Original article

Derivative ultraviolet spectrophotometric determinationof dexchlorpheniramine maleate in tablets in presence of coloring agents

Nilton S. Viana Jr., Ligia M. Moreira-Campos, Cristina D. Vianna-Soares *

Faculdade de Farmácia, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Belo Horizonte, MG 31270-910, Brazil

Received 28 February 2005; received in revised form 16 August 2005; accepted 25 August 2005

Available online 13 October 2005

Abstract

Formulation excipients can frequently affect the drug analysis in pharmaceuticals yielding background interference by ultraviolet spectro-photometry. Sample separation procedures to diminish such interferences are usually recommended as sample pre-treatment, however it canbe difficult to eliminate them and they can still persist. In addition, these procedures can be time consuming and laborious to perform.Excipients, like dyeing agents can also be present in a formulation and yield color to drug solution. This work reports the successful devel-opment of a derivative ultraviolet spectrophotometry for dexchlorpheniramine maleate (DPM) determination in solid dosage forms, in spite ofthe color imparted to tablets solution. Standard curves obtained by second order derivative ultraviolet spectrophotometry showed linearitywith a correlation coefficient of 0.9999 in the concentration range of 9.75–32.5 µg ml−1 DPM in 0.1 mol l−1 sulfuric acid, using zero-peak (ZP)and peak–peak (PP) methods. The average relative standard deviation range was between 0.26% and 1.08% and 0.18% and 0.63% for ZP andPP methods, respectively. Application of the method in tablet samples resulted in coefficients of variation in the range of 0.83–1.40%, and0.63–0.83% for ZP and PP methods, respectively. Recovery test percentage values obtained were between 96.95% and 105.61% for the testedtablet samples.© 2005 Elsevier SAS. All rights reserved.

Keywords: Dexchlorpheniramine; Derivative ultraviolet spectrophotometry; Pharmaceutical preparations; Quality control

1. Introduction

The histamine (H1) receptor antagonists are representedby the anti-allergic, anti-motion sickness and anti-itch drugs.They are represented by several groups of drugs, such as, etha-nolamine, ethylenediamine, alkylamine, piperazine, phenothi-azine and piperidine derivatives. First generation drugs, devel-oped around 1940, such as, pyrilamine maleate (anethylenediamine salt) can both stimulate and depress the CNS[1]. Drugs, which pass easily through the blood–brain barrier(BBB), for instance, diphenhydramine (an ethanolamine) aremore prone to cause sedation, a common adverse reaction.On the other hand, second generation drugs, introduced inthe 1980’s (e.g. loratadine, acrivastine) are relatively largeand less lipophilic molecules, cross the BBB less easily, andact more selectively at the H1 receptors. They produce a mod-erate degree of sedation and anticholinergic effects com-

pared to first generation antihistamines. Third generationdrugs (as fexofenadine, desloratadine, and levocetirizine), areclaimed to be free of sedation. One of the most potent anti-allergic drugs of the alkylamines group is the prototype, chlo-rpheniramine (first generation agent). It does not show anti-emetic effects and gastrointestinal effects are less intense.However, it is prone to produce moderate drowsiness and driv-ing ability impairment immediately after oral administration.After 8 h, no significant difference from placebo was found[2]. Dexchlorpheniramine (DPH), a propylamine used in thesalt form of maleate (DPM, Fig. 1) is a dextro isomer of chlo-rpheniramine and is twice more potent than the racemic mix-

* Corresponding author. Tel./fax: +55 31 3499 6975.E-mail address: cviannas@ufmg.br (C.D. Vianna-Soares). Fig. 1. DPM chemical structure (390.86 g mol−1, pka 9.0).

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0014-827X/$ - see front matter © 2005 Elsevier SAS. All rights reserved.doi:10.1016/j.farmac.2005.08.009

ture. These drugs find their greatest use in the symptomatictreatment of allergic rhinitis [3,4]. They are useful in the acuterather than chronic form of urticaria (hives). In mild andchronic allergy, the dosage of the drug must be increasedbecause of tolerance development. It is also a common ingre-dient in cold medicines. Several pharmaceutical dosage formsof DPH alone (most commonly, plain or coated tablets,2–6 mg per dose unit or oral solutions, 0.4–2 mg per 5 ml) orin association are found in the market. The associations nor-mally contain active ingredients such as betametasone, phe-nylephrine hydrochloride, pseudoephedrine hydrochloride,haloperidol, pyridoxine, guaiphenesine, noscapine hydrochlo-ride or propantheline bromide. In Brazil, the reference prod-uct, Polaramine®, plain or coated tablets and syrup are manu-factured by Schering Plough. Several other companiesmanufacture the pharmaceutical as similar or generic prod-ucts [5,6].

