Why is 70% ethanol used not 100%?

At a 70% concentration, however, the ethanol takes longer to evaporate and this longer contact time means that it is able to penetrate cells more effectively . The water in a 70% grade is also important in denaturing proteins. And 70% ethanol also tends to be cheaper and is less flammable [1].

Why do we use 70 ethanol for sterilization?

70% denatured alcohol penetrate the cell wall more completely which permeates the entire cell, coagulates all proteins, and therefore the microorganism dies . Extra water content slows evaporation, therefore increasing surface contact time and enhancing effectiveness.

How is phenol used in microbial control?

Phenol and phenol derivatives agents kill most bacteria, most fungi, and some viruses, but are usually ineffective against endospores. They alter membrane permeability and denature proteins .

Why is 70 percent alcohol better than 100?

Even though you may think the higher concentration is more effective, experts say 70% is actually better for disinfecting. It has more water, which helps it to dissolve more slowly, penetrate cells, and kill bacteria . The disinfecting power of rubbing alcohol drops at concentrations higher than 80%-85%.

How do alcohols control microbial growth?

Alcohols make up another group of chemicals commonly used as disinfectants and antiseptics. They work by rapidly denaturing proteins, which inhibits cell metabolism, and by disrupting membranes, which leads to cell lysis . Once denatured, the proteins may potentially refold if enough water is present in the solution.

Which two alcohols are effective in microbial control?

Alcohols. Although several alcohols have been shown to be effective antimicrobials, ethyl alcohol (ethanol, alcohol) , isopropyl alcohol (isopropanol, propan-2-ol) and n-propanol (in particular in Europe) are the most widely used (337).

Why can bacteria still grow under 100% ethanol?

Therefore, the ethanol has to pass the bacterial membrane/wall to get into the bacteria - if you use 100% ethanol instead, the bacteria get 'sealed' and they will survive ... Another mechanism is the high osmotic pressure of ethanol/water-mixtures; and the 70% has the highest one.

Why don't we use 100% ethanol?

Is hygroscopic, meaning it absorbs water out of the air, which means it can damage engines due to excess water content if not handled carefully . Attacks the seals and hoses used in most vehicles -- cars require special design to resist polar solvents in order to use ethanol blending levels greater than 10-15%

Why we use 70 ethanol instead of 100 in DNA extraction?

Usually, about 70 percent of ethanol solution is used during the DNA washing steps. This allows the salts to dissolve while minimizing DNA solubility . The last 100 percent ethanol wash which is mainly employed helps to promote convenient ethanol evaporation from DNA pellet, thus preventing any carryover.

Why do we use 70% ethanol instead of 90% or 95% ethanol to sterilize the cell spreader?

Since a 70% ethanol solution has more water, it takes longer to dissolve and thus can actually penetrate the bacterial cell wall and kill it .

Why is 100% ethanol ineffective?

100 % ethanol would evaporate very rapidly and therefore would not be able to kill the bacteria. However, if it is 70% or 80% ethanol and 20% or 30% water, the ethanol would not evaporate very rapidly. Since the ethanol is not evaporating rapidly, it will enter the cell and denature proteins inside the cell.

Efficacy of 75% alcohol in pretreatment of the Andersen sampler in trapping maximum airborne microbes (2024)

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PMC7781652

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Aerobiologia (Bologna). 2021; 37(1): 171–178.

Published online 2021 Jan 5. doi:10.1007/s10453-020-09668-2

PMCID: PMC7781652

PMID: 33424104

Yan Zhao,¹ Jingwei Zhang,¹ Shumei Wang,² Lingqi Yu,² Hao Yu,¹ Yuwen Wang,¹ and Lihong Feng¹

Author information Article notes Copyright and License information PMC Disclaimer

Associated Data

Supplementary Materials

Abstract

The present study was conducted to evaluate the effects of the pretreatment methods and sampling time on the sampling of airborne bacteria in hospitals. Methods for the pretreatment of Andersen samplers, namely, non-sterilized, 75% ethanol and autoclaving sampled for 5min, 10min and 15min in the general ward and class 1000 clean operating department, respectively, were studied. Statistical analysis was used to compare the differences in sampling results of airborne bacteria under different pretreatment methods, sampling time and environmental conditions. In the first test, the sampling results of the airborne bacteria obtained by pretreatment of the sampler with 75% ethanol and without pre-treatment were not very different, and the sampling results showed a certain declining trend with the extension of the sampling time. In the second test, the pretreatment effect of autoclaving was significantly better than that of 75% ethanol, and the sampling time had no effect on the sampling results. After removing the influencing factors of the environment, the results were consistent with the results of the second test. It was observed that the Andersen samplers should not be pretreated with 75% ethanol before airborne microbes sampling. The pretreatment should be carried out by autoclaving, and the sampling time has little effect on the sampling results.

