CN110987834A - Method for detecting antibacterial performance of ceramic - Google Patents
Method for detecting antibacterial performance of ceramic Download PDFInfo
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- CN110987834A CN110987834A CN201911174317.6A CN201911174317A CN110987834A CN 110987834 A CN110987834 A CN 110987834A CN 201911174317 A CN201911174317 A CN 201911174317A CN 110987834 A CN110987834 A CN 110987834A
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- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
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Abstract
The invention discloses a method for detecting antibacterial performance of ceramics, which comprises the steps of respectively carrying out spectrophotometric detection on mixed liquor in which ceramics to be detected are soaked, mixed liquor in which the ceramics to be detected are not soaked and frozen, obtaining absorbance data, and then calculating according to a formula to obtain the antibacterial rate of the ceramics to be detected, thereby realizing the rapid detection of the antibacterial performance of the ceramics. The detection method provided by the invention is simple and easy to implement, simple and convenient to operate, rapid, high in accuracy and good in repeatability, and meanwhile, the antibacterial performance of the ceramic can be visually observed through the color change of the bacterial liquid.
Description
Technical Field
The invention relates to the technical field of detection of antibacterial performance of materials, in particular to a method for detecting antibacterial performance of ceramics.
Background
With the frequent occurrence of food-borne food poisoning caused by pathogenic bacteria, people have a deep concept of disinfection and antibiosis and pay more and more attention to antibacterial products. As a ceramic product material which is most widely used for home decoration, the antibacterial performance of the ceramic product material is attracted by market attention, and a wide variety of antibacterial ceramic products are brought to the market. In the face of continuously developing new antibacterial ceramic products, the inspection department needs to objectively evaluate the antibacterial ceramic products, so that consumers can use the antibacterial ceramic products safely. According to the traditional antibacterial ceramic detection methods, JIS Z2801 and ISO 22196, diluted bacterial suspension is contacted with a sample and cultured for a period of time, and then the number of bacteria before and after culture is detected to calculate the antibacterial rate of the sample, so that the traditional antibacterial ceramic detection methods have the characteristics of long detection period (at least two days), complex operation, insignificant comparison of antibacterial effects of products and the like. The lag behind of the detection method for the antibacterial activity of the ceramic greatly influences the development of the antibacterial ceramic, and the establishment of a method for rapidly detecting the antibacterial performance of the ceramic is particularly necessary.
Disclosure of Invention
The invention aims to provide a method for rapidly detecting antibacterial performance of ceramic. Therefore, the invention provides a method for detecting antibacterial performance of ceramic, which comprises the following steps:
s1: soaking the ceramic to be tested in the mixed solution for culturing for 3-6 h to serve as an experimental group; culturing the mixed solution with the same volume for the same time to obtain a blank group; taking the mixed solution with the same volume, preserving at the temperature of 0.5-3 ℃, and taking the mixed solution as a control group, wherein the preservation time is the same as the culture time; the mixed solution comprises a prefabricated bacterial suspension, a lactose cholate culture solution and an acid-base indicator;
s2: respectively taking the mixed solution of the cultured experimental group and the mixed solution of the cultured blank group with the same volume as the test solution X and the test solution Y, and simultaneously taking the mixed solution of the preserved control group with the same volume as the test solution Z; respectively centrifuging the test solution X, the test solution Y and the test solution Z to respectively obtain supernatant X1Supernatant Y1And supernatant Z1Separately test the supernatant X1Supernatant Y1And supernatant Z1The antibacterial rate of the ceramic to be detected is obtained according to a formula by the absorbance at the maximum absorption wavelength of the acid-base indicator, so that the antibacterial performance of the ceramic to be detected is detected;
wherein the preformed bacterial suspension is prepared by adding a slant culture of a nutrient agar culture medium of escherichia coli into water and uniformly mixing; the formula is R (%) -100 (A-B)/(A-C), R is the antibacterial rate of the ceramic to be tested, and A is supernatant Y1Absorbance at the maximum absorption wavelength of the acid-base indicator, B is the supernatant X1In the acid-base meaningAbsorbance at the maximum absorption wavelength of the indicator, C is the supernatant Z1Absorbance at the absorption maximum wavelength of the acid-base indicator.
Wherein, the pottery that awaits measuring can be applicable to the ceramic tile at least, and general ceramic tile is the cuboid that has thinner thickness, and for the antibacterial property of guaranteeing the test ceramic tile upper surface, the other five faces of the ceramic tile that will await measuring except that the upper surface are wrapped up with waterproof adhesive tape, and the antibacterial property of other faces has no actual meaning.
