Simultaneous Determination of Pyridoxine, Meclizine and Buclizine in Dosage Formulations and Human Serum by RP-LC

Rapid liquid chromatographic procedures for analytical quality control of pharmaceutical preparations and human serum containing antihistamine drugs, meclizine and buclizine alone or in combination with pyridoxine are proposed, using acetonitrile:water (80:20) as a mobile phase (pH adjusted to 2.6), methylparaben as internal standard and UV detection was made at 230 nm. The results obtained showed a good agreement with the declared content. The method shows good linearity in the range of 30–10,000 ng mL^?1 for pyridoxine and 25–10,000 ng mL^?1 for meclizine and buclizine serum concentrations with a correlation coefficient 0.9999 (inter- and intra-day CV < 3.91%). The recovery was >97.8%. The proposed method may be used for the quantitative analysis of meclizine and buclizine alone or in combination with pyridoxine from raw materials, in bulk drugs, dosage formulations and in serum.     

Simultaneous Determination of Pyridoxine,Meclizine and Buclizine in Dosage Formulations and Human Serum by RP-LC

Rapid liquid chromatographic procedures for analytical quality control of pharmaceutical preparations and human serum containing antihistamine drugs, meclizine and buclizine alone or in combination with pyridoxine are proposed, using acetonitrile:water (80:20) as a mobile phase (pH adjusted to 2.6), methylparaben as internal standard and UV detection was made at 230 nm. The results obtained showed a good agreement with the declared content. The method shows good linearity in the range of 30–10,000 ng mL⁻¹ for pyridoxine and 25–10,000 ng mL⁻¹ for meclizine and buclizine serum concentrations with a correlation coefficient 0.9999 (inter- and intra-day CV < 3.91%). The recovery was >97.8%. The proposed method may be used for the quantitative analysis of meclizine and buclizine alone or in combination with pyridoxine from raw materials, in bulk drugs, dosage formulations and in serum.

     

Spectrofluorimetric determination of some water-soluble vitamins

Two simple and sensitive spectrofluorimetric methods were developed for determination of three water-soluble vitamins (B1, B2, and B6) in mixtures in the presence of cyanocobalamin. The first one was for thiamine determination, which depends on the oxidation of thiamine HCl to thiochrome by iodine in an alkaline medium. The method was applied accurately to determine thiamine in binary, ternary, and quaternary mixtures with pyridoxine HCl, riboflavin, and cyanocobalamin without interference. In the second method, riboflavin and pyridoxine HCl were determined fluorimetrically in acetate buffer, pH 6. The three water-soluble vitamins (B1, B2, and B6) were determined spectrofluorimetrically in binary, ternary, and quaternary mixtures in the presence of cyanocobalamin. All variables were studied in order to optimize the reaction conditions. Linear relationship was obeyed for all studied vitamins by the proposed methods at their corresponding lambda(exc) or lambda(em). The linear calibration curves were obtained from 10 to 500 ng/mL; the correlation ranged from 0.9991 to 0.9999. The suggested procedures were applied to the analysis of the investigated vitamins in their laboratory-prepared mixtures and pharmaceutical dosage forms from different manufacturers. The RSD range was 0.46-1.02%, which indicates good precision. No interference was observed from common pharmaceutical additives. Good recoveries (97.6 +/- 0.7-101.2 +/- 0.8%) were obtained. Statistical comparison of the results with reported methods shows excellent agreement and indicates no significant difference in accuracy and precision.     

Development of a validated HPLC method for the determination of B-complex vitamins in pharmaceuticals and biological fluids after solid phase extraction

An HPLC method was developed for the simultaneous determination of seven water-soluble vitamins, viz. thiamine, riboflavin, nicotinic acid, nicotinamide, pyridoxine, cyanocobalamin, and folic acid, in multivitamin pharmaceutical formulations and biological fluids (blood serum and urine). Separation was achieved at ambient temperature on a Phenomenex Luna C18 (150 x 4.6 mm) analytical column. Gradient elution was performed starting at a 99:1 A:B v/v composition, where A: 0.05 M CH3COONH4/CH3OH (99/1) and B: H2O/CH3OH (50/50), at a flow rate of 0.8 mL/min. After a 4-min isocratic elution the composition was changed to 100% of B in 18 min and elution continued isocratically for 8 min. Detection was performed with a photodiode array detector at 280 nm. Each vitamin was quantitatively determined at its maximum wavelength. Spectral comparison was used for peak identification in real samples. Detection limits were in the range of 1.6-3.4 ng, per 20-microL injection, while linearity held up to 25 ng/microL. Accuracy, intra-day repeatability (n = 6), and inter-day precision (n = 7) were found to be satisfactory. Theobromine (2 ng/microL) was used as internal standard. Sample preparation of biological fluids was performed by SPE on Supelclean LC-18 cartridges with methanol-water 85/15 v/v as eluent. Extraction recoveries from biological matrices ranged from 84.6% to 103.0%.     

