Performance test methods for textile antibacterial fabrics

As antibacterial finishing of protective functional textiles, there are various performance testing methods. Various qualitative and quantitative testing methods for the antibacterial properties of textiles are introduced in detail; at the same time, the key factors affecting the testing results of the antibacterial properties of textiles are discussed.

1. Selection of test strains

Species tested included bacteria and fungi. Among bacteria, Gram-positive bacteria and Gram-negative bacteria are mainly used, and among fungi, molds and ringworms are mainly used.

In the evaluation of the antibacterial performance of textile antibacterial fabrics, the selection of bacterial strains must be scientific and representative. Staphylococcus aureus is a highly resistant pathogenic bacterium among asporal bacteria and can be used as a representative of Gram-positive bacteria. Bacillus megaterium is a common pathogenic bacterium among spore-like bacteria; Bacillus subtilis is easy to form spores and has strong resistance, so it can be used as a representative of spore bacteria. Escherichia coli is widely distributed and has been used in various experiments as a representative strain of common Gram-negative bacteria. Aspergillus flavus and Chaetomium globus have been included in my country’s national standards (GB2423.16-81) as prescribed strains for anti-mold tests. Some other selected molds are common molds that corrode textiles or polymer materials. Candida albicans is a common opportunistic pathogenic fungus in human skin and mucous membranes. It is sensitive to drugs and has the characteristics of a fungus. The colonies resemble bacteria but not bacteria and are different from molds. Because they have colonies that resemble bacteria, they are easy to count and observe. They are often used as Representative of fungi.

Therefore, in order to assess whether textile antibacterial fabrics have broad-spectrum antibacterial effects, a more reasonable choice is to mix representative bacterial strains in a certain proportion for testing. For the antibacterial properties of most current antibacterial products, only Staphylococcus aureus, Escherichia coli and Candida albicans are often selected as representatives of Gram-positive bacteria, Gram-negative bacteria and fungi respectively. But in fact, it is far from enough to only use these three bacteria to represent the antibacterial properties of fabrics.

In addition, since most fungi cannot count the number of colonies, the antifungal performance of textiles is mainly evaluated by observing the growth of fungi on the sample after a certain period of time under certain temperature and humidity conditions after the sample is exposed to the fungus. Assessment, and for the assessment of the degree of fungal growth, the British standard BS6085-81 is used for grade assessment.

2. Textile antibacterial performance testing method

Textile antibacterial fabric performance testing methods mainly include qualitative testing methods, semi-quantitative testing methods and quantitative testing methods.

2.1 Qualitative testing method

Qualitative testing methods mainly include the American AATCC Test Method90 (Halo Test, halo method, also called agar plate method), JISZ2911-1981 (antimicrobial test method), AATCC-30 (evaluation of anti-fungal and anti-corrosion properties of textile materials) And GB/T 20944.1-2007 (Evaluation of Antibacterial Performance of Textiles Part 1: Agar Plate Diffusion Method), etc. Qualitative testing methods include inoculating test bacteria on the fabric and observing the growth of microorganisms on the fabric with the naked eye. It is based on the activity of the antibacterial agent leaving the fiber and entering the petri dish. It is generally suitable for dissolution antibacterial finishing, but is not suitable for wash-resistant antibacterial finishing. The advantages are low cost and fast speed, but the disadvantage is that it cannot quantitatively measure antibacterial activity and the results are not accurate enough.

2.1.1AATCC-90 test method

It is a rapid qualitative method for antibacterial efficacy screening of antibacterial agents. The principle is: inoculate the test bacteria on the agar medium and then stick it closely to the sample. After incubating at 37°C for 24 hours, use a magnifying glass to observe the growth of the fungi and the size of the halo in the sterile area around the sample, and compare it with the test situation of the control sample. This method can process a large number of samples at one time, and the operation is relatively simple and takes a short time. However, there are also some problems. For example, although it is stipulated that the test bacterial liquid should be cultured within a certain period of time, there is no clear regulation on the bacterial concentration. In addition, the width of the blocking band represents the diffusion and antibacterial efficacy, which is meaningful for comparison with standard fabrics, but cannot be used as a quantitative assessment of antibacterial activity.