According to the last editions of the American pharma-copeia (USP28) and of the British pharmacopoeia (BP2003)non-aqueous titrimetric methods with visual [7] or potentio-metric [8] endpoints are recommended for determination ofdexchlorpheniramine maleate (DPM) in raw material, respec-tively. The use of ultraviolet spectrophotometry at 264 or265 nm for DCM determination in syrup (oral solution) ortablets has been recommended by USP 28 [7]. However, themethod is applied only after several alkaline and/or acidicextractions of the samples with ether or hexane, and back-extractions of the aqueous portions with the same organicsolvent. British pharmacopoeia [8] recommends gas chroma-tography for DPM determination in oral solution. These meth-ods are either very laborious, time consuming, or expensive.In addition, they deal with easily flammable and nocuousorganic solvents.

Derivative ultraviolet spectrophotometry has successfullybeen applied to drugs alone or in association. This techniqueis an alternative method to determine drugs with low specificabsorptivity, substances under the influence of increased back-ground absorption, or drugs in association, wherein overlapsand absorption addition occur. First order (D1) derivative ultra-violet spectrophotometry [9–13] or first order differentialpulse polarography [14] has been reported in the literature todetermine DPM in presence of drug mixtures or in pharma-ceutical preparations. Nevertheless, tablets and syrups maycontain dyes, which impart color to solution and causetroubles even in the first order derivative spectral analysis.

This work reports a feasible derivative spectrophotomet-ric technique for the evaluation of DPM in tablets pharma-ceutical preparations using the approach of the second orderderivative (D2), zero-peak (ZP) and peak–peak (PP) methodsin the presence of coloring agents.

2. Experimental

2.1. Chemicals and solutions

Chemical reference substance (CRS) was of pharmaceu-tical grade and reagents were of analytical grade. DPM stan-

dard, used as received, was obtained from a local pharmaceu-tical company. Solutions were prepared in 0.1 mol l−1 sulfuricacid. DPH test and reference tablet samples (label claimed2 mg of DPM per unit dose) were purchased in local pharma-cies from two different pharmaceutical companies (A, B).Sample A placebo was prepared and furnished by the manu-facturer.

2.2. Apparatus

An HP8453 double-beam ultraviolet–visible spectropho-tometer equipped with HP laser printer and electronic deriva-tive mode was employed with 1 cm quartz cuvettes. Deriva-tive scan speed used was low with a constant 2 nm slit width.The wavelength interval and the ordinate scale employed areindicated on each spectrum of figures. A Branson 3210 ultra-sonic water-bath was used to aid tablet dissolution.

2.3. Methods

2.3.1. Preparation of calibration curvesDPM stock solution was adequately prepared to obtain

standard solutions in the concentration range 9.75–32.5 µg ml−1. Derivative ultraviolet method was calibratedfor the second order. Differential wavelength was automati-cally obtained. Ordinate range for second derivative readings(D2, absorbance nm−2) was arbitrarily obtained according topeak length. Readings were taken in triplicate by seven dif-ferent groups of measurements (k = 7, n = 21) as follows. ZP(265 nm, 269 nm, 273 nm, 276 nm) and peak–peak (PP, 265–269 nm, 269–273 nm, 273–276 nm) quantitative measure-ments were evaluated by the peak amplitudes readings induplicate. DPM standard solutions in the concentration20 µg ml−1 prepared in 0.1 mol l−1 sulfuric acid were used forcomparison with samples in the precision test.

2.3.2. Preparation of pharmaceutical sampleTablet samples A and B, label claimed 2 mg of DPM per

dosage unit. The average weight was determined with 20 tab-lets, which were grounded in a mortar until fine powder.Accu-rately weighed amount of powder equivalent to 2 mg of DPMwas quantitatively transferred to a 100 ml calibrated flask withthe aid of 30 ml 0.1 mol l−1 sulfuric acid. Ultrasonic water-bath was used for 5 min to aid complete dissolution. The solu-tions were diluted to volume and final concentration wasequivalent to 20 µg ml−1 DPM.

