Preparation of Additol Antiseptic and Disinfectant Solution

Introduction

Terpineol is a naturally occurring mono terpene alcohol and is extracted from leaves of Pinus by distillation fraction method. It has balsamic turpentine like aroma and has excellent antibacterial activity.^1-4

Basil oil is an essential oil which has significant biological activity against insect pests. It is also called as sweet basil essential oil, which is obtained from the leaves of Ocimum basilicium, basil herb and extracted by steam distillation or solvent evaporation method using Clevenger apparatus.^5-6

Propylene glycol (IUPAC name: propane -1,2-diol) is a viscous colourless liquid, also nearly odourless but possesses a faint sweet taste with a chemical formula of CH₃CH (OH) CH₂OH. It exhibits bactericidal activity at 100% concentration.^7-8

Liquid caramel is a colouring agent, which is a water-soluble colouring made by heat treatment of carbohydrates.⁹ Castor oil is used in the preparation of soap solution and is extracted from castor beans of Ricinus communis plant by mechanical pressing or solvent evaporation.¹⁰ Sodium hydroxide (NaOH) is also used in the preparation of soap solution. It is an essential ingredient in soap making, and causes saponification.^11-12

Materials and Methods

The following materials were used in the present study.

Apparatus

·          Sterilized stainless steel container

·          Test tubes

·          Beakers

·          Volumetric flasks

·          Measuring cylinders

·          Thermometer

·          Magnetic stirrer

·          Induction stove

Chemicals and Reagents

Castor oil and Sodium hydroxide (NaOH) were used in the preparation of soap solution. We used Terpineol oil, Basil oil, Propylene glycol, and liquid caramel in further preparation.13-14

Preparation of soap solution

Castor oil was transferred to stainless steel container (1.5 L). It was heated on an induction stove until the temperature reached to 80⁰C; then at 80⁰C sodium hydroxide solution was added, stirred and continuously heated. As it formed a thick mass, the heating was continued at slow heat. It again formed a thick solution and at this point the solution was tested for saponification by taking a drop of the formed sap into a test tube containing water. The soap solution was obtained once the saponification test was passed.¹⁵

Then warm water was added slowly to the formed mixture up to a volume of 1L using purified water and this solution was used for making the antiseptic liquid.

Antiseptic liquid was prepared by following steps:

• Soap solution was taken in a beaker (1000 ml) and was kept for stirring on a magnetic stirrer.
•       Terpineol oil and Basil oil were dissolved in propylene glycol.
•     The above terpineol mixture was then added to the soap solution while stirring, and the stirring was  continued for 30 minutes.
•     After 30 minutes, liquid caramel was added to the solution while stirring was continued for 5 more minutes.
•      The final   solution   was   taken   up   for   several disinfectant tests. Then the solution was transferred to a suitable bottle and was labelled.

   Results

Validation report

The methods used for disinfectant testing were:

·          Carrier test

·          Suspension test

·          Capacity test

·          Practical test

·          In-use test

Carrier test

It is the oldest test, also called as R. Koch’s test. The disinfectant solution prepared was transferred to the petri plate containing Staphylococcus aureus bacteria culture in a drop wise manner. No growth occurred and thus the prepared disinfectant was found to be more effective.

Capacity test:

The test performed was Kelsey-Sykes test. Kelsey- Sykes test is a triple challenge test. Here, the capacity of disinfectant was determined by three successive consecutive additions of bacterial suspension until the disinfectant capacity to kill was exhausted.

Suspension test:

Here the sample of bacterial culture was suspended into the disinfectant solution. After exposure, it was verified by subculture of bacterial culture. After subculture, if no growth occurs, it indicates a more effective disinfectant.

Practical test

Filter paper test

Here the discs of filter paper were used. Filter paper was soaked with a chemical disinfectant. They were placed on agar plate and incubated. Clear zone appeared around the disc, and the disinfectant solution was found to be effective.

In-use test

It was done in laboratory to detect contamination of disinfectant. 1 ml of sample disinfectant was added to 9 ml of diluent. 10 drops (0.02 ml each) of diluted sample were placed on each of two nutrient agar plates. One was incubated at 37^oC for 3 days. The other was incubated at room temperature for 7 days. No colonies were found indicating no contamination in disinfectant.

Rideal Walker coefficient test (RWC) test

The test was done with parent culture, incubated for 24 to 48 hours and compared with standard phenol solution. It was checked at different intervals of 2.5 min, 5 min, 7.5 min and 10 minutes and test proved to be effective as value was found to be more than one.

Discussion

The prepared Additol solution was subjected to different tests. When a sample of solution was transferred to petri plate, no growth occurred and in the capacity test, bacterial suspension was made until the total was exhausted with three challenge numbers.

In the next test, after the solution was exposed, it was verified by bacterial subculture. Filter paper soaked on agar plate produced clear zone around the disc.

Disinfectant contamination with two nutrient agar plates incubated at 37⁰C for three days showed no colonies. Additol solution was compared with standard phenol solution at different intervals of times and was proved to be effective. The results were in good agreement, and were performed under the testing guidelines. Test was carried out to determine the antimicrobial activity, and the obtained results were enclosed by standard limits.

By following the standard procedure, under the FDA testing guidelines, prepared product Additol was compared with marketed product, Dettol by using Riedel-Walker Co-efficient test and the procured values were within the range.

Conclusion

The proposed Additol 1000 ml was evaluated for the disinfectant activity following the standard procedure. The proposed disinfectant exhibited good disinfectant activity against the test microorganisms.