Evaluation Of Thin-layer Chromatograms Research Articles

Stability indicating spectrophotometric and spectrodensitometric methods for the determination of diatrizoate sodium in presence of its degradation product

Three sensitive, selective, and precise stability indicating methods for the determination of the X-ray contrast agent, diatrizoate sodium (DTA), in the presence of its acidic degradation product (highly cytotoxic 3,5 diamino metabolite) and in pharmaceutical formulation were developed and validated. The first method is a first derivative (D1) spectrophotometric one, which allows the determination of DTA in the presence of its degradate at 231.2nm (corresponding to zero crossing of the degradate) over a concentration range of 2–24μg/mL with mean percentage recovery 99.95±0.97%. The second method is the first derivative of the ratio spectra (DD1) by measuring the peak amplitude at 227nm over the same concentration range as D1 spectrophotometric method, with mean percentage recovery 99.99±1.15%. The third method is a TLC-densitometric one, where DTA was separated from its degradate on silica gel plates using chloroform:methanol:ammonium hydroxide (20:10:2 by volume) as a developing system. This method depends on quantitative densitometric evaluation of thin layer chromatogram of DTA at 238nm over a concentration range of 4–20μg/spot, with mean percentage recovery 99.88±0.89%. The selectivity of the proposed methods was tested using laboratory-prepared mixtures. The proposed methods have been successfully applied to the analysis of DTA in pharmaceutical dosage forms without interference from other dosage form additives. The results were statistically compared with the official US pharmacopeial method. No significant difference for either accuracy or precision was observed.

Stability‐indicating spectrophotometric and spectrodensitometric methods for the determination of diacerein in the presence of its degradation product

Three sensitive, selective, and precise stability-indicating methods for the determination of the novel osteoarthritis drug, diacerein (DIA) in the presence of its alkaline degradation product (active metabolite, rhein) and in pharmaceutical formulation were developed and validated. The first method is a first derivative (D(1) ) spectrophotometric one, which allows the determination of DIA in the presence of its degradate at 322 nm (corresponding to zero crossing of the degradate) over a concentration range of 4-40 µg/mL with mean percentage recovery 100.21 ± 0.833. The second method is the first derivative of the ratio spectra (DD(1) ) by measuring the peak amplitude at 352 nm over the same concentration range as (D(1) ) spectrophotometric method, with mean percentage recovery 100.09 ± 0.912. The third method is a TLC-densitometric one, where DIA was separated from its degradate on silica gel plates using ethyl acetate:methanol:chloroform (8:1.5:0.5 v:v:v) as a developing system. This method depends on quantitative densitometric evaluation of thin layer chromatogram of DIA at 340 nm over a concentration range of 1-10 µg/spot, with mean percentage recovery 100.24 ± 1.412. The selectivity of the proposed methods was tested using laboratory-prepared mixtures. The proposed methods have been successfully applied to the analysis of DIA in pharmaceutical dosage forms without interference from other dosage form additives and the results were statistically compared with reference method.

Spectrophotometric and spectrodensitometric methods for the determination of rivastigmine hydrogen tartrate in presence of its degradation product

Three sensitive, selective and precise stability-indicating methods for the determination of the anti-Alzheimer's drug, rivastigmine hydrogen tartrate (RIV) in the presence of its alkaline degradation product (major metabolite, NAP 226-90) and in pharmaceutical formulation were developed and validated. The first method is a second derivative (D(2)) spectrophotometric one, which allows the determination of RIV in the presence of its degradate at 262 nm (corresponding to zero crossing of the degradate) over a concentration range of 50-500 microg/ml with mean percentage recovery 100.18 +/- 0.628. The second method is the first derivative of the ratio spectra (DD(1)) by measuring the peak amplitude at 272 nm over the same concentration range as (D(2)) spectrophotometric method, with mean percentage recovery 99.97 +/- 0.641. The third method is a TLC-densitometric one, where RIV was separated from its degradate on silica gel plates using methanol:butanol:H(2)O:ammonia (5:4:1:0.01 v:v:v) as a developing system. This method depends on the quantitative densitometric evaluation of thin layer chromatogram of RIV at 263 nm over a concentration range of 20-160 microg/spot, with mean percentage recovery 100.19 +/- 1.344. The selectivity of the proposed methods was tested using laboratory-prepared mixtures. The proposed methods have been successfully applied to the analysis of RIV in pharmaceutical dosage forms without interference from other dosage form additives and the results were statistically compared with reference method.