Electronic supplementary material

The online version of this article (10.1007/s10453-020-09668-2) contains supplementary material, which is available to authorized users.

Keywords: Pretreatment methods, Hospital disinfection, Airborne microbe, Andersen sampler, Clean operating department

Introduction

Airborne transmission is one of the most frequent causes of hospital infections (Eames et al. 2009; Kozajda et al. 2019). Dangerous infectious coronavirus disease 2019 (COVID-19) spread via airborne during aerosol generating procedures (Doremalen et al. 2020), and 20% of COVID-19 infections occurring in health care personnel in hospital of Italy (Remuzzi et al. 2020). Some studies have also reported that bacteria and viruses are present in aerosol particles (Kirby et al. 2016; Pichon et al. 2019; Núñez et al. 2017).

The airborne microbe test is a frequently used sampling method for preventing indoor infections in public places and hospital wards. This testing is an effective way to identify air pollution in the hospital as soon as possible and to prevent the occurrence of nosocomial infection. The total number of bacterial colonies refers to the number of collected bacterial colonies grown on the nutrient agar plate under certain conditions (such as aerobic, nutritional, pH, temperature and time conditions) (Li et al. 2019; Harp 2018). Certain conditions refer to culture at 35–37°C for 48h under aerobic conditions (General Administration of Quality Supervision, Inspection and Quarantine of the People's Republic of China. 2010). Therefore, it is difficult for anaerobic or micro-aerobic bacteria with special nutritional requirements and nonmesophilic bacteria to reproduce under these conditions, because the existing conditions could not satisfy the physiological requirements for their growth.

Andersen six-stage sampler is a sensitivity instrument which could size and counted airborne particle aerosols, which was of great value in controlling sources of airborne disease (Andersen 1958; Buttner et al. 1993). Andersen sampler will not be soiled or contaminated by proper handling and operation, except for very special detection studies or work with pathogenic aerosols; it does not require cleaning or sterilization between runs, and only need passing very clean air before sampling to eliminate particles that can clog the holes in the stages. However, there are numerous pathogenic aerosols in the hospital environment, and frequent sampling of airborne microbe in the hospital environment will pollute the Anderson sampler (Charifa et al. 2017; Joost et al. 2019; Sedighe et al. 2019). Therefore, before using the Anderson sampler for air sampling in a hospital environment, it is necessary to perform a proper pretreatment on the sampler (Sung et al. 2014). China's architectural technical code for hospital clean operating department and indoor air quality standard also requires aseptic operation when using the Andersen sampler for airborne microbe sampling (Ministry of Housing and Urban–Rural Development of the People’s Republic of China 2013; General Administration of Quality Supervision, Inspection and Quarantine of the People's Republic of China. 2002). Pretreatment of the Andersen sampler is also part of the aseptic operation, but the current standards have not yet clearly defined the pre-treatment method of the Anderson sampler. The use of 75% ethanol as cleaning pre-treatment of the Andersen stages was introduced by manufacturers of many sampler models, often used in the daily sampling of airborne microbe in the hospital environment. Moreover, the diameter of airborne particle aerosols usually determined the number of the deposition step, in the case of Andersen multistage. However, it is often found that the sampling result of the old Anderson sampler is higher than that of the new one in our daily sampling work. Therefore, the processing effect of the pretreatment methods of Andersen sampler is worthy of attention.

Currently, little research has been performed on the effects of cleaning and disinfecting Andersen samplers on sampling. Noro et al. (1998) used an Andersen sampler to sample and count bacterial cells and airborne dust and found that a clean environment was effective in reducing airborne environmental contamination. The effect of different pretreatment methods on the sampling results of Andersen samplers has not been reported. In this study, the differences in sampling results with two types of disinfection of an Andersen sampler were compared in a general hospital ward and a class 1000 operating clean room. The effects of sampling time and disinfection methods on the sampling results of airborne microbes were discussed. Evidence is provided for selecting a reasonable sampling time and effective disinfection method for infection detection in hospitals.

Methods

The study was conducted at a general tertiary hospital in Tianjin as the experimental site. A common ward and a class 1000 operating clean room were selected randomly. Airborne microbes were sampled with Andersen six-stage sampler (QT-30, SKC, USA).

