Simple Extractive Colorimetric Determination of Tramadol Hydrochloride by Acid-Dye Complexation Methods in Solid Dosage Form

 

R. Kalaichelvi1* and E. Jayachandran2

1K.C.Reddy Institute of Pharmaceutical Sciences, Jangamguntla Palem, Medikonduru Mandal, Guntur-522 348, India 2S.C.S. College of Pharmacy, Harapanahalli-583131. Devanagari Dist, Karnataka

*Corresponding Author E-mail: rkselvi123@rediffmail.com

 

ABSTRACT:

Two simple and sensitive ion-pairing spectrophotometric methods have been described for the assay of tramadol hydrochloride in pure form and in pharmaceutical formulations. The developed methods involve formation of colored chloroform extractable ion-pair complexes of the drug with Bromocresol Green (BCG) and Bromothymol Blue (BTB) in acidic medium. The extracted complexes showed absorbance maxima at 430, 424 nm for BCG, BTB, respectively. Beer’s law is obeyed in the concentration ranges 5-40, 2-10 μg/mL with molar absorptivity of 7.2 x 103, 1.8 x 104, L mole-1 cm-1 for BCG and BTB, respectively. These methods have been successfully applied for the assay of drug in pharmaceutical formulations. No interference was observed from common pharmaceutical adjuvants. Results of analysis were validated statistically and through recovery studies.

 

KEYWORDS: Tramadol hydrochloride, Spectrophotometry, Validation

 

INTRODUCTION:

Tramadol is an orally administered non-steroidal anti-inflammatory drug1-2, which possesses good analgesic properties and good tolerability profile in a variety of painful conditions. The chemical name for tramadol hydrochloride is (±) cis-2-[(dimethylamino)methyl]-1-(3-methoxyphenyl)cyclohexanol hydrochloride. Literature survey reveals that reports are available for estimation of tramadol hydrochloride by Potentiometric3, Kinetic spectrophotometric4, thin layer chromatography and densitometry5, capillary isotachophoresis6, HPTLC7, UV spectrophotometric and HPLC8,9 in pharmaceutical formulations and HPLC10-13 methods in plasma and urine. The objective of the present study was to develop simple, precise, accurate and economic analytical methods with the better detection range for estimation of tramadol hydrochloride in bulk, pharmaceutical formulations.

 

Experimental procedures:

Instruments:

The spectrophotometric measurements were carried out using An Elico UV/Visible double beam spectrophotometer SL-164 with matched quartz cells of 10 mm path length.

 

Reagents:

All chemicals were of analytical reagent grade of E.Merck unless otherwise specified. Doubly distilled water was used to prepare all solutions. Freshly prepared solutions were always employed. Potassium hydrogen phthalate buffer solution of pH 4 was also prepared. 0.1% (w/v) BCG and BTB were prepared. Tablets containing 100 mg active material were kindly supplied from local pharmacy stores.

 

Analytical method development:

Method using BCG:

From the 100 µg mL-1 solution, 0.5, 1, 2, 3 and 4 mL was transferred to a series of separating funnels and 2 mL of pH-4 buffer was added to each and then 1mL of 0.1 % w/v BCG was added and shaken well and 10 mL of chloroform was added to each and shaken well and kept for few minutes. The chloroform layer was separated and treated with anhydrous sodium sulphate and the absorbance of the solution at 430 nm was measured against reagent blank. Final concentrations of analysed solutions were 5 µg mL-1 to 40 µg mL-1. The standard calibration plot was prepared to calculate the amount of the analyte drug in unknown samples.

 

Method using BTB:

From the 100 µg mL-1 solution, 0.2, 0.4, 0.6, 0.8 and 1 mL was transferred to a series of separating funnels and 2 mL of pH-4 buffer was added to each and then 1mL of 0.1 % w/v BTB was added and shaken well and 10 mL of chloroform was added to each and shaken well and kept for few minutes. The chloroform layer was separated and treated with anhydrous sodium sulphate and the absorbance of the solution at 424 nm was measured against reagent blank. Final concentrations of analysed solutions were 2 µg mL-1 to 10 µg mL-1. The standard calibration plot was prepared to calculate the amount of the analyte drug in unknown samples.