2.3.3. PrecisionPrecision repeatability test was performed by measuring

derivative absorbances for D2 through ZP and PP amplitudesin the same day and experimental conditions. Intermediateprecision was performed by measuring derivative absor-bances for D2 through ZP and PP amplitudes in different days.Duplicate readings of 10 determinations of tablet solutions(20 µg ml−1) were performed for the precision tests.

2.3.4. RecoveryRecovery test was performed using the standard addition

method by measuring D2 absorbances at all peaks or valleys.

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Different volumes (5,0, 10,0 and 15,0 ml) of a 20 µg ml−1

DPM standard solution were added to a 200 ml calibratedflask containing 20 µg ml−1 DPM sample solutions. The theo-retical final concentration values in each flask were 15, 20 and25 µg ml−1. Sample solutions were individually prepared withconcentration 20 µg ml−1 in 0.1 mol l−1 sulfuric acid. Resultsare the average of triplicate readings of all determinations inthe tablet solutions.

3. Results and discussion

Common drug analysis techniques such as volumetricmethods are rapid and practical for quantitative determina-tion of bulk drugs in raw materials. Finished products mayusually require sample extraction or matrix elimination pre-vious to quantitative analysis.

Ultraviolet spectrophotometric methods have widely beenemployed in the quality control analyses of pharmaceuticalpreparations due to their rapidity and simplicity. However,they can be affected by matrices interferences or other drugsspectra superposition. Derivative ultraviolet spectrophotom-etry, nevertheless, can be applied to many finished productswith the advantage of solving subtle problems due to detailedslope differentiation and yielding a clear spectrum [15,16].

DPM is present in many pharmaceutical dosage forms asthe only drug or in association with other drugs. Many authorshave reported DPM determination by first order derivativespectrophotometry. However, they have not described whetherdiluted DPM solutions were colored or not. This presents agreat problem for ultraviolet spectrophotometric determina-tions due to colored background interference. The use of D2

derivative ultraviolet spectrophotometry to determine DPMin the presence of coloring agents is herein described.

Selection of the derivative parameters for DPM analysiswere standardized based upon the elimination of backgroundinterferences and the peaks profile of the curves. Standard,sample A and its placebo solution curves were scanned in theregion 235–320 nm in the zero, first and second derivativeorders. Selectivity was evaluated whether peaks and troughsof the derivative spectra of standard and sample solutions (insame concentration) were coincident and placebo furnishedno interference in the given derivative ultraviolet region.

The differential wavelength for DPM determination wasautomatically chosen. The antihistaminic spectrum wasobtained with a reasonable signal-to-noise ratio (SNR) yield-ing a smooth curve with good resolution [16]. Calibrationgraphs for D2 (Fig. 2) were found linear in the establishedconcentration range 9.75–32.5 µg ml−1 with a correlation coef-ficient of 0.9999 for all tested methods. Simple linear regres-sion analysis of measurements utilized to obtain statisticalresults of calibration was evaluated through the least-squaresmethod [17].

Fig. 2 presents the overlay of second order ultraviolet spec-tra of DPM standard samples (P) in the concentration of 9.75–32.5 µg ml−1 in 0.1 mol l−1 sulfuric acid. As demonstrated inthe figure, the spectra present two characteristic maxima andtwo minima. Maxima are represented at D269 and D276 andminima are shown at D265 and D273. Besides all the ZP indi-vidual absorptions readings, the absorptions at PP can alsobe measured at D265

269, D269273 and D273

276 by adding the respectiveabsolute readings at each one of the considered zero-peakmeasurements. This study shows the absorbance intensitiesin all these wavelengths at ZP or PP and points out whichmethod could appropriately be selected for determination ofDPM in the presence of coloring agents as excipient in tab-lets. The regression equation, Y = a + bX, in which,Y = second derivative absorbance (absorbance nm−2) andX = DPM concentration (µg ml−1), and its statistical param-eters are represented in Table 1. The average relative stan-dard deviations for ZP and PP (0.18–1.08%), D2 determina-tions are also represented in Table 1. All methods ofquantitation used provided low relative standard deviationsbelow 2.0%.

