INTRODUCTION:

MF is a corticosteroid (glucocorticoid) having an anti-inflammatory activity. The exact mechanism of MF on allergic rhinitis has not been recognized. Corticosteroids showed a wide range of effects on various cell types such as mast cells, eosinoplills, neutrophills, macrophages, and lymphocytes^1,2. The chemical name of MF is (11β, 16α)-9, 21-dichloro-11hydroxy-16-methyl-3,20-dioxopregna-1,4-dien17-yl 2-furoate. It is white or off-white crystalline powder, soluble in acetone, dichloromethane, and slightly soluble in ethanol. Practically insoluble in water^3,4,5. It is official in Indian and British pharmacopoeia.

Literature survey indicate that many analytical methods are reported for determination of MF using HPLC, HPTLC, RP-HPLC, in different solvents but no spectroscopic method was available for estimation of MF in bulk and pharmaceutical dosage form (cream) using dichloromethane as solvent^6,7.

MATERIALS AND METHODS:

Reagents and chemicals:

MF with 99.56% purity as at work standard was obtained as gift sample from ZCL CHEMICALS Limited, Ankleshwar, India. Dichloromethane was purchased from Fisher Scientific, Mumbai. Ethanol, acetone and octanol was purchased from Merck. Distilled water and all other chemicals used were analytical grade, cream formulation was purchased from local pharmacy. A double-beam Shimadzu-1800 UV- Visible spectrophotometer, with spectral bandwidth of 2nm, along with of ±0.5nm wavelength accuracy for all the weighing throughout the research work, weighing balance Shimadzu AUX220 was used.

Stock Solution (Standard) of Mometasone Furoate:

Stock solution (1000µg/ml) of MF was prepared in dichloromethane.

Spectral characteristics of Mometasone Furoate:

Solution of Mometasone Furoate (5µg/ml) was prepared by relevant dilution of standard solution and scanned at 200nm to 400nm.

Preparation of Calibration curve:

Appropriate dilutions of the stock solution of MF was done with dichloromethane to yield calibration standard solutions with concentrations 1, 2, 4, 6, 8, and 10μg/ml. All the solutions scanned in rang of 200-400nm for determining the absorption maxima. The λ_max of MF was recorded 246.0nm. Beer’s lambert’s range for MF was selected and standard curve of the drug was plotted^8,9.

Method for the determination Mometasone Furoate in cream:

Accurately weighed 1g of sample equivalent to 1mg of MF was dissolved in volumetric flask (100ml) with dichloromethane. 4ml of above solution taken and diluted up to 10ml in volumetric flask, the solution was filtered using Whatman filter paper¹⁰. The resultant solution was of 4μg/ml.

Method Validation³:

The method was validated with respect to LOD, Accuracy, linearity, LOQ and Precision as per the ICH Q2 (R1) guidelines.

Linearity:

The standard curve was obtained with six different concentrations of reference standard solution in range of 1-10µg/ml for the UV-Visible spectroscopic method. All the values were performed in triplicate. The linearity was evaluated by linear regression analysis, which be situated to calculate the coefficient of correlation and slope¹¹.

Precision:

Precision was checked by taking five replicates of 4µg/ml for repeatability. For inter-day and intra-day (2, 4, 6µg/ml) where analysed for three times for consecutive three days and three thrice a day respectively. For Intermediate Precision: (Ruggedness) six repeats of 4µg/ml were analysed by different analysts. The data were analysed at 246nm. The percentage Relative Standard Deviation (% R.S.D.) of the analytical response were calculated^12,13,14.

Accuracy:

Accuracy was performed at three levels (75%, 100% and 125%) by taking 4µg/ml as 100%. To determine accuracy standard solution was added^16,17.