Spectrophotometric and spectrodensitometric determination of Clopidogrel Bisulfate with kinetic study of its alkaline degradation

Four sensitive, selective and precise stability-indicating methods for the determination of Clopidogrel Bisulfate (CLP) in presence of its alkaline degradate and in pharmaceutical formulations were developed and validated. Method A is a second derivative (D 2) spectrophotometric one, which allows the determination of CLP in presence of its alkaline degradate at 219.6, 270.6, 274.2 and 278.4 nm (corresponding to zero-crossing of the degradate) over a concentration range of 4–37 μg mL −1 with mean percentage recoveries 99.81 ± 0.893, 99.72 ± 0.668, 99.88 ± 0.526 and 100.46 ± 0.646, respectively. CLP can be determined in the presence of up to 65% of its degradate. D 2 method was used to study the kinetic of CLP alkaline degradation that was found to follow a first-order reaction. The t 1/2 was 6.42 h while K (reaction rate constant) was 0.1080 mol/h. Method B is the first derivative of the ratio spectra (DD 1) spectrophotometric method, by measuring the peak amplitude at 217.6 and 229.4 nm using acetonitrile and CLP can be determined in the presence of up to 70% of its degradate. The linearity range was 5–38 μg mL −1 with mean percentage recoveries 99.88 ± 0.909 and 99.70 ± 0.952, respectively. Method C based on the determination of CLP by the bivariate calibration depending on simple mathematic algorithm which provides simplicity and rapidity. The method depends on quantitative evaluation of the absorbance at 210 and 225 nm over a concentration range 5–38 μg mL −1 with mean percentage recovery 99.27 ± 1.115. CLP can be determined in the presence of up to 70% of its degradate. Method D is a TLC-densitometric one, where CLP was separated from its degradate on silica gel plates using hexane:methanol:ethyl acetate (8.7:1:0.3, v/v/v) as a developing system. This method depends on quantitative densitometric evaluation of thin layer chromatogram of CLP at 248 nm over a concentration range of 0.6–3 μg/band with mean percentage recovery 99.97 ± 1.161. CLP can be determined in the presence of up to 90% of its alkaline degradate. The selectivity of the proposed methods was checked using laboratory prepared mixtures. The proposed methods have been successfully applied to the analysis of CLP in pharmaceutical dosage forms without interference from other dosage form additives and the results were statistically compared with the official method.

Application of spectrophotometric, densitometric, and HPLC techniques as stability indicating methods for determination of Zaleplon in pharmaceutical preparations

Spectrophotometric, spectrodensitometric and HPLC are stability indicating methods described for determination of Zaleplon in pure and dosage forms. As Zaleplon is easily degradable, the proposed techniques in this manuscript are adopted for its determination in presence of its alkaline degradation product, namely N-[4-(3-cyano-pyrazolo[1,5a]pyridin-7-yl)-phenyl]- N-ethyl-acetamide. These approaches are successfully applied to quantify Zaleplon using the information included in the absorption spectra of appropriate solutions. The second derivative (D 2) spectrophotometric method, allows determination of Zaleplon without interference of its degradate at 235.2 nm using 0.01N HCl as a solvent with obedience to Beer's law over a concentration range of 1–10 μg ml −1 with mean percentage recovery 100.24 ± 0.86%. The first derivative of the ratio spectra ( 1DD) based on the simultaneous use of ( 1DD) and measurement at 241.8 nm using the same solvent and over the same concentration range as (D 2) spectrophotometric method, with mean percentage recovery 99.9 ± 1.07%. The spectrodensitometric analysis allows the separation and quantitation of Zaleplon from its degradate on silica gel plates using chloroform:acetone:ammonia solution (9:1:0.2 by volume) as a mobile phase. This method depends on quantitave densitometric evaluation of thin layer chromatogram of Zaleplon at 338 nm over a concentration range of 0.2–1 μg band −1, with mean percentage recovery 99.73 ± 1.35. Also a reversed-phase liquid chromatographic method using 5-C8 (22 cm × 4.6 mm i.d. 5 μm particle size) column was described and validated for quantitation of Zaleplon using acetonitrile:deionised water (35:65, v/v) as a mobile phase using Paracetamol as internal standard and a flow rate of 1.5 ml min −1 with UV detection of the effluent at 232 nm at ambient temperature over a concentration range of 2–20 μg ml −1 with mean percentage recovery 100.19 ± 1.15%. The insignificance difference of the proposed methods results with those of the reference one proved their accuracy and precision.

