Disinfectant for Urinary Infection Caused by Escherichia coli by Using Natural Oils

According to the WHO analysis, antibacterial treatment which is preferred for treating peptic ulcers and oral health problems are considered as a major improvement in the treatment but are responsible for several health problems that include vomiting, diarrhea and the increase in the bacterial resistance is a major concern. So, there is an urgent need for a good therapeutic alternative which is effective. Usage of essential oils to treat different ailments is found to be extremely effective with no side effects. Thus, essential oils are gaining im-portance in treatments due to their antibacterial, antifungal, antiprotozoal properties. Essential oils are also used as aroma therapeutic agents. They are com-plex mixture of low molecular weight compounds that include terpenoids, phenylpropanoids, oxygenated derivatives (aldehydes, ketones, alcohols, esters) which greatly differ in composition in different essential oils are the major com-ponents present. The bioactive properties and their medical potential are widely used in pharmaceutical industries. In this study, we use these essential oils to prepare a disinfectant and analyze the antimicrobial properties using the agar diffusion method. Our major concern in this study is to decrease the growth of Escherichia coli which causes urinary infection, by using disinfectant prepared using a mixture of essential oils of particular concentration.


Introduction
Disinfectant is considered as the agents when applied to inanimate objects inhibit the growth of microorganisms. When studied about the traditional medicines, Journal of Biosciences and Medicines and also in case of surgical conditions [1]. Thus the properties of these oils were studied and found to have antipathological activity. These are secondary metabolites of the plants and are prepared using steam distillation and solvents. About 43% of people find aromatherapy to be effective in relieving stress. Peppermint oil relieves people from migraine; Lavender oils helps to treat insomnia conditions in people with heart ailments. Thus, the idea of usage of these valuable essential oils in disinfectant preparation of ideal concentrations [2] and used to prevent the contamination by Escherichia coli. Urinary infection is more prevalent in the females compared to males; the risk factors include usage of poor hygienic washrooms, menopause and pregnancy (Wiley Online Library). The bacteria enter the urinary tract through urethra and multiply in bladder. The bacteria responsible for bladder related urinary infection is Escherichia coli [3]. By using essential oils such as clove oil, rose oil, cinnamon powder and peppermint oil of ideal concentration (500 µg/ml), using ethanol as a solvent [4], the disinfectant was prepared and agar diffusion method was performed to find the growth of E. coli to be decreased, thus maintaining hygienic conditions [5].

Preparation of Different Oils
Commercial oils were taken which have undergone FDA approved purity checks and a mixture of ideal concentration (500 µg/ml) by using ethanol as the solvent. Figure 1 shows the commercial oils used [6].

Processing and Evaluation of Disinfectant
To find out which bacteria is present in major constituent in washrooms, the petri dishes are prepared under sterilized condition in laminar air flow chamber, the lid is opened to allow the contamination of the bacteria. Figure 2 shows the colonies of the bacteria in the petri dishes after incubation. Figure 3 shows that   the petri dishes are incubated (37 degree Celsius) for 24 hours. The bacteria is isolated and identified by biochemical test where a test kit named KI1001 AND KI1002 is used. The petri dish is swabbed aseptically by the isolated bacteria and by following the guidelines the agar diffusion method was performed by using the disinfectant made using essential oils [7]. The growth of the Escherichia coli was no more than 20%. The inhibition ranges from 6.4 mm to 11 mm. The lipopolysaccharides of the Gram negative bacteria might be responsible for quite good resistance of the bacteria.
S. Harini et al.

Staining Techniques
The staining techniques are done to identify the type of the bacteria [8] present. Figure 4 shows the staining done using Gram staining and found out that it is Gram negative bacteria.

Staining for Examination
Different staining techniques like simple stain, Gram stain and Negative stain were performed.

Biochemical Test Kit
A kit that consists of different tests and by examining the color change the isolated bacteria is identified. The kits used are KI1001 and KI1002.

Clove Oil Preparation
Take 20 g of cloves in a mortar and pestle. Grind the cloves and put it in a white cloth. Tie it in one end of the cloth. Put it in the olive oil in small container. Place the small container in water containing big container. Boil for 2 hours. Allow the oil to cool and take the oil and crush the cloth to get the clove extract [9].

Preparation of Rose Oil
Fill the rose petals in the container with oil. Place the container in the big container containing water. Heat the container for 2 hours and cool it, then again heat it for 2 hours for 6 times. Cool down and extract the oil from the rose petals.

Preparation of Mint Oil
Crush the peppermint leaves with your hand or spoon. Put the leaves in the container. Fill the jar with olive oil covering the leaves. Let the oil steep for 2 days in sun and shake every 12 hours and keep it in the dark place.

Preparation of Cinnamon Powder
Take about 20 g of cinnamon in the mortar and pestle and grind it to get in powder form.

Preparation of Borneo Camphor Powder
Take about 10 g of Borneo camphor and smash it to make it into powder form.
Lavender oil, Tea tree oil, Glycerol, apple cider vinegar is also used.
The oils and the materials are taken in different proportions and mixed using the mixer for 10 -15 minutes to make it into homogenous phase.
This disinfectant can be used in the floor of restrooms and also as aerosols.
Commercial oils obtained is FDA approved purity check was used for the preparation of the disinfectant and zone of inhibition was measured [10].

Procedure for the Biochemical Test Kit
Two kits KI1001 and KI1002 were taken. It consists of nearly 12 tests in each kit.
KB0010 was gram negative rods and KB0020 was gram positive rods.
In KI1001-the tests that are performed to identify the bacterial colonies in-

Figures and Tables
Gas chromatography results that were reviewed from a few research journals, Table 1 and Table 2 provide the information about the major antibacterial compound present in the essential oils were studied. Table 3 represents the Single factor ANOVA is performed using Excel and the F value is calculated.
Since F critical value is greater than F value, there is not much significant difference between the different concentrations of oils taken. The values taken were the number of bacterial cells [11] that was resistant to the application of different concentrations of oils. The bacterial cells were counted using colony counters [12].
This implies that when different concentrations of the disinfectants were pre-

Limitation
Although essential oils were found to have no side effects, when applied directly to skin in case of migraine, insomnia patients. It causes health problems that include rashes, asthma attacks (in case of aroma therapy), headaches and allergic reactions. Compound with high phenol content like cinnamon cause skin irritations. Swallowing of essential oils is not advisable.

Results
All the petri dishes that were taken in the wash rooms were contaminated by bacteria, and it was identified as Escherichia coli. Figure

Discussion
Poor hygienic condition resulted in urinary infection problems in youngsters; to avoid the problem the disinfectant without the presence of ethanol is prepared. 80% of students find it hygienic after usage of the disinfectant. This is prepared by the usage of natural oils which consists of antimicrobial properties and odor properties.