INTRODUCTION:

Eletriptan hydrobromide is a second generation triptan drug and it intended for treatment of migraine headache^1,2. It is used as an abortive medication, and blocks migraine attack which is already in progress. EHB chemically known to be 3-[(-1-methylpyrrolidin-2-yl) methyl]-5-(2-phenylsulfonylethyl)-1H-indole. It is selective at 5-HT1B/1D receptor agonist; thought to be due to the agonist effects at the 5-HT1B/1D receptors located on intracranial blood vessels (including arteriovenous anastomoses) and sensory nerves of the trigeminal system that results in cranial vessel constriction and inhibition of proinflammatory neuropeptide release³.

Literature survey reveals few chromatographic methods to determine the EHB by HPLC^4-9, TLC¹⁰and Liquid Chromatography coupled with tandem mass spectroscopy¹³,simultaneous determination of EHB with other anti-migraine drugs^11,12, determination of EHB in plasma using Forced degradation studies , development of stability indicating method¹⁴, Spectrophotometric method^15,16,determination of Process related impurities in Eletriptan using UPLC method¹⁷, TLC-Densitometric method¹⁸, Fluorimetric and Colorimetric method¹⁹, Capillary Electrophoresis²⁰, and Thermal diffractometric studies²¹.UV^22-24methods, RP-HPLC²⁵,and a spectrophotometric method²⁶ were also reported

The analytical useful functional groups in EHB have not been fully exploited for designing suitable visible spectrophotometric methods and so still offer a scope to develop more visible spectrophotometric methods with better sensitivity, precision and accuracy. The author has made some attempts in this direction and succeeded in developing two methods for determination and validation of EHB in Pharmaceutical dosage forms.

A reported UV spectrophotometric method has been adopted for the determination of EHB in pharmaceutical formulations (tablets), which has been made use of reference method to compare the results obtained by the proposed visible spectrophotometric methods.

Fig.1.Structure of EHB

Experimental

Materials and Methods

Instruments Used

A Schimadzu UV-Visible spectrophotometer 1801 with 1 cm matched quartz cells was used for all spectral and absorbance measurements. A Systronics digital pH meter 361 was used for pH measurements.

Procedure of Assay of EHB in formulations

An accurately weighed amount of formulation (tablets) equivalent to 100 mg of drug was dissolved in 20 ml of distilled water, shaken well and filtered. The filtrate was further diluted to 100 ml with distilled water to get 1 mg/ml solution of drug in formulations.

One ml of this solution was furthered diluted to 25 ml to get 40 µg/ml solution. The absorbance of the solution was determined λ_max223 nm (Fig. 2). The quantity of the drug was computed from the Beer’s law plot (Fig. 3) of the standard drug in distilled water.

Fig.2. Absorption Spectra of EHB in Methanol (UV Reference method)

Fig 3. Beer’s Law plot of EHB in methanol (UV Reference method)

Preparation of standard Drug solution

The stock solution (1 mg/ml) of Eletriptan Hydrobromide (EHB) was prepared by dissolving 100 mg of it in 100 ml of millipore-distilled water. A portion of this stock solution was diluted stepwise with the distilled water to obtain the working standard EHB solution of concentrations 24 µg/ml and 60µg/ml for the proposed methods.

Recommended Procedures:

After systematic and detailed study of the various parameters involved like effect of volume of KMnO₄ required for oxidation, time and temperature for oxidation, volume of FGFCF solution, Effect of volume of 1.0M Na₂SO₄ solution, Keeping time after addition Na₂SO₄ solution, order of addition of reagents on color development and stability period after final dilution, the following procedure was recommended for the determination of EHB in bulk samples for method-A. Similarly based on the study of the various parameters involved like effect of volume of AM solution and conc.H₂SO_4,the order of addition of reagents on colour development, temperature, shaking time, solvent for final dilution and stability period after final dilution, the following procedure was recommended for the determination of EHB in bulk samples for method-B

Method-A

Into a series of 25 ml tubes containing aliquots of standard EHB solution (0.1-0.6 ml, 24 µg/ml), 0.5 ml of KMnO₄ solution was added and the total volume in each tube was brought to 5 ml with distilled water and kept aside for 10 min at laboratory temperature. Then 4.0 ml each of FG FCF solution and sodium sulfate solution were added successively and kept aside for 5 min. The volume was made up to the mark with distilled water. The absorbance was measured at λ_max 622 nm against distilled water. The decrease in absorbance corresponding to consumed permanganate and in turn the drug concentration was obtained by subtracting the decrease in absorbance of the test solution (dye – test) from that of the blank solution (dye – blank).(Fig.4).The amount of EHB in sample solution was obtained from the Beers-Lambert’s plot Fig.5.

Method-B

Aliquots of standard EHB solution (1.0 – 6.0 ml, 60 µg/ml) were transferred into a series of 10 ml calibrated tubes. Then 1.0ml of 2% ammonium molybdate and 4 ml of conc. sulphuric acid were added to each tube and the contents were heated for 20 min. in a boiling water bath. After cooling, the volume was made up to 10 ml with ethanol. The resulting absorbance of the green colour was measured at λ_max 641nm against a reagent blank (Fig.6). The amount of EHB in sample solution was obtained from the Beers-Lambert’s plot Fig.7.