Determination of water-soluble vitamins by liquid chromatography with a parallel dual-electrode electrochemical detector

A method for the determination of water-soluble vitamins (ascorbic acid, pyridoxine hydrochloride, pyridoxal hydrochloride, pyridoxamine dihydrochloride, p-aminobenzoic acid, folic acid) by liquid chromatography, with a parallel dual-electrode electrochemical detector, is described. One electrode was controlled at +0.80 V (vs. SCE), the other at +1.20 V (vs. SCE). The possibility of interference by eight other water-soluble vitamins (riboflavin, nicotinamide, cyanocobalamin, menadione, dextro calcium pantothenate, thiamine, nicotinic acid, dextro biotin) was studied. These vitamins did not interfere when a parallel dual-electrode detector system was used. The estimation of five of the vitamins was studied in detail. The linear ranges found were 10 ng-1.2 mug for pyridoxine hydrochloride, 2 ng-2mug for pyridoxal hydrochloride, 10 ng-3 mug for pyridoxamine dihydrochloride, 5-200 ng for folic acid and 0.6-200 ng for p-aminobenzoic acid, the limits of detection being 3, 0.6, 1, 2 and 0.06 ng respectively. Application of the technique to the estimation of vitamin B(6) in tablets is illustrated. The results indicate that the vitamin B(6) in these tablets existed in the pyridoxine hydrochloride form and the B(6) content agreed well with liquid chromatograph by spectrophotometric analysis.     

Chemometric methods for the simultaneous determination of some water-soluble vitamins

Two spectrophotometric methods, derivative and multivariate methods, were applied for the determination of binary, ternary, and quaternary mixtures of the water-soluble vitamins thiamine HCI (I), pyridoxine HCI (II), riboflavin (III), and cyanocobalamin (IV). The first method is divided into first derivative and first derivative of ratio spectra methods, and the second into classical least squares and principal components regression methods. Both methods are based on spectrophotometric measurements of the studied vitamins in 0.1 M HCl solution in the range of 200-500 nm for all components. The linear calibration curves were obtained from 2.5-90 microg/mL, and the correlation coefficients ranged from 0.9991 to 0.9999. These methods were applied for the analysis of the following mixtures: (I) and (II); (I), (II), and (III); (I), (II), and (IV); and (I), (II), (III), and (IV). The described methods were successfully applied for the determination of vitamin combinations in synthetic mixtures and dosage forms from different manufacturers. The recovery ranged from 96.1 +/- 1.2 to 101.2 +/- 1.0% for derivative methods and 97.0 +/- 0.5 to 101.9 +/- 1.3% for multivariate methods. The results of the developed methods were compared with those of reported methods, and gave good accuracy and precision.     

Simultaneous determination of a ternary mixture of doxylamine succinate, pyridoxine hydrochloride, and folic acid by the ratio spectra-zero-crossing, double divisor-ratio spectra derivative, and column high-performance liquid chromatographic methods

Three simple, rapid, and accurate methods, i.e., the derivative ratio spectra-zero-crossing method (method I), double divisor-ratio spectra derivative method (method II), and column reversed-phase high-performance liquid chromatographic (RP-HPLC) method (method III) were developed for the simultaneous determination of doxylamine succinate (DOX), pyridoxine hydrochloride (PYR), and folic acid (FA) in their ternary mixtures and in tablets. In methods I and II, the calibration graphs were linear in the range of 2.5-80, 1.0-40, and 1.0-30 microg/mL for DOX, PYR, and FA, respectively. In the HPLC method, the separation of these compounds was performed using mobile phase consisting of 0.05 M phosphate buffer (pH 6.3)-methanol-acetonitrile (50 + 20 + 30, v/v/v), and UV detection was performed at 263 nm. Linearity was observed between the concentrations of the analytes and peak areas [correlation coefficient (r) > or =0.9998] in the concentration range of 1.0-200, 4.0-600, and 4.0-600 microg/mL for DOX, PYR, and FA, respectively. The standard deviation of retention time in method III was 0.011, 0.015, and 0.016 for DOX, PYR, and FA, respectively. The precision studies for all of the methods gave relative standard deviation values of <2%. The results obtained from the methods were statistically compared by means of Student's t-test and the variance ratio F-test. It was concluded that all of the developed methods were equally accurate, sensitive, and precise. These methods could be applied to determine DOX, PYR, and FA in their combined dosage forms.     

