, shaking up, placing the centrifuge tube in a water bath at 60 ℃ for reaction for 5min, dropwise adding reagent D until yellow appears and disappears, adding reagent E and reagent F, performing vortex mixing, standing for 10min, adding reagent G, performing vortex mixing, and standing for 5 min.
Step 2, colorimetric determination
And after the reaction is finished, carrying out colorimetric determination on the sample solution, determining the total content of ethylene oxide and 2-chloroethanol in the sample, and judging the result within 10 min.
Further, in the step 1, the amount of the added reagent A is 10mL-30mL, the amount of the added reagent B is 1-3 drops, the amount of the added reagent C is 1-3 drops, the amount of the added reagent E is 8-15 drops, the amount of the added reagent F is 8-15 drops, and the amount of the added reagent G is 8-15 drops.
Further, the colorimetric determination method in step 2 is to compare the color with a standard color gradation solution or a color chart by visual colorimetry, judge the total content of ethylene oxide and 2-chloroethanol in the sample by comparing with the standard color gradation solution or the standard color gradation chart, and determine that the sample is positive if the color is deeper than the detection limit, otherwise, determine that the sample is negative.
Further, the standard color gradation solution or the standard color chart is prepared according to the following steps:
(1) preparing a 2-chloroethanol standard working solution with a known concentration by using a reagent A as a solvent;
(2) adding a reagent B into the standard working solution, taking supernatant after water bath, adding a reagent C into the supernatant, shaking up, carrying out water bath at 60 ℃, dropwise adding a reagent D until yellow appears and disappears, adding a reagent E and a reagent F, mixing uniformly, standing, adding a reagent G, mixing uniformly, and standing to obtain the standard color gradation solution;
(3) and printing according to the color value of each standard color gradation solution to obtain the standard color comparison card.
As some illustrative examples, the standard color-scale solution or the standard color chart is prepared by the following steps:
(1) preparing 2-chloroethanol standard working solution with the concentration of 0 mu g/mL, 0.005 mu mol/mL, 0.010 mu mol/mL, 0.020 mu mol/mL, 0.040 mu mol/mL, 0.100 mu mol/mL and 0.200 mu mol/mL by using the reagent A;
(2) respectively putting a proper amount of the standard solution into a 50mL centrifuge tube, adding a proper amount of reagent B, putting the centrifuge tube into a water bath for 20min, putting 2mL of supernate into a 15mL centrifuge tube, adding reagent C, covering a cover, shaking up, putting the centrifuge tube into a water bath at 60 ℃ for reacting for 5min, dropwise adding reagent D until yellow appears and disappears, adding reagent E and reagent F, carrying out vortex mixing, standing for 10min, adding reagent G, carrying out vortex mixing, and standing for 5min to obtain a standard color gradation solution.
(3) And printing according to the color value of each standard color gradation solution to obtain a standard color comparison card.
Compared with the prior art, the invention has the beneficial effects that:
(1) through repeated determination, the detection kit and the detection method provided by the invention take less time than 40min, and are obviously shorter than the prior art.
(2) Obviously, the method realizes the simultaneous one-step measurement of the total amount of the ethylene oxide and the 2-chloroethanol in the sample to be measured.
(3) Obviously, the detection kit and the method of the invention are free from the limit of expensive instruments, and have simple and convenient operation and low cost.
In conclusion, the method is simple and convenient to operate, low in cost, high in speed and high in sensitivity, can be used for rapidly detecting ethylene oxide and 2-chloroethanol residues in medical instruments on site and detecting the ethylene oxide and the 2-chloroethanol residues simultaneously, does not need special instruments and equipment, can obtain an experimental result within 40 minutes, and is suitable for rapid detection on site.
Drawings
FIG. 1 is a standard color chart obtained in example 1.
Detailed Description
Unless otherwise specified, the instruments and reagents used in the practice of the present application are commercially available. The technical scheme of the application can be normally purchased and reproduced commercially by a person skilled in the art. The present invention will be described in detail below with reference to specific embodiments, but the present invention is not limited thereto.