Limit of detection (LOD) and Limit of quantification (LOQ)¹⁸:

The LOD and LOQ were calculated using following formulae;

LOD = 3 × (standard deviation of y-intercept/slope of the calibration curve)

LOQ = 10 × (standard deviation of y-intercept/slope of the calibration curve)

Where,

SD = standard deviation of response and

S = average of the slope of the calibration curve

RESULT AND DISCUSSION:

Solubility:

Table 1: Results showing solubility of Mometasone Furoate in different solvents

S. No.	Solvent	MF
1	Water	Practically insoluble
2	Ethanol	Slightly soluble
3	Dichloromethane	Very soluble
4	Acetone	Soluble
5	Octanol	Soluble

Identification Test:

After 15 minutes of addition of sulphuric acid yellow colour was developed, which disappeared after addition of 10ml water and no fluorescence was observed under UV^19,20.

The observation states that the drug sample of Mometasone Furoate

IR Spectroscopy:

IR spectroscopy was performed using solid API sample and IR spectrum was obtained. It was then compared with standard IR spectrum of Mometasone Furoate and different groups present were identified on basis of wave number^21,22.

Figure 1: IR spectrum of Mometasone Furoate

Figure 2: Standard IR Spectra (Source: Bio-Rad Laboratories, Inc. Spectrabase)²³

Table 2: Inference of IR spectrum

Bond

Frequency Range

(cm^-1)

Peak observed

(cm^-1)

C-------H (Alkanes)

2850-2970

1340-1470

2940.68

1465.68/1392

C-------H (Aromatic Gr.)

690-900

785.15

718

885

C======O(Ketone )

1690-1760

1723

C--------O(Alcoholic)

1050-1300

1075.47

Selection of Wavelength Maximum:

Spectral features of MF Solution (5µg/ml) and MF was primed by applicable dilution of standard working solution. The solution was scanned in the spectrum mode from 200 nm to 400 nm²⁴.

Figure 3: Graph showing wavelength maximum for Mometasone Furoate

The wavelength maximum (λ max) for Mometasone Furoate was found to be 246 nm. The cut-off value of dichloromethane for UV analysis is 232nm.

Linearity:

Linearity for different concentration of drug was performed and r²value was noticed to be 0.9993 for Mometasone Furoate.

The regression equation was found to be: y = 0.0857x - 0.0017

Table 3: Preparation of Linearity Solution

S. No.	Stock soln. to be taken in ml (100ppm)	Final volume (ml)	Final Concentration in µg/ml
1	1	100	1
2	2	100	2
3	4	100	4
4	6	100	6
5	8	100	8
6	10	100	10

The calibration curve of Mometasone Furoate was found to be linear at 1-10 µg/ml. The graph gave a regression coefficient of 0.9993.

Figure 3: Calibration curve of Mometasone Furoate

Table 4: Linearity data of Mometasone Furoate

Concentration (µg/ml)	Absorbance *	SD*	Percent RSD
1	0.085	0.0005	0.58
2	0.170	0.0015	0.88
4	0.338	0.0005	0.12
6	0.505	0.0026	0.51
8	0.698	0.001	0.14
10	0.849	0.0005	0.05

*Average of 3 replicates

Table 5: Regression Analysis Data for calibration Curves

Parameters(units)	MF
Linearity range (µg/mL)	1-10 µg/ml
Regression Coefficient (R²)	0.9993
Slope	0.0857
Intercept	-0.0017

Precision:

Repeatability:

Precision was performed and % RSD was observed as 1.69.

Table 6: Repeatability data for Mometasone Furoate

Conc. (µg/ml)	Sample No.	Absorbance*	Average Abs.	SD
4	1	0.358	0.358	±0.002
		0.356
		0.360
	2	0.344	0.342	±0.0032
		0.339
		0.345
	3	0.348	0.344	±0.002
		0.346
		0.340
	4	0.345	0.345	±0.0025
		0.348
		0.343
	5	0.355	0.353	±0.0062
		0.358
		0.346
AVERAGE ABSORBANCE			0.3484
AVERAGE SD			±0.00318
RSD			0.91%

*Average of 3 replicates

Intra-Day:

The %RSD was found to be >1% for intra-day.