Fig. 3 shows the overlay of zero, first and second deriva-tives absorption curves of tablet sample A and its placebocompared to the standard solution curve with correspondentDPM concentration 20 µg ml−1. The presence of backgroundinterference in zero order spectra is attested due to the addi-tional absorbance seen in sampleA, in comparison to the stan-dard solution. It is important to observe that the first orderderivative is not a good choice to determine DPM in thissample because the exact superposition does not occur. Sec-ond order derivative order is more appropriate for DPM quan-titative analysis. Fig. 4 shows the overlay spectra of zero andsecond derivative absorption curves of tablet samples A, B

Fig. 2. Overlay of second derivative absorption curves of standard solutions in the concentration points 9.75, 13.0, 16.25, 19.5, 22.75, 26.0, 29.25, 32.5 µg ml−1

DPM in 0.1 mol l−1 sulfuric acid.

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and standard solution with correspondent DPM concentra-tion 20 µg ml−1. Sample B curve shows greater backgroundinterference than sample A in zero order, which seemed to betotally eliminated in the second order curve.

It is known that higher derivative orders yield sharper peakamplitudes, but also yield lower derivative absorbance unitsof the bandwidth for a given curve [16]. The best selection ofa derivative order or method is when there is a greater reduc-tion of the broad interfering [18]. For the purpose of this inves-tigation nevertheless, all peaks were considered for quantita-tive evaluation. Thereafter, statistical analysis was performedin all methods of quantitation. To verify the capability of

regression equations to predict the absorbance behavior ofDPM in dosage forms, the method was tested for precisionand recovery. The results are shown in Tables 2 and 3, respec-tively. The method precision was evaluated through the repeat-ability test, performed with two replicates of 10 determina-tions compared to standard solutions. Statistical analysisshowed low values of relative standard deviation (R.S.D.)ranges for repeatability ZP methods (0.83–1.40%) and forPP methods (0.63–0.83%) in tablets sample A with labeledclaim values between 95.77% and 98.43%. Sample B showedthe following percentage results D265 = 106.11%, D269 =116.15%, D273 = 101.86%, D276 = 97.31% for ZP methods,

Table 1Statistical resultsa for calibration graphs of DPM by second order derivative ultraviolet spectrophotometry using zero-peak (ZP) and peak–peak (PP) methods

Method ZP PPParameterk (nm) 265 269 273 276 265–269 269–273 273–276b 4.5 × 10−6 2.2 × 10−4 2.8 × 10−4 1.7 × 10−4 6.7 × 10−4 5.00 × 10−4 4.50 × 10−4

a –4.05 × 10−6 1.10 × 10−4 3.22 × 10−5 3.29 × 10−5 1.07 × 10−4 7.73 × 10−5 6.72 × 10−7

%R.S.D. 0.26 1.08 0.53 0.98 0.51 0.63 0.18t –0.155 2.021 –0.968 0.874 1.393 1.088 0.038

a Average of two determinations; B, angular coefficient; A, intercept; %R.S.D., relative standard deviation (N = 8); t, Student’s t parameter for P = 0.05.

Fig. 3. Overlay of zero (a), first (b) and second derivative (c) absorption curves of tablet solutions samples A (----) and standard solution curve (—) withcorrespondent concentration of 20 µg ml−1 DPM in 0.1 mol l−1 sulfuric acid. Sample A placebo is represented by curve (.....).

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and D265269 = 109.50%, D269

273 = 108.32% and D273276 = 100.13%

for PP methods. Repeatability and intermediate precisionresults for sample A showed low R.S.D. values (< 2.0%) indi-cating good precision within and between days.

Statistical analysis for sample A and B was performedusing Prism Graph Pad software (San Diego, CA, USA). Thet-test was performed for deciding whether or not two particu-lar average absorbance samples are different (P < 0.05,approximate t Table value 2.145 for n – k = 21 – 7 = 14degrees of freedom). In all pair groups the statistic t weresmaller in magnitude than the critical value (t Table value).Results showed no significant difference in the second deriva-tive absorbances samples [17].