Bacteria are identified by the action of various enzymes produced by bacterial cells, which decompose sugars depending on the kind of bacteria. When the coliform group bacteria exist in the presence of bile salt or other equivalent selection factors, lactose can be decomposed and acid can be generated by growth at 37 ℃, the pH value of a liquid culture medium is reduced, the color of an indicator in the culture solution is changed, and the culture solution keeps original color because other bacteria do not decompose lactose and do not generate acid. Generally, the acid production quantity is in positive correlation with the detection rate of coliform groups, and the color of the culture solution gradually changes into yellow along with the increase of the concentration of escherichia coli; the higher the concentration of Escherichia coli, the more pronounced the yellow color, and the stronger the absorbance of the culture solution to blue at a wavelength of 465 nm. Bromcresol purple, chlorophenol red and bromophenol red are used as indicators sensitive to acid-base development, the pH color change range is 5.0 (yellow) -7.0, and the color-changed yellow culture solution has the maximum absorbance at the wavelength of 465nm, so that the bacterial activity can be quantitatively detected by measuring the absorbance of the recovered bacterial solution, and the quantitative evaluation on the antibacterial activity of the material is realized.
Preferably, in S1, before the ceramic to be tested is immersed in the mixed solution, sterilization and drying are further performed, where the sterilization and drying specifically include: and soaking the ceramic to be tested in an ethanol solution for 5min, then taking out, and drying at 60 ℃.
Preferably, the temperature during the culture in S1 is 35-37 ℃.
Preferably, the nutrient agar culture medium slant culture of the escherichia coli is prepared by inoculating the escherichia coli on the nutrient agar culture medium slant and culturing for 18-24 hours.
Preferably, the acid-base indicator is a bromocresol purple solution, a bromophenol red solution, or a green phenol red solution.
Preferably, the lactose bile salt culture solution is prepared by dissolving a lactose bile salt fermentation culture medium in water and then sterilizing; the bromocresol purple solution, the bromophenol red solution and the green phenol red solution are respectively prepared by dissolving bromocresol purple, bromophenol red and green phenol red in a sodium hydroxide solution.
Preferably, the lactose bile salt culture solution is prepared by dissolving 35g of lactose bile salt fermentation culture medium in 1000mL of water and sterilizing at 121 ℃ for 15 min; the bromocresol purple solution is prepared by dissolving 0.1g of bromocresol purple in 20mL of 0.02mol/L sodium hydroxide solution and then adding water to dilute to 100 mL; the bromophenol red solution is prepared by dissolving 0.1g of bromophenol red in 20mL of 0.02mol/L sodium hydroxide solution and then adding water to dilute to 100 mL; the green phenol red solution is prepared by dissolving 0.1g of green phenol red in 20mL of 0.02mol/L sodium hydroxide solution, and then adding water to dilute to 100 mL.
Preferably, the concentration of the pre-formed bacterial suspension is 105～106CFU/mL。
Preferably, the mixed solution comprises 1mL of prefabricated bacterial suspension, 50mL of lactose cholate culture solution and 2mL of bromcresol purple solution, and the bromcresol purple solution can be replaced by bromphenol red solution or green phenol red solution.
Has the advantages that: the detection method provided by the invention is simple and easy to implement, simple and convenient to operate, rapid, high in accuracy and good in repeatability, and meanwhile, the antibacterial performance of the ceramic can be visually observed through the color change of the bacterial liquid.
Detailed Description
The concept and technical effects of the present invention will be clearly and completely described in the following embodiments to fully understand the objects, aspects and effects of the present invention. It should be noted that the embodiments and features of the embodiments in the present application may be combined with each other without conflict.
Example 1:
preparation work:
(1) preparation of test bacterial suspensions: taking out the slant of the nutrient agar culture medium of the escherichia coli by using sterile inoculating loop for 24 hours, adding 1 mu L of fresh culture into sterile water for dilution, shaking and uniformly mixing, and placing in a refrigerator at 4 ℃ for storage for later use.
(2) Preparing a sample to be tested: taking 1 tile with activity value of 0.1 detected by JIS Z2801, cutting into 50 x 50mm, winding with waterproof adhesive tape to remove other 5 surfaces of the glaze, soaking in ethanol solution for 5min for sterilization, taking out the sample, and drying in an oven at 60 deg.C.