Table 7: Intra-Day Precision Data for Mometasone Furoate

Drug	Conc. (µg/ml)	Absorbance*	SD	PERCENT RSD
MF	2	0.170	±0.0011	0.64
	4	0.336	±0.0019	0.56
	6	0.507	±0.0042	0.82

*Average of 3 replicates

The method precision gave results obtained within 1% RSD suggesting the method is precise.

Inter-Day:

The % RSD was instituted to be >2% for inter-day.

Table 8: Inter-Day precision data for Mometasone Furoate

Drug	Conc. (µg/ml)	Absorbance*	SD	PERCENT RSD
MF	2	0.165	0.0021	1.27
	4	0.345	0.004	1.15
	6	0.502	0.0035	0.69

*Average of 3 replicates

The method precision gave results obtained within 2% RSD suggesting manner is precise.

Intermediate Precision: (Ruggedness):

Table 9: Results for Ruggedness

S. No.	Conc.	Mometasone Furoate
S. No.	Conc.	Analyst I	Analyst II
1	4 mg/ml	0.345	0.336
2		0.354	0.332
3		0.339	0.341
4		0.341	0.335
5		0.345	0.331
6		0.351	0.342
Mean ± SD		0.3458±0.0057	0.3361±0.0045
RSD		1.64%	1.33%

a: Mean of six determinations

b: Ruggedness studies was conceded out using altered analysts.

The results for the evaluation of gave RSD beneath 2% which suggests that the method is rugged to changes²⁵.

Accuracy:

Accuracy was performed and %recovery was originated to be 99.14% to 100.31% for Mometasone Furoate.

Table 11: Result of Assay

Drug	Label Claim (μg/ml)	Concentration Found (μg/ml)
MOMETASONE FUROATE	4	4.15
		3.97
		3.99
		4.00
		4.09
AVERAGE CONCENTRATION±SD		4.04±0.07681
%RSD		1.9
%ASSAY		101.0

Table 12: Summary of Validation Parameters for Mometasone Furoate

S. No.	VALIDATION PARAMETER	MOMETASONE FUROATE
1	LINEARITY
	Regression equation	y = 0.0857x - 0.0017
	Regression coefficient	0.9993
2	RANGE (µg/ml)	1 - 10
3	ACCURACY (Percent recovery)	99.14 to 100.31
4	PRECISION (Percent RSD)
	Repeatability	0.91
	Intra-day	0.56 - 0.82
	Inter-day	0.69 – 1.27
	RUGGEDNESS	1.33 – 1.64
5	LOD (µg/ml)	0.031
6	LOQ (µg/ml)	0.106
7	PERCENT ASSAY	101.00

CONCLUSION:

Thus, it can be determined that techniques in present research work were simple, sensitive and reproducible when checked for validation parameters like accuracy, precision and ruggedness for monotonous purpose of Mometasone Furoate in bulk along with pharmaceuticals (cream).

ACKNOWLEDGEMENT:

The authors express heartfelt and special thanks to ZCL Chemicals Limited, Ankleshwar, India, who provided the drug. Authors as well obliged to Material Research Centre (MRC), Malviya National Institute of Technology, Jaipur for their plentiful support in experimental and analytical work.

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Received on 17.11.2019 Modified on 27.12.2019

Research J. Pharm. and Tech. 2020; 13(7): 3093-3097.

DOI: 10.5958/0974-360X.2020.00548.X

Recovery Level	Initial Sample Conc.(μg/ml)	Conc. of Standard Drug Added (μg/ml)	Total Conc. (μg/ml)	Absorbance*	Amount of Drug Recovered (μg/ml)	%Recovery (In %)
75%	4	3	7	0.597	7.04	100.51
				0.595	7.02	100.23
				0.589	6.94	99.14
100%	4	4	8	0.685	8.08	101.00
				0.675	7.96	99.52
				0.688	8.11	101.31
125%	4	5	9	0.770	9.08	100.82
				0.762	8.99	99.97
				0.768	9.06	100.66