Simultaneous determination of nicotinamide,pyridoxine hydrochloride,thiamine mononitrate and riboflavin in multivitamin with minerals tablets by reversed-phase ion-pair high performance liquid chromatography

A reversed-phase ion-pair high performance liquid chromatographic method (HPLC) has been developed and validated for the routine analysis of nicotinamide, pyridoxine hydrochloride, thiamine mononitrate and riboflavin in multivitamin with minerals tablets. HPLC separation of the vitamins was performed on a Hypersil C(18) column and detected by ultraviolet absorbance at 280 nm. The use of methanol-aqueous 0.5% acetic acid solution (18:82, v/v; containing 2.5 mM sodium hexanesulfonate, pH = 2.8) as the mobile phase at a flow-rate of 1.2 mL/min enables the baseline separation of the four analytes free from interferences with isocratic elution at 30 degrees C. The analysis time was 17 min per injection. The method was linear in the ranges of 5-90, 2.5-90, 5-95 and 25-450 micro g/mL for thiamine mononitrate, riboflavin, pyridoxine hydrochloride and nicotinamide, respectively. The average coefficients of variation of within- and between-day assays were 2.2 and 3.6% for thiamine mononitrate, 1.8 and 2.4% for riboflavin, 1.3 and 1.7% for pyridoxine hydrochloride and 1.0 and 1.5% for nicotinamide, respectively. The average recoveries of thiamine mononitrate, riboflavin, pyridoxine hydrochloride and nicotinamide were 97.0, 97.2, 98.9 and 100.4% for the tablets, respectively. The method has been successfully applied to the simultaneous determination of nicotinamide, pyridoxine hydrochloride, thiamine mononitrate and riboflavin in multivitamin with minerals tablets.     

Determination of tryptophan in raw materials, rat brain and human plasma by RP-HPLC technique

This paper describes tryptophan (TRP) estimation in raw human plasma and rat brain by reversed-phase high-performance liquid chromatography (RP-HPLC). Estimation was carried out on a Purospher STAR C18 column using water-acetonitrile (90:10 v/v, at pH 2.7) mixture at a rate of 1.5 mL/min as mobile phase. Eluents were monitored at 273 nm by an ultraviolet detector. The method was linear (R(2) > 0.999), precise (intra-day and inter-day precision <2%) in the range of 0.25-20 μg/mL. The detection and quantification limits were 0.0144 μg/mL and 0.0437 μg/mL, respectively. In human plasma, Day 1 and Day 2 precision were 0.054-2.29% and 1.66-3.7%; whereas precisions in rat brain were 1.23-2.3% and 0.677-4.2%, respectively. The method was applied to study TRP level in human smokers and in arthritic rat brain. An efficient RP-HPLC method was developed for TRP determination that worked for clinical and research purposes.     

Nanocrystalline Metallosilicate Modified Electrodes for the Simultaneous,Sensitive, and Selective Determination of Riboflavin,Rutin, and Pyridoxine

Nanocrystalline metallosilicate modified glassy carbon electrodes were fabricated for the simultaneous determination of vitamins. Among these, nanocrystalline zirconosilicate exhibited the highest activity with a linear range from 30 nM–500 µM for riboflavin and 120 nM–600 µM for rutin and pyridoxine. Sensitivity values of 2.8, 1.49, and 1.13 µA/µM cm² and lower detection limits of 5 nM, 30 nM, and 30 nM for riboflavin, rutin, and pyridoxine, respectively, were found. The proposed sensor is stable and reproducible (RSD<3.5 %). The analytical performance of this sensor was demonstrated in the pharmaceutical preparations (multivitamin tablets) with satisfactory recovery (97–103 %).     