2.1.2AATCC-90 spray method

The AATCC-90 spray method is one of the improvements to the AATCC-90 test method. It sprays a certain amount of TNT reagent on the cultured sample and observes the growth of bacteria on the sample with the naked eye. The principle of color development is that the TNT reagent is reduced by the succinate dehydrogenase of the test bacteria to generate an insoluble red pigment and appear red, thereby achieving the purpose of determining antibacterial properties. The advantage of this method is that no matter whether the sample has an inhibition zone, as long as there is bacterial growth on the plate, it will appear red.

2.1.3AATCC-90 colorimetric method

The AATCC-90 colorimetric method is also an improvement on the AATCC-90 test method. A certain amount of TNT reagent is added to the bacterial eluate on the sample after culture to develop color. After 15 minutes, the absorbance at 525nm is measured with a spectrophotometer. , to find the number of viable bacteria. However, the above two methods are not suitable for test bacteria without succinate dehydrogenase.

2.1.4J ISZ2911 Antifungal Test Method

This method is the early result of the first research idea in mold detection. Its basic principle is: evenly spray a certain amount of mixed spore suspension on the sample and culture medium, culture it for a certain period of time, and regularly observe the mold growth of the sample. The anti-fungal performance of the sample is graded according to the growth of the fungus.

2.1.5AATCC 30 test method

AATCC-30 is an evaluation of the anti-fungal and anti-corrosion properties of textile materials. The resistance of textile materials to mold and rot was determined to evaluate the effectiveness of fungicides on the antimicrobial properties of textile materials. It is divided into several methods such as soil burial method, agar plate method and humidity bottle method. The soil burial method refers to burying a sample (of a certain size) in mud for a certain period of time and then measuring the fracture strength of the sample. This method uses the fracture strength lost after the sample is buried in the soil to characterize its anti-fungal ability. The agar plate method is a method used to evaluate the ability of fabrics to resist this type of bacteria. This method is to evenly drop a certain amount of aqueous solution dispersed with Aspergillus spores on an agar plate containing culture medium, then place a sample disc treated with a non-ionic wetting agent on it, and evenly drop a certain amount of it on the sample disc. The above aqueous solution is placed at a certain temperature for a period of time, and then the growth of mold on the sample is observed. It is characterized by the mold area on the sample disk. The humidity bottle method is to suspend the pretreated sample strip in a wide-mouth bottle with certain ventilation, containing a certain amount of aqueous solution dispersed with a certain number of bacterial spores, and place it at a certain temperature for a period of time. This method also uses the mold area on the sample strip for characterization.

2.1.6GB/T 20944.1-2007 Evaluation of antibacterial properties of textiles Part 1: Agar plate diffusion method

This standard is a newly introduced qualitative test method for the antibacterial performance of textiles in China. Two layers of agar culture medium are injected into the plate, the lower layer is sterile culture medium, and the upper layer is inoculation culture medium. The sample is placed on the two layers of culture medium and incubated for a certain period of time. , based on the degree of bacterial reproduction at the contact point between the culture medium and the sample, the antibacterial performance of the sample is qualitatively evaluated.

2.2 Semi-quantitative testing method

The most commonly used semi-quantitative testing method for antibacterial properties of textiles is the parallel streaking method.

The parallel streaking method is a semi-quantitative experimental method for the antibacterial efficacy of textiles. It can be used to qualitatively test the antibacterial properties of antibacterially finished textile materials relatively quickly and conveniently. It can be used to determine the antibacterial ability of textiles with diffusible antibacterial agents. Replaced the cumbersome A A T C C-1 00.