Example 1 reagent preparation and Standard Gray Scale solution and Standard colorimetric card preparation
1. Reagent preparation
Respectively preparing a sodium hydroxide solution (reagent A) with the concentration of 0.1mol/L, a potassium iodide solution (reagent B) with the concentration of 0.2mol/L, a mixed solution of periodic acid and hydrochloric acid (reagent C) with the concentrations of 10G/L and 6mol/L, a sodium thiosulfate solution (reagent D) with the concentration of 40G/L, an ethylene diamine tetraacetic acid disodium solution (reagent E) with the concentration of 100G/L, a 4-amino-3-hydrazine-5-mercapto-1, 2, 4-triazocene solution (reagent F) with the concentration of 5G/L and a potassium periodate solution (reagent G) with the concentration of 15G/L.
2. Standard color gradation solution and standard colorimetric card manufacturing method
Preparing 2-chloroethanol standard working solution with the concentration of 0 mu mol/mL, 0.005 mu mol/mL, 0.010 mu mol/mL, 0.020 mu mol/mL, 0.040 mu mol/mL, 0.100 mu mol/mL and 0.200 mu mol/mL by using a reagent A as a constant volume solution; corresponding to the total amount of ethylene oxide and 2-chloroethanol in the sample: 0mmol/kg, 0.1mmol/kg, 0.2mmol/kg, 0.4mmol/kg, 0.8mmol/kg, 2.0mmol/kg, 4.0 mmol/kg.
Respectively taking 20mL of standard working solution, placing the standard working solution into a 50mL centrifuge tube, adding 3 drops (about 150 mu L) of reagent B, placing the solution into a water bath for 20min, taking 2mL of supernatant into a 15mL centrifuge tube, adding 3 drops (about 150 mu L) of reagent C, covering the centrifuge tube with a cover, shaking up, placing the centrifuge tube into a water bath at 60 ℃ for reaction for 5min, dropwise adding reagent D until yellow appears and disappears, adding 8 drops (about 500 mu L) of reagent E and 8 drops (about 500 mu L) of reagent F, standing the mixture for 10min after vortex mixing, adding 3 drops (about 150 mu L) of reagent G, and standing the mixture for 5min after vortex mixing to obtain a standard color gradation solution.
Printing a standard color comparison card: and printing a standard color comparison card according to the color value of each standard color level solution, and performing plastic package storage on the outer surface of the standard color comparison card, wherein the finally manufactured color level card is shown in fig. 1.
Example 2: example of Rapid measurement of the Total amount of ethylene oxide and 2-chloroethanol
The reagent used in this example was prepared as in example 1.
1. Detection method
Step 1, preparing a sample solution;
accurately weighing 1.0G +/-0.01G of sample to be detected, placing the sample to be detected in a 50mL centrifuge tube, adding 20mL of reagent A, adding 3 drops (about 150 mu L) of reagent B, placing the sample in a 60 ℃ water bath for 20min, taking 2mL of supernate into a 15mL centrifuge tube, adding 3 drops (about 150 mu L) of reagent C, covering a cover, shaking uniformly, placing the sample in a 60 ℃ water bath for reaction for 5min, dropping the reagent D until yellow appears and disappearing right, adding 8 drops (about 150 mu L) of reagent E and 8 drops (about 150 mu L), standing for 10min after vortex mixing, adding 3 drops (about 150 mu L) of reagent G, and standing for 5min after vortex mixing.
Step 2, colorimetric determination
After the reaction is finished, the sample solution is subjected to colorimetric determination by using the standard colorimetric card prepared in example 1, the total content of ethylene oxide and 2-chloroethanol in the sample is determined, and the result is interpreted within 10 min. A positive sample is obtained if the color is darker than the detection limit, and a negative sample is obtained if the color is not darker than the detection limit.
2. Examples of detection
20 parts of medical instruments extracted from different manufacturers are detected, wherein each part comprises two identical parallel samples, one sample is detected according to the detection method (short for quick detection method), and the other sample is detected by adopting a laboratory detection method, and the laboratory method is selected from GB/T16886.7-2015 part 7 of biological evaluation of medical instruments: residual ethylene oxide sterilization (laboratory methods for short).