Separation of water-soluble vitamins by reversed-phase high performance liquid chromatography with ultra-violet detection: application to polyvitaminated premixes

Nine water-soluble vitamins: [thiamine (B1), ascorbic acid (C), nicotinamide (PP), pyridoxine (B6), calcium pantothenate (B5), folic acid (B9), cyanocobalamin (B12), riboflavin (B2) and biotin (B8)] were separated on a YMC-Pack Pro C18 column (250 mm x 4.6 mm, 5 microm particle size) in a single run with a gradient elution of mobile phase consisting of 0.025% trifluoroacetic acid pH 2.6 (solvent A) and acetonitrile (solvent B). The separation was achieved within 17 min with a flow rate of 0.8 ml min(-1) and the detection was performed at two wavelengths (210 and 275 nm). The calibration graphs plotted with six concentrations of each vitamin were linear with a regression coefficient R2 > 0.995. The method was applied for the quantification of vitamins B1, C, PP, B6, B5, B9 B2 and B8 in polyvitaminated premixes (premixes) used for the fortification of infant nutrition products. The sample preparation involves an aqueous extraction of vitamins and two different samples dilution were used prior the LC-analysis. The specificity of the method was demonstrated by the retention characteristics, UV spectra and by comparing the peak purity with the standard of each vitamin. The repeatability of the method was evaluated at different level of concentrations on 12 premixes and the coefficients of variation (CVr) were below 6.5%. The values of the intermediate precision (CV1) were below 9.6% (n = 6). The concentrations of vitamins found in premixes with our method were comparable to the declared values, since no bias was found between the two sets of results at 95% confidence. The simplicity of the procedure should make it highly desirable for quality control of premixes in the food industry.     

Kinetic and thermodynamic studies on ligand substitution reactions and base-on/base-off equilibria of cyanoimidazolylcobamide, a vitamin B12 analog with an imidazole axial nucleoside

Ligand substitution reactions of the vitamin B12 analog cyanoimidazolylcobamide, CN(Im)Cbl, with cyanide were studied. Cyanide substitutes imidazole (Im) in the alpha-position more slowly than it substitutes dimethylbenzimidazole in cyanocobalamin (vitamin B12). The kinetics of the displacement of Im by CN- showed saturation behaviour at high cyanide concentration; the limiting rate constant was found to be 0.0264 s(-1) at 25 degrees C and is characterized by the activation parameters: DeltaH(not =) = 111 +/- 2 kJ mol(-1), DeltaS(not =) = +97 +/- 6 J K(-1) mol(-1), and DeltaV(not =) = +9.3 +/- 0.3 cm3 mol(-1). These parameters are interpreted in terms of an I(d) mechanism. The equilibrium constant for the reaction of CN(Im)Cbl with CN- was found to be 861 +/- 75 M(-1), which is significantly less than that obtained for the reaction of cyanocobalamin with CN- (viz. 10(4) M(-1)). pKbase-off for the base-on/base-off equilibrium was determined spectrophotometrically and found to be 0.99 +/- 0.05, which is about 0.9 pH units higher than that obtained previously in the case of cyanocobalamin. In addition, the kinetics of the base-on/base-off reaction was studied using a pH-jump technique and the data obtained revealed evidence for an acid catalyzed reaction path. The results obtained in this study are discussed in reference to those reported previously for cyanocobalamin.     

Simultaneous Determination of Caffeine and Pyridoxine in Energy Drinks using Differential Pulse Voltammetry at Glassy Carbon Electrode Modified with Nafion®

A new voltammetric method for simultaneous determination of caffeine and pyridoxine present in different types of commercial energy drinks has been developed. This electroanalytical method is based on anodic oxidation of these biologically active nitrogen‐containing heterocycles at glassy carbon electrode covered with thin layer of sulfonated fluoropolymer Nafion® using differential pulse voltammetry in 0.1 M Britton‐Robinson buffer of pH 4.5 at potential step 5 mV, potential amplitude 70 mV, and scan rate 50 mV/s. Linear ranges for caffeine and pyridoxine determination were 63.1–600 μM and 7.5–200 μM with the detection limits of 18.9 and 2.2 μM, respectively. It was validated using high‐performance liquid chromatography with spectrophotometric detection. Obtained results have shown that voltammetric approach is very simple and low‐cost analytical method which can be used for routine determination of caffeine and pyridoxine in energy drinks.     

Application of liquid chromatography to the simultaneous determination of acetylsalicylic acid, caffeine, codeine, paracetamol, pyridoxine, and thiamine in pharmaceutical preparations.