Stability-Indicating Methods for the Determination of Rofecoxib

Two stability indicating methods have been developed for determining rofecoxib in the presence of its degradation product. The first suggested method is high performance liquid chromatography (HPLC), in which analysis is carried out using hypersil BDS C18 column (250 × 4.6 mm I.D.) with mobile Phase consisting of 0.05 M phosphate buffer (pH 3.5) and acetonitrile (70:30 v/v). A linear relationship was obtained between the detector response at 225 nm and the corresponding concentration of the studied rofecoxib in the concentration range (1–6 μg / 10 μl) with mean % recovery of 99.80 ± 0.405. The second method depends on the quantitative densitometric evaluation of thin layer chromatograms (HPTLC) with mobil phase consisting of toluene: chloroform: methanol (60: 35: 5 v/v/v) by using fluorescent high performance silica gel 60 plate. A linear relationship was obtained between peak area and the concentration of the cited drug in the range 1–6 μg/spot with a mean % recovery of 99.79 ± 0.185. The suggested methods are precise, accurate, rapid and prove their specificity in the presence of its degradation products. Both procedures are successfully applied to determine the drug in the presence of its degradation product and also in their pharmaceutical formulations.

Stability indicating methods for the determination of aceclofenac

Five new selective, precise and accurate methods for the determination of aceclofenac in the presence of its degradation product; diclofenac are described. Method A utilizes third derivative spectrophotometry at 242 nm. Method B is RSD 1 spectrophotometric method based on the simultaneous use of the first derivative of ratio spectra and measurement at 245 nm. Method C is a pH-induced difference (Δ A) spectrophotometry using UV measurement at 273 nm. Method D is a spectrodensitometric one, which depends on the quantitative densitometric evaluation of thin layer chromatogram of aceclofenac at 275 nm. Method E is RP-HPLC that depends on using methanol: water (60:40 v/v) as mobile phase at a flow rate of 1 ml/min and UV detection at 275 nm. Regression analysis of a beer's plot showed good correlation in the concentration ranges 5–40, 10–40, 15–50, 50–200, 1–50 μg/ml for methods A, B, C, D and E, respectively. These methods are suitable as stability indicating methods for the determination of aceclofenac in presence of its main degradation product, diclofenac. The proposed methods were applied for the analysis of the drug in its pharmaceutical formulation and the results obtained were compared with those obtained with the official B.P. method.

Simultaneous determination of terbinafine HCL and triamcinolone acetonide by UV derivative spectrophotometry and spectrodensitometry

A binary mixture of terbinafine hydrochloride and triamcinolone acetonide was determined by three different methods. The first one concerned with determination of both drugs using first derivative (D 1) spectrophotometric technique at 297 and 274 nm over concentration ranges of 5–30 and 4–24 μg ml −1, with mean percentage accuracies 99.90±0.67 and 100.25±0.49, respectively. The second method depends on ratio-spectra 1st derivative (RSD 1) spectrophotometry at 298 and 248 nm over the same concentration ranges with mean percentage accuracies 100.22±0.51 and 99.93±0.56, respectively. The spectrodensitometric analysis provides a rapid and precise method for the separation and quantitation of both terbinafine hydrochloride and triamcinolone acetonide. The method depends on the quantitative densitometric evaluation of thin layer chromatogram of terbinafine hydrochloride and triamcinolone acetonide at 283 and 238 nm over concentration ranges of 5–25 and 2.5–22.5 μg spot −1, with mean percentage accuracies 100.66±0.51 and 100.27±0.73, respectively. The suggested procedures were checked using laboratory prepared mixtures and were successfully applied for the analysis of their pharmaceutical preparations. The three methods retained their accuracy and precision when applying the standard addition technique. The results obtained by applying the proposed methods were statistically analysed and compared with those obtained by a reported method.

