INTRODUCTION:

Eugenol^1-2 is chief chemical constituent of clove oil. Eugenol has number of pharmacological actions such as Analgesic, Antioxidant, Anesthetic, and Antimicrobial Anticancer Anti-inflammatory and Anticonvulsant. Eugenol has it’s applications in the fragrance and flavoring industries also. Number of novel drug delivery systems of eugenol have been reported for a variety of applications. Thus development of a validated UV spectrophometic method for Eugenol will be always be advantageous as the method is simple and rapid.

Fig. 1: Chemical structure of Eugenol.

MATERIAL AND METHODS: ^{3- 5}

Materials:

Eugenol (pure) was gifted by S.H. Kelkar and Co. Pvt. Ltd. Mumbai, India. Ethanol was purchased from Research Lab Fine chem. Industries, Mumbai.

Equipments:

Single pan Electronic Balance (ohaus corp, USA AR0640 and UV spectrophotometer (pharmaspec 1700, shimadzu, Japan) was employed in the study.

Preparation of standard solutions:

Eugenol (100mg) was taken and dissolved in Ethanol and then volume was made up 100 ml with Ethanol. The dilutions were made by ethanol to obtain standard solution in the range 15-50 µg/ml. The absorbance of the resultant solutions was determined at the λmax of 282 nm.

Accuracy:

Three concentrations as 20 µg/ml, 30 µg/ml, and 40 µg/ml were prepared nine measurements are carried out and averaged. Result was compared to certificate of analysis.

Precision:

The precision of the method was assessed by analyzing eugenol for intra-assay precision and intermediate precision

A) Intra-assay precision:

Intra-assay precision was assessed by analyzing eugenol in three different concentrations (20, 30, 40µg/ml) of three times a day. The % RSD was calculated for absorbance.

Fig. 2: UV spectrum of eugenol in ethanol

B) Intermediate Precision:

Intermediate precision was carried out by doing full analysis of 30 µg/ml concentration by change in analysts within the same laboratory followed by calculation of % RSD.

Linearity and Range:

The Calibration curve was plotted using the concentration range of 15-50µg/ml .The absorbance of the solutions was determined at 282 nm. A calibration curve was constructed by plotting absorbance Vs concentration was determined.

Ruggedness:

Ruggedness was measured by the triplet measurements of 15µg/ml and 30µg/ml by change in operators in same lab. The % RSD was calculated for absorbance.

Robustness:

Robustness was measured by the triplet measurement of 30µg/ml concentration by slight change in concentration of solvent ethanol from 100% to 95 % followed by calculation of %RSD

RESULT AND DISCUSSION:

UV -Specrophotometry:

The UV method for the estimation of Eugenol in ethanol was validated . Fig 2 shows the UV spectrum of Eugenol in ethanol.

Accuracy:

Accuracy was investigatigated by analyzing three concentrations (20, 30, 40µg/ml) the average % purity of Eugenol concentration in sample was found to be 98.87%

Precision:

A) Intra assay precision:

Intra assay precision was assessed by analyzing Eugenol in three different concentration (20, 30, 40µg/ml) of Eugenol three times a day. The % RSD was found to be 1.78%.

B) Intermediate precision:

Intermediate precision was assessed by analyzing Eugenol by change in analyst. The %RSD was found to be 1.30%

Linearity and range:

The absorbance of the prepared standard solutions (15-50µg/ml) was determined at 282 nm the mean absorbance was found to be .3131-1.016.the plot of absorbance versus concentration (fig.3) obeyed Beer-Lambert’s law in above concentration range with regression coefficient of 0.991

Ruggedness:

Ruggedness was assessed by change in operator and change in concentration the % RSD was found to be 0.52%

Robustness:

Robustness was measured by slight change in concentration of solvent ethanol. The % RSD was found to be 1.74 %

Table1: % purity data for accuracy of UV method in ethanol

Concentration(µg/ml)	Absorbance	% Purity
20	0.3748	97.05
20	0.3798	98.35
20	0.3776	97.80
30	0.5436	93.86
30	0.5615	96.96
30	0.5622	97.06
40	0.7964	103.15
40	0.7950	102.97
40	0.7912	102.47

Fig.3: Calibration curve of eugenol in ethanol by UV spectrophotometry (282nm)

Table 2: Linearity and Range

Concentration(µg/ml)	15	20	25	30	35	40	45	50
Absorbance	0.3131	0.424	0.488	0.6538	0.7421	0.7964	0.915	1.016

Table 3: Validation Parameters

Parameters Done	Standard Values	Practical Values
Accuracy	≥95%	98.87
Intra-assay precision	RSD≤2%	1.78%
Intermediate precision	RSD≤2%	1.30%
Linearity And Range	Linear responses in calculated range Intercept=0 Coefficient of correlation=0.9991	Linear responses in calculated range i.e.15-50µg/ml Intercept=0 Coefficient of correlation = 0.9914
Ruggedness	RSD≤2%	0.52%
Robustness	RSD≤2%	1.74%

Table 4: Optical parameters:

Parameter	Data
λmax	282nm
E 1%,1 cm	193
Range	15-50µg/ml
Regression Equation	y =0.0205x
Correlation coefficient	0.9914
Slop	0.0205
Intercept	0

REFERENCES:

1. Promod K, Ansari SH, Ali J. Eugenol: A natural compound with versatile pharmacological actions. Nat prod commun 5(12); 1999-2006.

2. Pomod K, Ansari SH, Ali J. Development and validation of UV spectrophotometric method for the quantitative estimation of eugenol. Asian J. Pharma. 2013; 3(2); 58-61.

3. Snyder LR, Kirland JJ, Glajch JL. Practical HPLC method development.2^nd ed. willey inter science publication; p.686-713.

4. Sethi PD. Quantitative Analysis of Drug in pharmaceutical formulation.3^rd ed., CBS publication; p.50-64.

5. ICH-Guidelines Q2 (R1), Validation of Analytical procedures: Text and Methodology, Geneva, Switzerland: 2005.

Received on 06.11.2015 Accepted on 05.12.2015

Asian J. Pharm. Ana. 5(4): October- December, 2015; Page 178-180

DOI: 10.5958/2231-5675.2015.00028.9