This paper describes a rapid reversed-phase liquid chromatographic method, with UV detection, for the simultaneous determination of acetylsalicylic acid, caffeine, codeine, paracetamol, pyridoxine, and thiamine in pharmaceutical preparations. A reversed-phase C18 Nucleosil column is used. The mobile phase consists of 2 successive eluants: water (5 min) and acetonitrile-water (75 + 25, v/v; 9 min), both adjusted to pH 2.1 with phosphoric acid. Before determination acetylsalicylic acid is completely converted to salicylic acid by alkaline hydrolysis. Salicylic acid, caffeine, paracetamol, pyridoxine, and thiamine are all detected at 285 nm, whereas codeine is detected at 240 nm. Calibration curves were linear for salicylic acid, caffeine, paracetamol, and pyridoxine in the range of 50-500 mg/L, and for codeine and thiamine in the range of 50-1000 mg/L. The method was applied to the analysis of 13 fortified commercial pharmaceutical preparations. Recoveries ranged from 92.6 to 105.5%, with relative standard deviations of 1.1-5.8%.     

LC Simultaneous Determination of Thioctic Acid, Benfotiamine and Cyanocobalamin in Thiotacid Compound Capsules

A simple, selective, sensitive, precise, simultaneous liquid chromatographic analysis of capsules containing thioctic acid, benfotiamine and cyanocobalamin was described. Good chromatographic separation was achieved using a Zorbax C18 (4.6 cm × 250 mm, 5 μm) and a mobile phase consisting of acetonitrile–phosphate buffer pH 3.5 (15:85, v/v) at a flow rate of 0.9 mL min^?1. The ultraviolet detector was set a wavelength of 280 nm. Thioctic acid, benfotiamine and cyanocobalamin were eluted at 2.869, 3.752 and 13.689 min, respectively. The linear ranges for thioctic acid, benfotiamine and cyanocobalamin were 30–180, 4–24 and 0.025–0.150 μg mL^?1, respectively. The recoveries of thioctic acid, benfotiamine and cyanocobalamine in pharmaceutical preparation were all greater than 98% and their relative standard deviations were less than 2.0%. The limits of detection were 2.57, 0.19 and 0.003 μg mL^?1 for thioctic acid, benfotiamine and cyanocobalamin, respectively.     

Simultaneous determination of riboflavin, pyridoxine, nicotinamide, caffeine and taurine in energy drinks by planar chromatography-multiple detection with confirmation by electrospray ionization mass spectrometry

A new high-throughput method was developed to detect simultaneously riboflavin (Vitamin B(2)), pyridoxine (Vitamin B(6)), nicotinamide (Vitamin B(3)), caffeine and taurine in energy drinks by multiple detection. Ten samples of energy drinks and six samples of beverages containing caffeine were prepared by degassing in an ultrasonic bath for 20 min. After chromatography, multi-wavelength scanning was performed by: (1) UV-absorbance measurement at 261 nm for nicotinamide and 275 nm for caffeine, (2) fluorescence measurement at 366/>400 and 313/>340 nm for riboflavin and pyridoxine, respectively, and (3) Vis-absorbance measurement at 525 nm for taurine, after post-chromatographic derivatization with ninhydrin reagent. Calibrations were linear or polynomial with determination coefficients r(2)>0.999. Overall recoveries of the five compounds were between 81 and 106% at three different concentration levels. Repeatabilities (RSD, %) of all substances in matrix were determined to be between 0.8 and 1.5%. Intermediate precisions (RSD, %) ranged between 3.6 and 7.4% for riboflavin, 2.8 and 6.3% for nicotinamide, 2.5 and 4.4% for caffeine, 2.1 and 2.9% for taurine and 0.5 and 4.0% for pyridoxine at different concentration levels. Mass confirmation was performed by a single quadrupole MS in positive electrospray ionisation (ESI) scan mode for all substances except taurine (negative mode). This method offers a good alternative for routine analysis due to its simplicity and at the same time reliability.     

Potentiometric membrane sensors for the selective determination of pyridoxine hydrochloride (vitamin B6) in some pharmaceutical formulations