AATCC-147 is used to evaluate the antibacterial finishing of textile materials. It is a semi-quantitative analysis of the antibacterial performance of textile materials. The AATCC-147 method is to drop a certain amount of culture fluid (containing a certain number of spores of Staphylococcus aureus and other bacteria) into a petri dish containing a nutrient agar plate, so that five parallel stripes are formed on the surface of the agar. Then place the sample vertically on these culture fluid stripes, squeeze it gently so that it is in close contact with the agar surface, and place it at a certain temperature for a certain time. This method uses the width of the antibacterial zone around the stripe in contact with the sample to characterize the antibacterial ability of the fabric.

2.3 Quantitative testing method

At present, the main quantitative testing methods for the antibacterial performance of textiles are the flask shaking method and the absorption method. The flask oscillation method is to oscillate the textile in the bacterial liquid to make the bacteria come into contact with the antibacterial agent contained in the textile. According to the change in the number of viable bacteria contained in the bacterial liquid before and after oscillation, it is used as the main indicator of antibacterial performance. The absorption method is to add drops of bacterial liquid containing a specified concentration to the textile antibacterial fabric sample and the control sample without antibacterial agents. After incubating it for a certain period of time under specified conditions, the samples and control samples before and after incubation are treated with the specified method. Wash the eluate, and then count the viable bacteria in the eluate. The antibacterial performance was evaluated by comparing the changes in the number of viable bacteria before and after culture.

The operation of the oscillation method is relatively complicated, and the scope of application in general standards is limited to non-dissolvable textile antibacterial fabrics; although the operation of the absorption method is more complicated and time-consuming, because it is a quantitative method, it is suitable for both dissolution-type and non-dissolution-type antibacterial fabrics. Textile antibacterial fabrics are more suitable, and the test conditions are closer to the actual wearing conditions of the human body, so this is the current method with the most accurate test results. From a scientific and practical perspective, this method should be the main development direction for textile antibacterial performance testing in the future.

The standards for quantitative testing mainly include the following: American AATCCTest Method 100 (bacteria count determination method), J ISL1902-8 (1998) quantitative test method, FZ/T02021-92, improved Quinn test method, ASTM E 2149-2001 (Dynamic test method for antibacterial properties of fixed antibacterial agents), GB/T20944.2-2007 (Evaluation of antibacterial properties of textiles Part 2: Absorption method), etc. The advantages of quantitative testing methods are quantitative, accurate and objective, but the disadvantages are long time and high cost. The following introduces commonly used quantitative testing methods for textiles.

2.3.1AATCC Test Method 100 test method

This law was proposed by the Standards Committee of the American Textile Printing and Dyeing Association in 1961 and was basically finalized after revisions in 1965 and 1981. This method was proposed earlier, has a greater impact, and can quantitatively detect the bactericidal and bacteriostatic abilities of the sample. The principle is: inoculate the test bacteria on the test sample and the control sample, add a certain amount of neutralizing solution after exposure for a certain period of time, shake vigorously to wash out the remaining bacteria in the sample, and measure the bacterial concentration of the eluate using the dilution plate method. , compared with the control sample, calculate the percentage reduction of bacteria on the antibacterial fabric.

The disadvantages of this method are that too many specimens cannot be tested at one time and it takes a long time; for non-dissolving samples, the antibacterial performance cannot be evaluated; there are no details.�The composition of the neutralizing solution is specified; the bacterial solution is too rich in nutrients and is too different from the actual wearing conditions; the container is too large and difficult to operate.

On the basis of absorbing domestic and foreign experience, after a large number of experiments and improvements, a systematic quantitative testing method system has been formed, which can meet the needs of antibacterial performance testing of different textile antibacterial fabrics, that is, improved AATCC-100. The main points are as follows : Change the sample of the AATCC-100 method from a circle with a diameter of 4.8 cm to a square with a side length of about 1.8 cm, and put it into a 30 mL or 50 mL Erlenmeyer flask with a lid. Use 0.85% ice-cold saline (0~4℃) instead of AATCC broth to dilute the inoculum. Dilute the bacterial strain from approximately 108~109 cfu/mL to 1×105~2×105 cfu/mL to prepare an inoculum solution. Use 20 mL, 0.85% ice-cold physiological saline instead of neutralizing agent to wash the sample.