Sampling and processing of airborne microbes

Each level of the Andersen sampler was supported by a nutrient agar plate (90mm, 190530, Tianjin Nuofan Biological Technology Co., Ltd, China) and sampled at a flow rate of 28.3L/min. The trial flow control system consisted of a primary calibrator (4046, TSI Incorporated, USA), which controlled the sampling flow rate. For the first trial in the general ward, one Andersen sampler was sterilized with 75% ethanol, and the other Andersen sampler was not sanitized before sampling. For sterilization, the surface, back and sealing ring of each stage of the sampler, as well as around the air intake, were wiped with cotton balls soaked with 75% alcohol. The temperature of the general ward was 24°C, and the humidity was 55%. The door and window of the general ward were closed.

For the second trial in the general ward, one Andersen sampler was sterilized with 75% ethanol in the same way as the first trial. After removing the seal ring, each stage of another Andersen sampler was individually packaged and autoclaved (HV-110, HIRAYAMA, Japan) at 102.9kPa and 121°C for 30min. The atmospheric conditions and background of the general ward were the same as in the first trial. For the third trial in the hospital class 1000 operating clean room, one Andersen sampler was sterilized with 75% ethanol, and the other Andersen sampler was autoclaved at 102.9kPa and 121°C for 30min before sampling.

In the three experiments, the two samplers were sampled for 5min, 10min and 15min. Each sampling period was repeated six times. Each sampler was sterilized or autoclaved before each sampling period. Sampling for the next cycle was performed after each sampling cycle was completed.

Cultivation and counting of airborne microbes

After the completion of all samplings the petri plates were incubated in a 35–37°C incubator (LRH-250A, Guangdong Medical Instrument Factory, China) for 48h (General Administration of Quality Supervision, Inspection and Quarantine of the People's Republic of China. 2010). Colony forming units were counted after the completion of the culture, and the results were recorded and converted to CFU/m³ according to the dilution ratio and volume of the sampling gas. The reference formula is as follow:

$Total bacteria in the air (CFU / m^{3}) = \frac{Total number of bacteria on the six - sampling plate}{28.3 L / min \times sampling time} \times 1000$

Statistical analysis

Total bacterial growth was derived from nutrient agar plates and calculated in CFU/m³. All calculations were performed using Statistical Package for the Social Sciences Version 24.0 (IBM Corp., Armonk, NY, USA). As the variables of total bacterial colonies follow nonnormal distribution, a Wilcoxon rank-sum test was performed. The statistical results of the Wilcoxon rank-sum test of two independent samples were represented by the Z value, and the test results of several independent samples were represented by the χ² value. The testing level was α = 0.05.

Results

Bacterial counts from first trial at general ward

The results of first trial revealed that there was no significant statistical difference between the pretreatment with 75% ethanol and no pretreatment (Z = 0.791, P = 0.443). A total 642 colonies (Median) of the no pretreatment group was recorded ranging from 215CFU/m³ to 3152CFU/m³. A total 596 colonies (Median) of the 75% ethanol pretreatment group was recorded ranging from 217CFU/m³ to 2880CFU/m³. Further analysis of the results of sampling at different time periods also showed that there was no significant statistical difference between the sampling results of the two pretreatment methods (Z_5 min = 0.961, P_5 min = 0.394; Z_10 min = 0.801, P_10 min = 0.485; Z_15 min = 0.401, P_15 min = 0.699). With pretreatment of the Andersen air microbial sampler with 75% ethanol, no statistically significant differences were observed between different sampling periods (χ² = 5.801, P = 0.055). When the Andersen air microbial sampler was not pretreated, the 5min test results were significantly higher than those of the 10min and 15min tests, and the difference was statistically significant (χ² = 7.871, P = 0.020). The test results of the first airborne bacteria sampling in the general ward are shown in Fig.1 and Table Table11 (Table S1, Supporting Information).