Table 1: Optical and Regression characteristics, precision and accuracy of the proposed methods for EHB

S.No	Parameter	Method-A	Method-B
1	Wave length λ_max (nm)	622	641
2	Beer’s law limits (µg ml^-1)	4-24	10-60
3	Detection limits (µg ml^‑1)	0.0382	3.2909
4	Molar absorptivity (1 mole cm^-1)	6.8549 x 10⁵	1.2115 x 10⁵
5	Sandell’s sensitivity (µg cm^-2 / 0.001 absorbance unit)	1.3738 x 10^-2	1.3738 x 10^-2
6	Regression equation (Y = a + bC) Slope (b)	0.0588	0.0124
7	Standard deviation of slope (S_b)	8.2103 x 10^-4	3.4928 x 10^-4
8	Intercept (a)	-0.0111	0.0081
9	Standard deviation of intercept (S_a)	1.2789 x 10^-2	1.3602 x 10^-2
10	Standard error of estimation (S_e)	1.3738 x 10^-2	1.4611 x 10^-2
11	Correlation coefficient (r²)	0.9992	0.9968
12	Relative standard deviation (%)*	3.8593	0.9117
13	% Range of error (Confidence Limits) 0.05 level*	4.0507	0.9570
14	% Range of error (Confidence Limits)0.01 level	6.3526	1.5008
15	% Error in bulk samples**	0.454	0.15

Table-2: Assay and recovery of EHB in Pharmaceutical Formulations

Sample	Amount taken (mg)	Amount found by proposed methods		Reference Methods	Percentage recovery by proposed methods
Sample	Amount taken (mg)	Method-A	Method-B	Reference Methods	Method-A	Method-B
Tablet I	40	39.82 ±0.020 F=1.56 t=0.59	39.863 ±0.012 F=1.77 t=1.22	39.92 ±0.016	99.367 ±0.173	99.101 ±0.306
Tablet II	40	39.70 ±0.027 F=1.075 t=0.25	39.801 ±0.024 F=1.36 t=1.25	39.88 ±0.028	99.379 ±0.324	99.342 ±0.245

*Average ± standard deviation of six determinations; the t- and F- values refer to comparison of the proposed method with the reference method. Theoretical values at 95% confidence limit t=2.57, F=5.05.

**After adding 2 different amounts of the pure labeled to the pharmaceutical formulations, each value is an average of 3 determinations

Chemistry of the coloured species in the present investigation:

Method-A

EHB exhibits reducing property due to the presence of functional moieties vulnerable to oxidation selectively with oxidizing agents such as KMnO₄in method-A under controlled experimental conditions. When treated with known excess of oxidant, EHB undergoes oxidation, giving products of oxidation besides unreacted oxidant. It is possible to estimate the drug content colourimetrically, which is equivalent to reduced form of the oxidant formed. The unreacted oxidant can be estimated colourimatrically either by decrease in the intensity of dye colour due to disruption of chromophoric centers in the dye FGFCF. The nature of coloured species obtained with EHBis represented in the scheme-1

Scheme 1

Step –I:

EHB+ KMnO₄ oxidation products + KMnO₄ + Mn(II)

(unreacted)

Step – 2:

KMnO₄ + FG FCF FG FCF + Reacted FGFCF

(unreacted)

(coloured)

unreacted

Colourless mixture of unreacted compounds due to rupture of conjugate system (Reproducible but not stoichiometric as several alternate pathways possible)

Method-B

The tetrahedral anion MoO₄^2- in aqueous medium which is not a strongly oxidized form, on acidification (with conc. H₂SO₄) exists as isopolyanionic species as a result of polymerization and condensation reaction which is having oxidizing property Mo₆octahedral as exemplified by Mo₇O₂₄^6- and Mo₈O₂₆^4-. The acid hydrolysis products of paraxetine (catechol unit and formaldehyde) probably effect the reduction of 1, 2 or 3 oxygen atoms from exemplified molybdate, thereby producing one or more of the possible reduced species which has a characteristic intense colour.The reactions are described in the scheme 2

Scheme 2

RESULTS AND DISCUSSION:

Optimum operating conditions used in the procedure were established adopting variation of one variable at a time (OVAT) method. The optical characteristics such as Beer’s law limit, Sandell’s sensitivity, molar absorptivity, percent relative standard deviation, (calculated from the six measurements, Regression characteristics like standard deviation of slope (S_b), standard deviation of intercept (Sa), standard error of estimation (S_e) and % range of error (0.05 and 0.01confidence limits) were calculated and the results are summarized in Table-1.

Commercial formulations containing EHB were successfully analyzed by the proposed methods. The values obtained by the proposed and reference methods for formulations were compared statistically by the t-test and F-test and found not to differ significantly. As an additional demonstration of accuracy, recovery experiments were performed by adding a fixed amount of the drug to the pure analyzed formulations at three different concentration levels. These results are summarized in Table-2.

The proposed methods for EHB determination have many advantages over other analytical methods due to its rapidity, lower cost and environmental safety. Unlike HPLC, LC procedures, the instrument is simple and is not costly. Economically, all the analytical reagents are inexpensive and available in any analytical laboratory. The results presented above indicate that the proposed methods have good sensitivity, selectivity, precision and accuracy. Results of analysis in bulk form and in formulations reveal that the proposed methods are suitable for the estimation of EHB, as impurities and excipients present in them cause no interference virtually.

Fig 4.Absorption spectra of EHB: KMnO₄/FGFCF

Fig. 5 Beer’s plot of EHB:KMnO₄/FGFCF

Fig 6.Absorption spectra of EHB: AM/H₂SO₄

Fig.7. Beer’s plot of EHB: AM/H₂SO₄

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Received on 08.07.2020 Modified on 19.07.2020

Research J. Science and Tech. 2020; 12(3):195-200.

DOI: 10.5958/2349-2988.2020.00027.3