Use the following formula to calculate the antibacterial activity and bactericidal activity of the sample: Antibacterial rate = number of viable bacteria in the blank control sample after 18h – number of viable bacteria in the sample after 18h / number of viable bacteria in the blank control sample after 18h × 100% bactericidal rate = ” The number of viable bacteria in the blank control sample at 0″-the number of viable bacteria in the sample after 18 hours/the number of viable bacteria in the blank control sample at “0” Antibacterial testing can be performed, and the nutrients of the culture medium are suitable for the use conditions of the fabric.

2.3.2J ISL1902-8(1998) Quantitative Test Method

Japanese scholars proposed improved methods for the American AATCC100 method, such as bacterial count determination method, bacterial proliferation inhibition test method, agar plate method, improved bacterial proliferation inhibition test method, improved AATCC100 test method, etc. Based on this, the Japanese Industrial Standards Committee revised JISL1902- The 1990 standard was revised and the JISL1902-1998 standard was formulated.

2.3.3FZ/T02021-92 test method

FZT01021-92 Textile Industry Standard of the People’s Republic of China, Fabric Antimicrobial Performance Test Method (hereinafter referred to as FZ/T method); place the test sample and the control sample in an Erlenmeyer flask respectively (2 bottles of test sample, 1 bottle of control sample), After adding the indicator bacteria liquid, the bacteria on the control sample and the zero-time test sample were immediately washed with buffer and the bacterial count was measured. After 20 hours of incubation at a suitable temperature, the sample was also washed with buffer, and the bacterial count was measured, and then calculated. The percentage of bacterial reduction in the sample was obtained.

2.3.4 Modified Quinn test method

Experimental principle: Drop the test bacteria liquid directly on the fabric to be inspected, so that the bacteria are fully exposed to the fabric for a certain period of time, and then cover the culture medium to allow the remaining bacteria to grow. Compare the percentage reduction of bacterial load in antibacterial samples to judge their antibacterial ability. Use a magnifying glass to count colony forming units.

2.3.5ASTM E 2149-2001 test method

ASTME 2149-2001 is an oscillation test method. The test operation is simpler than the absorption method. It is currently an ideal test method. The oscillation method is to inoculate bacteria in a certain liquid. It does not have high requirements for the water absorption of the sample. For fiber, whether it is powdered, feathers, or uneven fabrics, any shape of the sample can be applied, and it can be used for non-dissolving fibers. It is very suitable for testing both antimicrobial fabrics and dissolvable antibacterial fabrics. This testing method can test not only fabrics, but also powdery and granular materials, as well as other surface-treated solid materials.

2.3.6GB/T 20944.2-2007 Evaluation of Antibacterial Performance of Textiles Part 2: Absorption Method

This standard is a newly introduced quantitative testing method for the antibacterial performance of textiles in China. The testing principle is to inoculate the sample and control sample with test bacterial solution. Carry out immediate elution and post-culture elution respectively, measure the number of bacteria in the eluate and calculate the antibacterial value or antibacterial rate to evaluate the antibacterial effect of the sample.

3.Key factors affecting textile antibacterial testing methods

This article mainly introduces the flask shaking method and absorption method among quantitative testing methods.

3.1 Key factors that affect the results of the flask shaking test The key factors that affect the test results of the flask shaking method include test strains, detergents, washing methods, test parameters, and determination of antibacterial effects.

3.1.1 Test strains

Even if microorganisms belonging to the same species are used, if the sources are different, the sensitivities to the antibacterial finishing samples will also be different, and the measured antibacterial values will also be different. Therefore, try to use standard strains when measuring. The growth of bacteria has certain regularity and can generally be divided into four growth phases: slow phase, logarithmic growth phase, stable phase and decline phase.