The construction and general performance characteristics of two novel potentiometric PVC membrane sensors responsive to the pyridoxine hydrochloride known as vitamin B6 (VB6) are described. These sensors are based on the use of the ion-association complexes of the pyridoxine cation with phosphomolybdate, and phosphotungstate counter anions as ion pair in a plasticized PVC matrix. The electrodes show a stable, near-Nernstian response for 6x10(-5)-1x10(-2) M VB6 at 25 degrees C over the pH range 2-4 with a cationic slope of 54.0+/-0.5 and 54.5+/-0.4 per concentration decade for pyridoxine-phosphomolybdate and pyridoxine-phosphotungstate respectively. The two electrodes have the same lower detection limit (4x10(-5) M) and the response times are 45-60 and 30-45 s in the same order for both. Selectivity coefficients for VB6 relative to a number of interfering substances were investigated. There is negligible interference from many cations, some vitamins and pharmaceutical excipients. Direct potentiometric determination of 15-2000 microg/ml pyridoxine shows an average recovery of 98.0% and 99.0% with relative standard deviation 1.5% and 1.2% at 100.0 microg/ml for pyridoxine-phosphomolybdate and pyridoxine-phosphotungstate electrodes, respectively. The determination of VB6 in some pharmaceutical preparations using the proposed electrodes gave an average recovery of 98.0 and 99.0% of the nominal value and a mean standard deviation of 1.1% and 0.9% (n=10) for pyridoxine-phosphomolybdate and pyridoxine-phosphotungstate electrodes, respectively. The results compare favorably with data obtained by the British Pharmacopoeia method.     

Quantitative profiling of biomarkers related to B‐vitamin status,tryptophan metabolism and inflammation in human plasma by liquid chromatography/tandem mass spectrometry

Vitamins B₂ and B₆ serve as cofactors in enzymatic reactions involved in tryptophan and homocysteine metabolism. Plasma concentrations of these vitamins and amino acids are related to smoking and inflammation, and correlate with other markers of immune activation. Large‐scale studies of these relations have been hampered by lack of suitable analytical methods. The assay described includes riboflavin, five vitamin B₆ forms (pyridoxal 5′‐phosphate, pyridoxal, 4‐pyridoxic acid, pyridoxine and pyridoxamine), tryptophan and six tryptophan metabolites (kynurenine, kynurenic acid, anthranilic acid, 3‐hydroxykynurenine, xanthurenic acid and 3‐hydroxyanthranilic acid), cystathionine, neopterin and cotinine. Trichloroacetic acid containing 13 isotope‐labelled internal standards was added to 60 µL of plasma, the mixture was centrifuged, and the resulting supernatant used for analysis. The analytes were separated within 5 min on a stable‐bond C8 column by a gradient‐type mobile phase containing acetonitrile, heptafluorobutyric acid and high concentration (650 mmol/L) of acetic acid, and detected using electrospray ionization tandem mass spectrometry (ESI‐MS/MS). The mobile phase ensured sufficient separation and high ionization efficiency of all analytes. Recoveries were 75–123% and within‐day and between‐day coefficients of variance (CVs) were 2.5–9.5% and 5.4–16.9%, respectively. Limits of detection ranged from 0.05 to 7 nmol/L. The method enables quantification of endogenous plasma concentrations of 16 analytes related to B‐vitamin status and inflammation, and may prove useful in large‐scale epidemiological studies. Copyright © 2009 John Wiley & Sons, Ltd.     

Ultramicrodetermination of cyanocobalamin in elemental diet by solid-phase extraction and high-performance liquid chromatography with visible detection

The ultramicrodetermination of cyanocobalamin (9 ng/g) in elemental diet containing 46 kinds of compounds, with concentrations at least 50–10⁶ times higher than that of cyanocobalamin, was performed by Sep-Pak C₁₈ purification and concentration of cyanocobalamin followed by HPLC with detection at 550 nm. The method is simple, rapid, sensitive and reproducible. The calibration graph was linear in the range of 0-0.2 μg. The recovery of cyanocobalamin was over 95% by the standard addition method. There was good agreement between the cyanocobalamin concentrations indicated and found.     

Simultaneous determination of codeine and pyridoxine in pharmaceutical preparations by first-derivative spectrofluorimetry

A method for the simultaneous determination of codeine and pyridoxine was developed, based on the measurement of their native fluorescence signals, by using first-derivative spectrofluorimetry to resolve the mixture. Codeine was measured at lambda(em) = 309 nm, and pyridoxine was measured at lambda(em) = 450 nm. Instrumental parameters were optimized, and the emission spectra were recorded between 275 and 475 nm, at lambda(ex) = 255 nm and excitation and emission slit widths of 2.5 and 10 nm, respectively. Systematic studies on the influence of species usually present along with the analytes (such as caffeine, ascorbic acid, paracetamol, and thiamine) were also performed. The calibration graphs were linear over the ranges of 0.5-7.0 and 0.1-1.0 microg/mL for codeine and pyridoxine, respectively, and the relative standard deviations (n = 10) were about 3%. The method was successfully applied to the determination of codeine and pyridoxine in solutions of synthetic mixtures and in synthetic and semisynthetic pharmaceutical formulations.