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Fig. 1

Airborne bacteria sampling results of the first trial in the general ward

Table 1

The three trials results of airborne microbe sampling in hospital (M, median)

Time	Frequency	No pretreatment		75% ethanol		Autoclave		Z	P
Time	Frequency	Range (CFU/m³)	M (CFU/m³)	Range (CFU/m³)	M (CFU/m³)	Range (CFU/m³)	M (CFU/m³)	Z	P
First test	18	215–3152	642	217–2880	596	–^*	–^*	0.791	0.443
Second test	18	–^*	–^*	288–1400	485	177–799	311	2.674	0.007
Third test	18	–^*	–^*	179–1152	298	5–36	15	5.130	0.000

Open in a separate window

^*Indicates that no such result

Bacterial counts from second trial at general ward

The results of second trial revealed that the airborne microbe sampling results with the sampler pretreated by autoclaving were significantly lower than those of the sampler pretreated with 75% ethanol (Z = 2.674, P = 0.007). A total 311 colonies (Median) of the autoclaving group was recorded ranging from 177CFU/m³ to 799CFU/m³. A total 485 colonies (Median) of the 75% ethanol pretreatment group was recorded ranging from 288CFU/m³ to 1400CFU/m³. Regarding sampling at different time intervals, the autoclaved sampler’s results of the airborne microbe sampling for 5min and 10min were significantly lower than the results of the sampler pretreated with 75% ethanol, but the results of sampling for 15min were no statistically differences (Z_5 min = 2.882, P_5 min = 0.004; Z_10 min = 2.242, P_10 min = 0.025; Z_15 min = 0.961, P_15 min = 0.337). The 5min sampling result of the pretreatment with 75% ethanol was significantly greater than that of the other two periods (χ² = 12.538, P = 0.002). For the pretreatment of the Andersen air microbial sampler by autoclaving, there was no statistically significant difference in the sampling results at different time points (χ² = 2.356, P = 0.308). The test results of the second airborne bacteria sampling in the general ward are shown in Fig.2 and Table Table11 (Table S2, Supporting Information).

Open in a separate window

Fig. 2

Airborne bacteria sampling results of the second trial in the general ward

Bacterial counts in class 1000 operation clean room

The results of the class 1000 operation clean room revealed that the airborne microbe sampling results of the sampler pretreated by autoclaving were significantly lower than those of the sampler pretreated with 75% ethanol (Z = 5.130, P = 0.000). A total 15 colonies (Median) of the autoclaving group was recorded ranging from 5CFU/m³ to 36CFU/m³. A total 298 colonies (Median) of the 75% ethanol pretreatment group was recorded ranging from 179CFU/m³ to 1152CFU/m³. The autoclaved sampler’s sampling results of 5min, 10min and 15min were significantly lower than the results of the sampler pretreated with 75% ethanol (Z_5 min = 2.882, P_5 min = 0.004; Z_10 min = 2.892, P_10 min = 0.004; Z_15 min = 2.892, P_15 min = 0.004). The sampling result of 5min in the class 1000 operating clean room of the pretreatment with 75% ethanol was significantly higher than that in the other two periods (χ² = 15.174, P = 0.001). For the sampler pretreated by autoclaving, there was no statistically significant difference in the sampling results at different time points (χ² = 3.797, P = 0.150). The test results of the airborne bacteria sampling in the hospital class 1000 operating clean room are shown in Fig.3 and Table Table11 (Table S3, Supporting Information).

Open in a separate window

Fig. 3

Airborne bacteria sampling results of the third trial in the hospital class 1000 operating clean room

The above three trials showed that 75% alcohol could not make the Anderson sampler achieve the expected pretreatment effect. Using the autoclaved Anderson sampler, sampling for 5min could correctly reflect the airborne microbes’ pollution level of the detected environment.

Discussion

Ethanol (75%) is a widely used medium-effect disinfectant. It has a good sterilizing effect on most bacteria and viruses, but it has no obvious sterilizing effect on bacterial spores (Xue 2010). In the first trial, there is little difference between the results of the pre-treatment of the Andersen air microbial sampler with 75% ethanol and the results of the non- sanitized sampler. The test results showed that 75% ethanol has a limited ability to rapidly sterilize the Andersen’s air microbial sampler. Other studies have found similar situations (Stewart et al. 2019; Sui et al. 2012). For example, in the observation of the disinfection effect of different chemical disinfectants, it was found that the microbial yield of the surface of the object disinfected by 75% ethanol was significantly lower than that of other chemical disinfectants. However, some previous research conclusions were contrary to ours. Sandle and Satyada (2015) pointed out the efficiency of 70% isopropyl alcohol for disinfection of the Andersen sampler and Trissel et al. (2007) indicated that repeated disinfection with IPA decreased the contamination rate. This may be because isopropyl alcohol contains two methyl groups, which greatly enhance its lipophilicity, while alcohol contains only one ethyl group. Ethyl is less lipophilic than methyl. 75% ethanol did not reflect the expected disinfection effect, which may be related to the external temperature (Mohanta and Jana 2016), defects with the disinfectant or the fact that the 75% ethanol disinfectant was not suitable for disinfection. In the first test, we also found that the sampling results showed a certain downward trend with increasing sampling time.