The detection results are shown in table 1, and it can be seen from table 1 that the detection results of the rapid detection method are consistent with those of the laboratory method, and the method can obtain the experimental results within 40 minutes without using special instruments and equipment, and is suitable for field rapid detection.
TABLE 1 quick test method and laboratory comparison results
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Claims (10)
1. A kit for rapidly detecting the total amount of ethylene oxide and 2-chloroethanol in a medical instrument is characterized in that:
the kit comprises a reagent A, a reagent B, a reagent C, a reagent D, a reagent E, a reagent F, a reagent G and a standard colorimetric card;
the reagent A is sodium hydroxide solution; the reagent B is a potassium iodide solution; the reagent C is a mixed solution of periodic acid and hydrochloric acid; the reagent D is a sodium thiosulfate solution; the reagent E is an ethylene diamine tetraacetic acid disodium solution; the reagent F is a 4-amino-3-hydrazine-5-sulfydryl-1, 2, 4-triazocene (AHMT) solution; the reagent G is potassium periodate solution.
2. The kit of claim 1, wherein: the concentration of the sodium hydroxide solution is 0.1-0.5 mol/L.
3. The kit of claim 1, wherein: the concentration of the potassium iodide solution is 0.05mol/L-0.2 mol/L.
4. The kit of claim 1, wherein: the concentration of periodic acid in the mixed solution of periodic acid and hydrochloric acid is 10g/L-30g/L, and the concentration of hydrochloric acid is 4mol/L-6 mol/L.
5. The kit of claim 1, wherein: the concentration of the sodium thiosulfate solution is 40g/L-80 g/L.
6. The kit of claim 1, wherein: the concentration of the ethylene diamine tetraacetic acid disodium solution is 100g/L-200 g/L.
7. The kit of claim 1, wherein: the concentration of the 4-amino-3-hydrazine-5-mercapto-1, 2, 4-triazene solution is 2g/L-8 g/L.
8. The kit of claim 1, wherein: the concentration of the potassium periodate solution is 10g/L-20 g/L.
9. A method for rapidly detecting the total amount of ethylene oxide and 2-chloroethanol in a medical device, the method comprising the steps of:
step 1, preparation of sample liquid:
adding a reagent A and a reagent B into the sample to be measured, and taking supernatant after water bath; adding the reagent C into the supernatant, shaking up, carrying out water bath at 60 ℃, and then dropwise adding the reagent D until the yellow color just disappears; then adding the reagent E and the reagent F, mixing uniformly, standing, adding the reagent G, mixing uniformly, and standing to obtain a sample solution;
step 2, colorimetric determination:
carrying out colorimetric determination on the sample solution and a standard color scale solution or a standard colorimetric card, determining the total content of ethylene oxide and 2-chloroethanol in the sample to be detected, and judging the result within 10 min;
the reagent A is sodium hydroxide solution; the reagent B is a potassium iodide solution; the reagent C is a mixed solution of periodic acid and hydrochloric acid; the reagent D is a sodium thiosulfate solution; the reagent E is an ethylene diamine tetraacetic acid disodium solution; the reagent F is a 4-amino-3-hydrazine-5-sulfydryl-1, 2, 4-triazocene solution; the reagent G is potassium periodate solution.
10. The method of claim 9, wherein: the standard color gradation solution or the standard color comparison card is prepared according to the following steps:
(1) preparing a 2-chloroethanol or ethylene oxide standard working solution with a known concentration by using the reagent A as a solvent;
(2) adding a reagent B into the standard working solution, taking supernatant after water bath, adding a reagent C into the supernatant, shaking up, carrying out water bath at 60 ℃, dropwise adding a reagent D until yellow appears and disappears, adding a reagent E and a reagent F, mixing uniformly, standing, adding a reagent G, mixing uniformly, and standing to obtain the standard color gradation solution;
(3) and printing according to the color value of each standard color gradation solution to obtain the standard color comparison card.
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