INTRODUCTION:

Simethicone, a silicon-based organic polymer, is widely found in drug formulations used in the treatment of conditions caused by excessive gas due to its anti-foaming properties. It is used as an active ingredient or excipient in different drug forms such as capsule, drop, suspension, chewable tablet and is preferred in the treatment of diseases such as heartburn, gastric ulcer, colic, sensitive bowel syndrome.^1-4

In literature studies, there are HPLC methods with UV or evaporative light scattering detectors (ELSD) for the determination of simethicone^4-7, but these methods are not found in raw material and finished product monographs. The absence of chromophore groups in the simethicone structure makes it difficult to work with the UV detector. The methods in the literature have been tried and the areas could not be taken as stated. Because it does not have a solubility in solvents such as water, alcohol and it does not have chromophore groups, it makes HPLC-UV detector studies difficult. Due to these features, evaporative light scattering detectors have been put into use for high-performance liquid chromatography for simethicone, which has working difficulties. An evaporative light scattering detector is commonly used for the analysis of compounds where UV detection might be a restriction and it can detect all compounds which are non-volatile and semi-volatile. However, since special detectors such as ELSD are not easily found in every sector and laboratory, it is necessary to add new methods to the literature for the determination of molecules such as simethicone.

In pharmacopeia research, there are analytical methods reported for many product forms such as tablets, capsules, oral suspension, chewable tablets which are containing simethicone in British (BP)⁸, US Pharmacopeias (USP)⁹. Almost all of the methods in these pharmacopeias are carried out by Fourier-transform infrared (FTIR) spectroscopy for the quantification or recognition of simethicone. Although this method is suitable for the raw material, the stability and robustness of the FTIR method are not as suitable as other analytical methods, given the effects of the placebo components for the finished product forms. Considering all these, a new gravimetric method was tried and validated to be an alternative to FTIR and HPLC methods. This method, which can be easily applied without the need for any derivatization and costly devices, is quite simple and economical.

The product we use in our study is an oral suspension produced by our company that contains aluminum hydroxide, magnesium hydroxide, and simethicone as active ingredients. Quantification of simethicone for this product had been studied and validated both by gravimetric method and FTIR spectroscopy specified in the finished product monographs. The results obtained by both methods were compared statistically.

MATERIAL AND METHODS:

Chemicals and reagents:

A high purity grade sample of SMT was purchased from USP and was supplied as raw material from Riocare India Pvt. Ltd. The suspension product had been improving by the World Medicine Pharmaceutical Industry and Trade Inc. (Istanbul, Turkey). Sodium hydroxide pellets, toluene, diethyl ether, hydrochloric acid and sodium sulphate anhydrous were purchased from Merck. HPLC grade water (0.05µc) produced by the Sartorius Stedim Biotech system.

Instrumentation:

A Shimadzu IRAffinity-1 FTIR spectrophotometer was used in the 400–4000 cm^-1 scan range. Each spectrum was automatically averaged over 20 scans obtained at a spectral resolution of 5 cm^-1. IR solution software was used for data processing and evaluation.

Preparation of solutions for FTIR method:

Place accurately weighed an amount of oral suspension, equivalent to 50mg of simethicone, to a suitable 120mL round bottom flask, add 40mL of 0.1N sodium hydroxide, and swirl to disperse. Add 25.0mL of toluene, close the bottle securely with a cap having an inert liner, and shake for 15 min on a reciprocating shaker (e.g., about 200 oscillations/min and a stroke of 38± 2mm). Place the mixture to a 125mL separator. Remove about 5 mL of the upper organic layer to a 15mL test tube containing 0.5g of anhydrous sodium sulfate. Cap the tube with a stopper having an inert liner, shake vigorously, and centrifuge the mixture until a clear supernatant is obtained.

In parallel, prepare the standard solution similarly. Dissolve 50mg of USP Polydimethylsiloxane RS in 25.0 mL of toluene, add 40mL of 0.1N sodium hydroxide, and add a volume of water equal to that of the specimen of oral suspension taken for the sample solution.

Determine solution absorption in 0.5-mm cells at absorption maximum wavelength of about 7.9μm with the help of an appropriate infrared spectrometer, using a reference solution for apparatus calibration.