The second trial results showed that the pretreatment effect of autoclaving was significantly better than that of 75% ethanol. Relevant research has indicated the same results; Lin et al. (2018) sterilized Bacillus subtilis spores loaded on an N95 filtering face piece respirator with different disinfection methods, and the results showed that the effect of high-temperature autoclaving was higher than that of other disinfection methods. Some studies have observed the morphology of live bacteria disinfected by 75% ethanol and disinfected by autoclaving with field emission scanning electron microscopy. After disinfection with 75% ethanol, it was found that the number of live bacteria on the surface of the sample was still large, and the shape of the bacteria was still relatively complete. However, after high-pressure steam sterilization, the number of viable cells on the surface of the sample was small, and the morphology was destroyed (Garibaldi et al. 2017; Darshan et al. 2019; Tan et al. 2013).

Different from the results of the 75% alcohol pretreatment in the first trial, this experiment revealed that the sampling time had an effect on the results with the 75% ethanol pretreatment but had little effect on the results with the autoclaving pretreatment. This situation may be because the two experiments were not performed at the same time, and the air pollution conditions of the general ward may have changed. However, the tendency of the total number of bacterial colonies to decrease as the sampling time increased was still apparent. The results of the second trial preliminarily determined that 75% ethanol is not suitable for disinfecting the Andersen air microbial sampler. But, the influence of the change in the environment during the sampling time should not be ignored. It was necessary to more accurately analyze the pretreatment effects of autoclaving and 75% ethanol on the sampling results while eliminating the factors of sampling environmental pollution conditions as much as possible.

The sampling environment could be effectively controlled, when the airborne bacteria sampling test was carried out in the class 1000 operating clean room and the self-cleaning degree of the sampler after pre-treatment would directly affect the sampling result. The results of the third test showed that the pretreatment effect of autoclaving on the Andersen sampler is significantly better than that of the sampler using 75% ethanol and further proved that the length of the sampling time has little effect on the results after autoclaving.

It was found that the sampling results of airborne microbes greatly affected by the self-cleaning degree of the Andersen sampler. If the Andersen sampler is not sterilized effectively before sampling, the authenticity of the sampling results will be seriously affected. Although the number of samples in this study is limited, it still can be suggested that the sampling in hospital environments, for the determination of the airborne microbe, should require a more rigorous pretreatment through autoclaving. However, the process of autoclaving pretreatment is complicated, so it is necessary to prepare several Anderson samplers for autoclaving pretreatment at the same time before sampling airborne microbes in multiple hospital wards.

In conclusion, this study found that the Andersen sampler needs to be pre-processed before sampling the airborne bacteria in the hospital indoor environment. Ethanol (75%) has a poor pretreatment effect on the sampler and should be avoided in daily sampling. Autoclaving has a good pre-treatment effect on the Andersen sampler, and could be widely applied to the pretreatment of the Andersen sampler before indoor airborne bacteria sampling (Table (Table1).1). However, since this study was only conducted in one hospital, it is inevitable that the results will have certain limitations. Therefore, further expansion of the scope of the experiment will help strengthen hospital disinfection management work. The effective pretreatment method for sampler disinfection should be correctly selected, which will also provide an accurate basis for more effective protection of the indoor air quality of the hospital and could be useful for operators who do not have sufficient scientific and cultural knowledge on aerobiological monitoring.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary file1 (DOC 58 kb)^{(54K, doc)}

Acknowledgements

We acknowledge the contribution of all the members who participated in the sampling and analysis.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

No ethical approval was required or sought for the study, because there was no interaction or intervention with human participants.

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Articles from Aerobiologia are provided here courtesy of Nature Publishing Group

Efficacy of 75% alcohol in pretreatment of the Andersen sampler in trapping maximum airborne microbes (2024)

Associated Data

Abstract

Electronic supplementary material

Introduction

Methods

Sampling and processing of airborne microbes

Cultivation and counting of airborne microbes

Statistical analysis

Results

Bacterial counts from first trial at general ward

Table 1

Bacterial counts from second trial at general ward

Bacterial counts in class 1000 operation clean room

Discussion

Electronic supplementary material

Acknowledgements

Compliance with ethical standards

References

FAQs

Why is 70% ethanol used not 100%? ›

Why do we use 70% ethanol instead of 90% or 95% ethanol to sterilize the cell spreader? ›