The individual growth cycles of different bacteria vary in length. For example, Staphylococcus aureus, a commonly used bacterium used in textile antibacterial fabric testing, reproduces much slower than Escherichia coli and Pneumoniae. The results obtained by inoculating test bacteria at different times (that is, the activity of the bacteria are different) are also different.

3.1.2 Detergents and washing methods

Unless it is a disposable product or a sample that does not need to be washed according to the actual use, it is generally washed once before processing the test sample, firstly to remove dust or impurities on the fabric, and secondly to eliminate excessive use of antibacterial agents.

Which detergent is used in the washing test and whether the residual detergent has been cleaned has a great impact on the results. Therefore, for washing�The procedures and detergents need to be standardized, otherwise the test data from different testing units will lack comparability.

3.1.3 Test parameters

Temperature is an important factor affecting microbial growth. If the temperature is too low, bacteria will stop growing; when the temperature rises, the chemical reactions and enzyme biochemical reactions in the cells are faster, and the growth rate is accelerated. Most bacteria are mesophilic microorganisms, and their optimum growth temperature is around 37°C. The reproduction of bacteria is greatly affected by temperature, so temperature consideration is indispensable.

Among the textile antibacterial fabric testing methods, except for the “Antibacterial Knitwear” industry standard, which sets the culture temperature of the bacterial liquid on the sample to (24±1)°C, the rest are all 37°C. In order to avoid the impact of temperature differences on bacterial growth during the test and make standards comparable, the test temperature should be unified. The concentration of the inoculum solution, the activity of the inoculum, the nutritional level of the inoculum, the culture method and the time of contact between the sample and the bacterial solution, etc., all have an impact on the test results. Therefore, stipulating unified test parameters can make test results reproducible and comparable within laboratories and between laboratories.

3.1.4 Determination of antibacterial effect

Currently, antibacterial results are expressed in two ways: percentage and logarithmic value. The percentages are easy to understand, but the antibacterial differences are not obvious. For example, 90%, 99%, 99.9%, 99.99%, 99.999%… The percentage values seem to be a little bit different, but if there is an extra 9 after the decimal point, the actual value will be 10 times different, and the test report is generally Only one digit after the decimal point is retained, so the antibacterial difference cannot be displayed. Moreover, bacteria grow in a geometric progression, so it is not reasonable to express it as a percentage. The logarithmic values can be expressed as 1, 2, 3, 4, and 5 respectively, which allows people to understand the antibacterial differences at a glance, but is not easy for laypeople to understand. Considering environmental and safety factors, excessive antibacterial effects cannot be pursued one-sidedly. Most antibacterial fabrics are in direct contact with human skin and need to be washed multiple times, so their safety and sustainability should be ensured. The notes on the scope of application of the Taiwan standard CNS14945 “Performance Evaluation of Antibacterial Fabrics for General Purpose Textiles” stipulate that before testing the antibacterial performance, the applicant must attach the skin irritation (pH value 1000 mg/kg The original certified laboratory report with no death or abnormal phenomena, or a copy of the third-party inspection report and warranty provided by the raw material manufacturer . For products that do not need to be washed or do not come into contact with the human body, the requirements in this regard can be reduced. Therefore, it is necessary to set a reasonable antibacterial effect judgment value based on actual use.

3.2 Key factors affecting the test results of absorption method

The key factors that affect the test results of the absorption method include four key factors: bacterial solution preparation, control sample, sample weight and culture time, which play a decisive role in the success of the experiment [11].

3.2.1 Preparation of bacterial solution

The preparation of bacterial solution is an initial but very critical step in the antibacterial test, which directly determines the quality of bacterial growth in the test. The current preparation methods are the two-step method represented by the American AATCC 100 and the three-step method represented by the Japanese JIS L 1902. The so-called two-step method is to cultivate bacteria in the following two steps:

In the first step, use an inoculation loop to take a strain of bacteria preserved in the inoculation loop, streak it into a nutrient agar plate, and incubate the plate at 37°C for a certain period of time;

In the second step, a typical bacterial colony is picked from the plate in the first step, inoculated into nutrient broth, and cultured at 37°C for a certain period of time; then, the bacterial solution in the second step is diluted to a specified concentration.