Preparation of solutions for gravimetric method:

Shake well the vial with suspension and accurately weighed amount of homogeneous suspension, equivalent to 20mL of suspension, is quantitatively transferred to a separation funnel. Add 15mL of concentrated hydrochloric acid and mix carefully. Extract with ether (5 x 50mL portions), filter through sodium sulfate anhydrous into a preliminarily weighed round bottom flask. Evaporate filtered extract and dry the residue at 80°C for 2 hours.

Likewise, prepare the standard solution. Weigh accurately Simethicone equivalent to 500.0mg SMT, is quantitatively transferred to a separation funnel. Add 15 mL of concentrated hydrochloric acid and mix carefully. Extract with ether (5 x 50mL portions), filter through sodium sulfate anhydrous into a preliminarily weighed round bottom flask. Evaporate filtered extract and dry the residue at 80°C for 2 hours. Determine the weight of the obtained dried residue.

Determine the weight of the obtained dried residue.

Amount of Simethicone was estimated using following equation,

m× ρ × 100

g/100mL =-----------------------------------------

Where:

m: Weight of dried residue, g;

ρ: Density of suspension, g / mL;

а: Weighed amount of suspension, g;

RESULTS AND DISCUSSION:

For our suspension product, SMT quantification analysis was performed with both FTIR method and the gravimetric method. Validation parameters were studied for these methods following the ICH guidelines¹⁰. Common parameters were evaluated statistically. For this, t-test and F-test were applied to the results obtained.

The FTIR spectrum of the peak, which is obtained by separating from noise and other components and which is the identifier for simethicone at 1263 cm^-1, is shown in Fig.1. By making use of the absorbance value of the SMT at this point, the relevant calculations were made. In gravimetry, a calculation based on the difference of weighing taken before and after extraction was made and validation parameters were examined.

Fig.1: Infrared spectra of Simethicone

Linearity was studied in both methods. For the gravimetric method; A linearity graph was drawn between the average amount of simeticon weighed for analysis and the amount obtained at the end of the analysis. The method showed good linearity with a correlation coefficient of 0.999 (Fig.2).

Fig.2: Linearity graph of gravimetric method

The linearity of the FTIR method was confirmed between 0.42-3.17mg mL^-1 concentrations. The slope at 1263 cm^-1 was 0.2636 with the intercept value of -0.0050 (Fig.3).

Fig.3: Linearity graph of FTIR method

Accuracy was assessed by preparing recovery solutions for both methods in the concentration range of 50%, 100 and 150%. These were analyzed against placebo and blank solutions to ensure that there was no interference. Each recovery solution was prepared three times. The results of percentage recovery were within the accepted limits from 98.0% to 102.0% and % RSD was not more than 2.0% as shown in Table 1.

Table 1: Accuracy results of the method

% Recovery	FTIR Method	Gravimetric Method
50%	101.23	100.11
100%	99.71	101.21
150%	99.86	99.54
Mean	100.16±1.11	100.29±0.97
RSD	1.45	1.26

To confirm the precision of the methods, 6 different sample solutions were prepared by the same analyst on the same day and studied by two methods. The Simethicone content was determined by FTIR method with a mean result of 2.50±0.04 g/100 mL (RSD 1.35%), by the gravimetric method with a mean result of 2.53±0.03 g/100 mL (RSD 1.30%), in good agreement with the label claim of 2.5 g/100 mL.

Table 2: Precision results of the method

	Method Precision		Intermediate Precision
	FTIR Method	Gravimetric Method	FTIR Method	Gravimetric Method
Mean	2.50±0.04	2.53±0.03	2.51±0.04	2.51±0.02
SD	0.03	0.03	0.04	0.02
RSD	1.35	1.30	1.61	0.94

Similarly, the intermediate-precision results performed by different analysts on different days are similar and their values are given in Table 2. Besides, the precision RSD results also meet the acceptance criteria of the ICH guidelines.

F- and t-test results:

Chemometric methods were used to examine the relationship between recovery, precision and intermediate precision results obtained by two methods, one spectrophotometric and the other gravimetric. First of all, ANOVA F-test was applied to the results obtained in each parameter. The F-test variance analysis was applied between the gravimetric results corresponding to the FTIR results and it was observed that the F values obtained at the α = 0.05 significance level were greater than 1 and thus there was an F distribution between the results obtained. According to the relationship between F and F critical, t-test was applied. The F-test results obtained for the recovery parameter, as seen in Table 3, F ≥ 1 and F < F critical two-tail were found. In this case, the " t-Test: two samples assuming equal variances" test was performed and again according to the results in Table 3 it was found that | t Stat | ≤ t is Critical two-tailed, it was emphasized that there is no difference between the results obtained by the two methods and therefore the recovery of the methods is equivalent.