(3) Preparing a culture solution: dissolving 35g lactose bile salt fermentation culture medium in 1000ml distilled water, sterilizing at 121 deg.C for 15min, and cooling.
(4) Preparing an indicator: 0.1g of bromocresol purple is taken and added with 20ml of sodium hydroxide solution with the concentration of 0.02mol/L for dissolution, and then the solution is diluted by sterile water to be constant volume in a 100ml volumetric flask to prepare 1g/L of bromocresol purple solution as an indicator for standby.
S1: adding the sterilized and dried tile sample and the mixed solution (comprising 1mL of prefabricated bacterial suspension, 50mL of lactose cholate culture solution and 2mL of bromocresol purple solution) into a sterile experiment bag a1, completely soaking the sample in the mixed solution, and performing shake culture on a shaking table at 36 +/-1 ℃ for 6 hours to obtain an experiment group; adding equal amount of mixed solution into the empty sterilization bag a2, and shake-culturing for 6h on a shaking table at 36 +/-1 ℃ to serve as a blank group; adding equal amount of the mixture into the empty sterilization bag a3, storing in a refrigerator at 1 deg.C for 6 hr to prevent proliferation of Escherichia coli, and using as control group;
s2: culturing the experimental group and the blank group for 6h and storing the control group for 6h, respectively adding 10mL of mixed solution into three centrifuge tubes in sterile bags of the experimental group, the blank group and the control group for centrifugation to respectively obtain supernatant X1Supernatant Y1And supernatant Z1(ii) a Test X1、Y1、Z1Absorbance at 465nm to obtain supernatant X of the experimental group1Absorbance at 465nm of 0.81, supernatant Y of blank1Absorbance at 465nm of 0.79; supernatant Z of control group1Absorbance at 465nm of 0.21; calculating according to the formula R (%) 100 × (A-B)/(A-C) of the antibacterial rate of the ceramic tile (R is antibacterial rate, A is absorbance of the blank group supernatant at 465nm, and B is absorbance of the experiment group supernatant at 465nmAbsorbance, C is the absorbance at 465nm of the control supernatant); r (%) ═ 100 ═ (0.81-0.79)/(0.81-0.21) ═ 1.3%; the antibacterial rate of the tile with the activity value of 0.1 is 1.3%.
Meanwhile, the color of the mixed liquor in the blank group and the mixed liquor soaked with the ceramic tiles with the activity value of 0.1 are changed from purple to yellow.
Example 2:
preparation work:
(1) preparation of test bacterial suspensions: taking out the slant of the nutrient agar culture medium of the escherichia coli by using sterile inoculating loop for 24 hours, adding 1 mu L of fresh culture into sterile water for dilution, shaking and uniformly mixing, and placing in a refrigerator at 4 ℃ for storage for later use.
(2) Preparing a sample to be tested: taking 1 tile with activity value of 2.7 detected by JIS Z2801, cutting into 50 x 50mm, winding with waterproof adhesive tape to remove other 5 surfaces of the glaze, soaking in ethanol solution for 5min for sterilization, taking out the sample, and drying in an oven at 60 deg.C.
(3) Preparing a culture solution: dissolving 35g lactose bile salt fermentation culture medium in 1000ml distilled water, sterilizing at 121 deg.C for 15min, and cooling.
(4) Preparing an indicator: 0.1g of bromocresol purple is taken and added with 20ml of sodium hydroxide solution with the concentration of 0.02mol/L for dissolution, and then the solution is diluted by sterile water to be constant volume in a volumetric flask of 100ml, and prepared into 1g/L of bromophenol red solution as an indicator for standby.