 

Estimation from formulations:

Tablets containing tramadol were successfully analyzed by the proposed methods. Twenty tablets of tramadol were accurately weighed and powdered. Tablet powder equivalent to 100 mg of tramadol was dissolved in 100 ml of distilled water and sonicated for 15 minutes, filtered and washed with distilled water, the filtrate and washings were combined and the final volume was made to 100 ml with distilled water. The solution was suitably diluted and analyzed as given under the assay procedure for bulk samples. The results are represented in Table 1. None of the excipients usually employed in the formulation of tablets interfered in the analysis of tramadol, by the proposed methods.

 

RESULTS AND DISCUSSION:

Spectral characteristics:

Absorption spectrum of the yellow drug-BCG and drug-BTB ion-pair complexes with their λmax at 430 nm, 424 nm shown in the Figure 1 and 2 respectively. The colorless blank is having practically negligible absorbance.

 

Figure 1. Absorption spectra of Tramadol Hydrochloride -BCG complex extracted into 10 mL chloroform:

 

Figure 2. Absorption spectra of Tramadol Hydrochloride -BTG complex extracted into 10 mL chloroform.

 

Optimization of variables:

Optimum conditions necessary for rapid and quantitative formation of colored ion-pair complexes with maximum stability and sensitivity were established by a number of preliminary experiments. Potassium hydrogen phthalate buffer was found to be suitable for these methods. Chloroform was preferred to other solvents (carbon tetrachloride, dichloromethane and ether) for these methods for its selective and quantitative extraction.

 

Optimum conditions were fixed by varying one parameter at a time while keeping other constant and observing its effect on the absorbance at 430 nm for BCG and 424 nm for BTB. The pH was studied by extracting the colored complex species at different absorbance was observed at the pH 4.0 and using 2 mL of buffer. A volume 0.1% (w/v) BCG and BTB was found to be optimal for complete complexation.

 

Analytical validation:

Accuracy:

Recovery studies were performed to judge the accuracy of the methods. Recovery studies were carried out by adding a known quantity of pure drug to pre-analyzed formulations and the proposed methods were followed. From the amount of drug found, percentage recovery was calculated. The results of analysis and recovery studies are given in Table 1.

 

Precision:

Repeatability was determined by using different levels of drug concentrations prepared from independent stock solution and analysed. Inter-day and intra-day variation and instrument variation were taken to determine intermediate precision of the proposed methods. Different levels of drug concentrations were prepared six different times in a day and studied for intra-day variation. Same protocol was followed for six different days to study inter-day variation. The relative standard deviation (in %) of the predicted concentrations from the regression equation was taken as precision (Table 3).

 

Linearity and range:

Beer’s law range, molar absorptivity, Sandell’s sensitivity, regression equation and correlation coefficient determined for the methods are given in Table 2. A linear relationship was obtained in the concentration range of 5 to 40 μg mL-1. for BCG and 2-10 μg mL-1 for BTB.

 

Detection limit (DL) and quantitation limit (QL):

The DL and QL of tramadol hydrochloride by the proposed methods were determined using calibration standards. DL and QL were calculated as 3.3r/S and 10r/S, respectively, where S is the slope of the calibration curve and r is the standard deviation of y-intercept of regression equation (Table 3).

 

Tablets analysis:

The proposed methods were applied to the determination of tramadol hydrochloride in commercial tablets. The


Table 1. Assay of tramadol hydrochloride in tablets.

Sample

(Tablet)

Labeled Amount

(mg/tab)

Amount Obtained (mg/tab)*

by proposed method

** % Recovery by the

proposed method

 

 

Method using BCG

Method using BTB

Method using BCG

Method using BTB

1

100

100.76

100.28

100.05

100.06

2

100

100.28

100.10

100.20

100.08

3

100

101.31

100.15

100.16

100.10

*Average of three determinations, ** After spiking the sample.

 

 

applicability of the proposed methods for the assay of tramadol hydrochloride in tablet formulation were examined by analyzing various formulations and the results obtained were tabulated in Table 1. Satisfactory results were obtained for drug and were in a good agreement with the label claims. The results were reproducible with low R.S.D. values. The average percent recoveries obtained were quantitative and indicating good accuracy of these methods. The results of analysis of the commercial tablets and the recovery study of drug suggested that there is no interference from any excipients present in tablets.