In Table 3, the recovery test results determined throughthe standards addition method [19] conducted in sample Aand placebo matrices are shown. Satisfactory values at D2

were obtained for the addition of standards in samples A andin its placebo. Recovery results from sample A was found inthe range of 96.95–105.61%. The standard addition to thesample A placebo only resulted in less variable recovery val-

ues in the range of 100.58–104.67% for ZP and PP methods.These results show that both methods presented good accu-racy.

Absorbance readings in a greater wavelength is preferredin detriment of all other lower wavelength readings, eventhough they show larger amplitude peaks. Regardless there isno significant difference between the second derivative absor-bance measurements in the described conditions; the recom-mended best feasible method for DPM determination is theZP at D273. It showed lower R.S.D. values for intermediateprecision and interference level (intercept value) for its cali-bration curve. In addition, it is positioned in a region of greaterwavelength and less interferences in the spectrum, and needsonly one absorbance measurement [20].

Despite the pink color yielded from tablet solutions, D2

derivative ultraviolet spectrophotometry was successfully usedwith no need for extractions or previous treatment of color-ing agents or excipients. The proposed method proved to bemore advantageous than traditional dosage methods becauseof the easy one-step, timesaving and low-cost analysis.

Fig. 4. Overlay of zero (a) and second derivative (b) absorption curves of tablet sample A (----), sample B (.....) and standard solutions (—) with correspondentconcentration of 20 µg ml−1 DPM in 0.1 mol l−1 sulfuric acid.

Table 2Statistical resultsa obtained in quantitative analysis of DPM in Sample A by second derivative ultraviolet spectrophotometry, zero-peak (ZP) and (peak–peak)PP quantitative methods

Method ZP PPParameterk (nm) 265 269 273 276 265–269 269–273 273–276Absorbance (× 10−3) 8.592 4.267 5.179 3.217 12.859 9.446 8.395Concentration (µg ml−1) 19.685 19.143 19.162 19.461 19.502 19.154 19.276Found (%) 98.43 95.72 95.81 97.31 97.31 95.77 96.38Average precision (%R.S.D.) 0.83 1.12 0.99 1.40 0.63 0.83 0.80Intermediate precision (%R.S.D.) 0.98 1.62 1.11 1.67 0.97 0.98 0.80

a Average of 10 determinations.

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4. Conclusion

The determination of DPM in colored tablets by secondderivative ultraviolet spectrophotometry with no interferenceof the coloring agent used was successfully achieved. Theinconvenient of a previous extraction or other sample pre-treatment was easily by-passed. The second order derivativeultraviolet spectrophotometric method described indicatesprecision, accuracy, simplicity, rapidity and a low-cost alter-native technique for DPM determination in tablets, due to itspotential in interferences elimination.

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Table 3DPM recovery resultsa. Standard addition tests for sample A and for its pla-cebo by second derivative ultraviolet spectrophotometry, zero-peak (ZP) and(peak–peak) PP quantitative methods

Method Standardadded

Standardadded

ZP or PP (µg ml−1) Found Recovery (µg ml−1) Found Recovery(k, nm) Sample A (µg ml−1) (%) Placebo (µg ml−1) (%)265 5.20 5.14 98.84 15.61 16.07 102.88

10.41 10.26 98.51 20.82 21.28 102.2115.61 15.86 101.56 26.02 26.87 103.26

269 5.20 5.28 101.48 15.61 16.03 102.6310.41 10.99 105.61 20.82 21.55 103.5015.61 15.45 98.93 26.02 26.75 102.77

273 5.20 5.24 100.61 15.61 16.07 102.8810.41 10.66 102.44 20.82 20.94 100.5815.61 15.99 102.37 26.02 26.63 102.33

276 5.20 5.05 96.95 15.61 16.34 104.6710.41 10.60 101.79 20.82 21.07 101.1915.61 15.41 98.70 26.02 27.22 104.61

265/269 5.20 5.19 99.71 15.61 16.05 102.8010.41 10.50 100.85 20.82 21.37 102.6315.61 15.72 100.69 26.02 26.83 103.10

269/273 5.20 5.26 101.00 15.61 16.05 102.7710.41 10.81 103.86 20.82 21.21 101.8815.61 15.75 100.83 26.02 26.68 102.53

273/276 5.20 5.16 99.22 15.61 16.17 103.5610.41 10.64 102.19 20.82 20.99 100.8115.61 15.77 100.97 26.02 26.86 103.20

a Average of two determinations.

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