S1: adding the sterilized and dried tile sample and the mixed solution (comprising 1mL of prefabricated bacterial suspension, 50mL of lactose cholate culture solution and 2mL of bromocresol purple solution) into a sterile experiment bag a1, completely soaking the sample in the mixed solution, and performing shake culture on a shaking table at 36 +/-1 ℃ for 6 hours to obtain an experiment group; adding equal amount of mixed solution into the empty sterilization bag a2, and shake-culturing for 6h on a shaking table at 36 +/-1 ℃ to serve as a blank group; adding equal amount of the mixture into the empty sterilization bag a3, storing in a refrigerator at 1 deg.C for 6 hr to prevent proliferation of Escherichia coli, and using as control group;
s2: after the experimental group and the blank group are cultured for 6h and the control group is stored for 6h, the experimental group, the blank group and the control group are respectively absentRespectively adding 10mL of mixed solution into three centrifuge tubes for centrifugation to obtain supernatant X1Supernatant Y1And supernatant Z1(ii) a Test X1、Y1、Z1Absorbance at 465nm to obtain supernatant X of the experimental group1Absorbance at 465nm of 0.81, supernatant Y of blank1Absorbance at 465nm of 0.22; supernatant Z of control group1Absorbance at 465nm of 0.21; calculating according to a calculation formula R (%) of the antibacterial rate of the ceramic tile, wherein R is the antibacterial rate, A is the absorbance of a blank group supernatant at 465nm, B is the absorbance of an experimental group supernatant at 465nm, and C is the absorbance of a control group supernatant at 465 nm; r (%). 100 ═ (0.81-0.22)/(0.81-0.21): 98.3%; the antibacterial rate of the tile with the activity value of 2.7 is 98.3%.
Meanwhile, the color of the mixed solution in the blank group changed from red to yellow, while the color of the mixed solution soaked with the tile having an activity value of 2.7 hardly changed.
Claims (9)
1. A method for detecting antibacterial performance of ceramic is characterized by comprising the following steps:
s1: soaking the ceramic to be tested in the mixed solution for culturing for 3-6 h to serve as an experimental group; culturing the mixed solution with the same volume for the same time to obtain a blank group; taking the mixed solution with the same volume, and storing at the temperature of 0.5-3 ℃, wherein the storage time is the same as the culture time, and taking the mixed solution as a control group; the mixed solution comprises a prefabricated bacterial suspension, a lactose cholate culture solution and an acid-base indicator;
s2: respectively taking the mixed solution of the cultured experimental group and the mixed solution of the cultured blank group with the same volume as the test solution X and the test solution Y, and simultaneously taking the mixed solution of the preserved control group with the same volume as the test solution Z; respectively centrifuging the test solution X, the test solution Y and the test solution Z to respectively obtain supernatant X1Supernatant Y1And supernatant Z1Separately test the supernatant X1Supernatant Y1And supernatant Z1Absorbance at the absorption maximum wavelength of the acid-base indicator according toThe antibacterial rate of the ceramic to be detected is obtained according to a formula, so that the antibacterial performance of the ceramic to be detected is detected;
wherein the preformed bacterial suspension is prepared by adding a slant culture of a nutrient agar culture medium of escherichia coli into water and uniformly mixing; the formula is R (%) -100 (A-B)/(A-C), R is the antibacterial rate of the ceramic to be tested, and A is supernatant Y1Absorbance at the maximum absorption wavelength of the acid-base indicator, B is the supernatant X1Absorbance at the maximum absorption wavelength of the acid-base indicator, C is the supernatant Z1Absorbance at the absorption maximum wavelength of the acid-base indicator.
2. The method according to claim 1, wherein in S1, before the ceramic to be tested is immersed in the mixed solution, the ceramic to be tested is further sterilized and dried, and the specific process of sterilizing and drying is as follows: and soaking the ceramic to be tested in an ethanol solution for 5-30 min, taking out, and drying at 60 ℃.
3. The method according to claim 1, wherein the temperature during the culturing in S1 is 35 to 37 ℃.
4. The method according to claim 1, wherein the nutrient agar culture medium slant culture of Escherichia coli is prepared by inoculating Escherichia coli to the nutrient agar culture medium slant and culturing for 18-24 h.
5. The method of claim 1, wherein the acid-base indicator is a bromocresol purple solution, a bromophenol red solution, or a green phenol red solution.
6. The method as claimed in claim 5, wherein the lactose cholate culture solution is prepared by dissolving lactose cholate fermentation medium in water and sterilizing; the bromocresol purple solution is prepared by dissolving bromocresol purple in sodium hydroxide solution.
7. The method as claimed in claim 6, wherein the lactose cholate culture solution is prepared by dissolving 35g of lactose cholate fermentation medium in 1000mL of water and sterilizing at 121 ℃ for 15 min; the bromocresol purple solution is prepared by dissolving 0.1g of bromocresol purple in 20mL of 0.02mol/L sodium hydroxide solution and then adding water to dilute to 100 mL.
8. The method of claim 7, wherein the concentration of the pre-formed bacterial suspension is 105～106CFU/mL。
9. The method of claim 8, wherein the mixture comprises 1mL of the preformed bacterial suspension, 50mL of the lactose bile salt broth, and 2mL of the bromocresol purple solution.
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