 

Table 2. Optical characteristics of proposed method

Parameters

BCG

BTB

λ max (nm)

430

424

Beer’s law limit (μg mL-1)

5 - 40

2 - 10

Sandell’s sensitivity (μg cm-2/0.001 absorbance unit)

2.4 × 10-5

6.05× 10-5

Molar absorptivity (L mol-1 cm-1)

7.2 x 103

1.8 x 104

Regression equation (Y = a + bc)

Slope (b)                                                                                                                                                      Intercept(a)

 

0.022

0.012

 

0.058

0.005

Correlation coefficient (r2)

0.999

0.999

 

Table 3. Summary of validation parameters

Parameters

BCG

BTB

Linearity and range (μg/ml)

5 - 40

2 -10

LOD (μg/ml)

1.0148

0.0356

LOQ (μg/ml)

3.3829

0.1188

Accuracy (% Recovery) ( n = 18)

100.14%

100.08%

%RSD

0.3088

0.2329

Precision (%RSD)

 

 

Intra-day (n = 6)

0.7711

0.7294

Inter-day ( n = 6)

0.5939

0.9354

Repeatability( %RSD) (n = 6)

0.3134

0.5175

 

 

CONCLUSION:

In summary, the proposed methods were simple, rapid, accurate, precise and inexpensive and can be used for routine analysis of tramadol in bulk and pharmaceutical formulations. The developed analytical methods will be useful for normal dissolution studies. The sample recoveries in all formulations were in good agreement with their respective label claims and thus suggested non-interference of formulations excipients in the estimation.

 

REFERENCES:

1.       Gutstein HG, Akil H. Opioid analgesics.In: Harrdman JG, Limbird LE, Gilman AG, editors. Goodman and Gillman’s The Pharmacological basis of Therapeutics. 10th ed. McGraw-Hill: London; 2001. p. 569-620.

2.       Rang HP, Dale MM, Ritter JM, Moor Rk, editors. Antiinflammatory Agents: Rang and Dale,s Pharmacology. 5th ed. Church Livingstone: London; 2003.p. 580.

3.       Ganjali M, Razavi T, Faridbod F, Riahi S, Norouzi P. Application of a New Tramadol Potentiometric Membrane Sensor as a Useful Device for Tramadol Hydrochloride Analysis in Pharmaceutical Formulation and Urine. Curr Pharm Analysis. 2009; 5: 28-33.

4.       Abdellatef HE. Kinetic spectrophotometric determination of tramadol hydrochloride in pharmaceutical formulation. J Pharm Biomed Anal, 2002; 29: 835-42.

5.       Krzek J, Starek M. Quality assessment for tramadol in pharmaceutical preparations with thin layer chromatography and densitometry. Biomed Chromatogr, 2004; 18: 589 – 59.

6.       Pospisilova M, Polasek M, Jokl V. Determination of tramadol in various dosage forms by capillary isotachophoresis. J Pharm Biomed Anal. 1998; 18: 446.

7.       Meyyanathan SN, Kumar P, Suresh B, Analysis of tramadol in pharmaceutical preparations by high performance thin layer chromatography. J Sep Sci. 2003; 26: 1359 – 1362.

8.       Küçük A, Farmaco KY. Determination of tramadol hydrochloride in ampoule dosage forms by using UV spectrophotometric and HPLC-DAD methods in methanol and water media. 2005; 60: 163-9.

9.       Kartinasari WF, Palupi T, Gunawan Indrayanto HPLC Determination and Validation of Tramadol Hydrochloride in Capsules. J Liq Chromatogr Relat Technol. 2005; 27: 737 – 744.

10.     Ceccato A, Chiap P, Hubert P, Crommen J. Automated determination of tramadol enantiomers in human plasma using solid-phase extraction in combination with chiral liquid chromatography. J Chromatogr B Biomed Sci Appl. 1997; 698 :161-70.

11.     Pedersen RS, Brřsen K and Nielsen F. Enantioselective HPLC method for quantitative determination of tramadol and O-desmethyltramadol in plasma and urine: Application to clinical studies. Chromatographia. 2003; 57: 5-6.

12.     Qu L, Feng S, Wu Y, Sichuan D, Bao XX, Ban YX. HPLC method for determination of tramadol hydrochloride in human plasma. 2003; 34: 574-5.

13.     Augustin C, Daniela M, Manuela C, Maria D, Camil V. Optimized HPLC method for tramadol and O-desmethyl tramadol determination in human plasma. J Biochem Biophys Methods. 2008; 70: 1304-1312.

 

 

 

 

 

Received on 25.03.2010       Modified on 24.04.2010

Accepted on 29.05.2010      © RJPT All right reserved

Research J. Pharm. and Tech.3 (4): Oct.-Dec.2010; Page 1096-1098