The three-step rule is to add another step to the two-step method, that is, take an appropriate amount of the bacterial liquid cultured in the second step, add it to the nutrient broth, and culture it at 37°C for a certain period of time, and then add the cultured bacteria to the broth. The bacterial solution is diluted to the specified concentration.

In the antibacterial test, bacteria with higher activity must be used to truly reflect the antibacterial performance of textile antibacterial fabrics. In experiments, the level of activity is generally judged based on the growth of bacteria on the control sample before and after culture. The more bacteria grow, the higher their activity is; conversely, the lower they are. The experimental data of the two-step method and the three-step method show that (as shown in Table 2), the three-step method greatly improves the bacterial activity in the bacterial solution, especially for Staphylococcus aureus with poor growth activity. However, Little change occurred for the more active E. coli . The three-step method avoids the risk of using preserved strains with low activity, ensures the reproducibility of the test, and truly reflects the antibacterial performance of textiles. However, compared with the two-step method, three consecutive cultures not only take too long, but also are cumbersome to operate. It is somewhat redundant for E. coli with higher activity, because the two-step method can also achieve the results of the three-step method. , that is, as long as the increase in the number of viable bacteria on the control sample meets the requirements. In order to facilitate the operation, the appropriate method should be selected according to the bacterial species.

3.2.2 Control sample

In antibacterial testing, the selection of control samples is extremely critical. This is because the calculation of the antibacterial effect is based on the control sample. The quality of bacterial growth on the control sample determines the evaluation result of the antibacterial effect.��. Some methods select fabrics of the same type as the sample but without antibacterial agents as control samples. This is mainly set for enterprises and is mainly used for comparative testing of screening processes. However, market-based testing cannot be performed, and companies often cannot provide control samples when submitting for testing. Therefore, no matter from the perspective of experimental operability or comparability, a control sample that is both representative and applicable to most samples should be selected.

Considering that textile antibacterial fabrics are generally pure cotton textiles such as underwear, underwear, socks, etc. that have more contact with human skin, in the experiment, the cotton lining fabric for textile color fastness test specified in GB 7565287 was selected as the control sample. . In the early stage of the test, the direct test without washing the cotton lining fabric showed that the bacteria on the lining fabric were cultured for 19 hours and the viable bacteria were counted according to the ten-fold dilution method. The data were serious. Violation of the tenfold dilution rule even occurs.

3.2.3 Sample amount The amount of sample has a significant impact on the results. This is because during the process of washing the viable bacteria on the cultured sample, the antibacterial agent contained in the sample is also washed out. The larger the sample size, the more antibacterial agents are eluted. These washed antibacterial agents will further inhibit the growth of viable bacteria remaining in the washed liquid, thereby reducing the number of remaining viable bacteria. The more the sample is used, the more the number of viable bacteria is reduced, and the better the antibacterial effect is. However, this result does not truly reflect the antibacterial performance of the textile. Therefore, unifying the sample dosage is very important in antibacterial testing.

3.2.4 Culture time

Culture time refers to the time from when bacteria grow and reproduce to when their number of viable bacteria reaches a maximum value under normal circumstances. Theoretically, this time period is 18h. Among the current methods, some choose 18 to 24 h, some choose 18±1 h, and some choose 20±2 h. Basically, they are selected within the range of 18 to 24 h.

4. Conclusion

So far, there is no unified international standard for antibacterial testing. There are certain differences in experimental strains, experimental instruments, experimental methods, and evaluation methods. Different detection methods also have an impact on the test results. The country should formulate unified, authoritative and scientific textile antibacterial fabric testing standards and evaluation systems as soon as possible to regulate and supervise this market with great development potential.