Table 3: F- and t- test results of accuracy

F-test two sample for variances			t-Test: Two Samples Assuming Equal Variances
	FTIR results	Gravimetric results		FTIR results	Gravimetric results
Mean	100.16	100.29	Mean	100.16	100.29
Variance	2.10	1.60	Variance	2.10	1.60
Observation	9	9	Observation	9	9
df	8	8	Cumulative Variance	1.85
F	1.31		Predicted Mean Difference	0
P(F<=f) one-tailed	0.36		df	16
F Critical two-tailed	3.44		t Stat	-0.20
			P(T<=t) one-tailed	0.42
			t Critical one-tailed	1.34
			P(T<=t) two-tailed	0.84
			t Critical two-tailed	1.75

Table 4: F- and t- test results of precision

F-test two sample for variances			t-Test: Two Samples Assuming Equal Variances
	FTIR results	Gravimetric results		FTIR results	Gravimetric results
Mean	100.08	101.03	Mean	100.08	101.03
Variance	1.82	1.73	Variance	1.82	1.73
Observation	6	6	Observation	6	6
df	5	5	Cumulative Variance	1.77
F	1.05		Predicted Mean Difference	0
P(F<=f) one-tailed	0.48		df	10
F Critical two-tailed	5.05		t Stat	-1.23
			P(T<=t) one-tailed	0.12
			t Critical one-tailed	1.37
			P(T<=t) two-tailed	0.25
			t Critical two-tailed	1.81

Table 5: F- and t- test results of intermediate precision

F-test two sample for variances			t-Test: Two Samples Assuming Equal Variances
	FTIR results	Gravimetric results		FTIR results	Gravimetric results
Mean	100.42	100.33	Mean	100.42	100.33
Variance	2.62	0.90	Variance	2.62	0.90
Observation	6	6	Observation	6	6
df	5	5	Cumulative Variance	1.76
F	2.92		Predicted Mean Difference	0
P(F<=f) one-tailed	0.13		df	10
F Critical two-tailed	5.05		t Stat	0.12
			P(T<=t) one-tailed	0.45
			t Critical one-tailed	1.37
			P(T<=t) two-tailed	0.90
			t Critical two-tailed	1.81

Similar calculations were made for the precision and intermediate precision parameters, and as with accuracy, it was observed that there was no difference between the results obtained by the two methods. F and t-test results obtained for these parameters are shown in Table 4 and Table 5.

CONCLUSION:

A new gravimetric method was optimized for the quantitative determination of simethicone in the oral suspension product which has simethicone, aluminum hydroxide and magnesium hydroxide as active pharmaceutical ingredients in the formulation. Quite simply, economical, time-saving this new method has been validated according to regulatory guidelines and compared with the FTIR procedure recommended by the US Pharmacopeia. The statistical F-test and t-test were applied to check the results obtained by the two analytical methods. No significant differences were observed. In the results obtained, it was seen that there were no significant differences between the methods at 90% confidence level, since | t Stat | ≤ t was critical two-tailed.

After extraction of simethicone from a dense matrix such as citric acid, peppermint oil, sodium methyl para-hydroxybenzoate, sodium propyl para-hydroxybenzoate, saccharin sodium, sorbitol 70% solution and mannitol with a simple method and quantification by an absolute method, a different product form was tried. This time, the gravimetric method was studied on the oral drop form, where only simethicone was used as a drug active ingredient in a dense placebo content. The results of the accuracy and precision parameters performed met the ICH criteria.

This proposed gravimetric method is relatively easy and economical to the analytical methods mentioned earlier. Besides being reliable and very fast, it is suitable for quality control analysis. Since it is an absolute method, it does not require linearity and therefore a calibration chart. With all these proofs and reasons, it is stated that this validated gravimetric analysis can be used in pharmaceutical products containing simethicone.

ACKNOWLEDGEMENT:

This work was supported by the World Medicine Pharmaceutical Industry and Trade Inc. The authors are thankful to the company, for providing necessary instrumental facilities and chemicals to carry out the research work.

CONFLICT OF INTEREST:

The authors declare no conflict of interest.

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Received on 22.06.2020 Revised on 06.08.2020

Asian J. Pharm. Ana. 2021; 11(1):17-21.

DOI: 10.5958/2231-5675.2021.00003.X