SPECTROPHOTOMETRIC AND SPECTRODENSITOMETRIC DETERMINATION OF CARBIMAZOLE

Two selective, sensitive and reproducible methods for the determination of carbimazole are described. The spectrophotometric determination of carbimazole was achieved through its reaction with a known excess of potassium bromate in bromide solution, then residual reagent was treated with sodium fluorescein in buffered medium at pH 5.5, where equivalent eosin is produced, which absorbed maximally at 517 nm. Stoichiometric study of the reaction showed that, carbimazole reacts with potassium bromate in the ratio of 1:1. The spectrophotometric method is linear over a range of 30–110 μg%. The spectrodensitometric analysis provides a rapid and precise method for the separation and quantitation of carbimazole. The method depends on the quantitative densitometric evaluation of thin layer chromatogram of carbimazole at 291 nm. It determines the drug in concentration range of 5–22.5 μg per spot. The two methods retained their accuracy and precision when applying the standard addition technique. The results obtained by applying the proposed methods were statistically analysed and compared with those obtained by the official B.P. method.

Determination of Phenytoin, Phenobarbitone, and Methyl Phenobarbitone in Combination

This work is concerned with the determination of a mixture of phenytoin, phenobarbitone and methylphenobarbitone by two different techniques. The first one depends on the simultaneous quantitative densitometric evaluation of thin layer chromatograms of the three drugs. The second is the application of quantitative 1H-NMR spectrometry for the simultaneous determination of phenobarbitone and methylphenobarbitone in the presence of phenytoin, while phenytoin was determined in this mixture by modifying the benzophenone spectrophotometric procedures.(1—4) The proposed procedures were proved using laboratory prepared mixtures of the three drugs and were successfully applied for their analysis in tablet form. The validity of the proposed procedures was further assessed by applying the standard addition technique.

Simultaneous determination of fluocortolone, fluocortolone caproate and nicotinic acid benzylester in pharmaceutical preparations by TLC

A direct quantitative thin-layer Chromatographic method for the determination of fluocortolone, fluocortolone caproate and nicotinic acid benzylester in the presence of each other in pharmaceutical preparations is described. The active ingredients were extracted from the preparations with an electronically controlled extraction apparatus within 15 min. Development of thin-layer chromatograms was carried out on silica gel 60 F254 precoated plates. All three active ingredients can be separated on one plate using one solvent system namely diethyl ether — hexane — benzene (75∶25∶1) and determined directly by the remission method using a densitometer. Fluocortolone and fluocortolone caproate were measured at 243 nm and nicotinic acid benzylester at 263 nm. Evaluation of thinlayer chromatograms takes place on-line from the linear calibration curves using an IBM 1800 computer. The described method is suitable for analyses of these substances in pharmaceutical preparations, such as ointments and creams and can be well reproduced with a coefficient of variation between 1.3–3.5% for the three active ingredients.

Dünnschichtchromatographische trennung mehrerer wirkstoffe aus salben und suppositorien und ihre anschliessende direkte quantitative analyse nach der remissionsmethode

Thin-layer chromatography of active compounds from ointments and suppositories followed by direct quantitative analysis by remissionThe paper describes a method for simultaneous thin-layer chromatographic separation of hydrocortisone, hydrocortisone acetate or hydrocortisone caproate alongside dibucaine hydrochloride, hexachlorophene and clemizole undecylate as well as clemizole hexachlorophenate in ointments and suppositories. Development of thin-layer chromatograms is carried out on silica gel 60 F-254 pre-coated plates. All four active ingredients can be separated on one silica gel plate using one solvent system and determined directly by the remission method using a densitometer. Hydrocortisone and its two esters are measured at 248 nm, dibucaine hydrochloride at 325 nm, hexachlorophene at 300 nm, and clemizole undecylate as well as clemizole hexachlorophenate at 275 nm. Evaluation of thin-layer chromatograms takes place on-line from a linear calibration curve using an IBM 1800 computer. The described method is very suitable for analyses of these active ingredients in drug forms, such as ointments or suppositories and is reproducible with coefficients of variation of 1.29–3.56%.

Direct Quantitative Photometry on Thin Layer Chromatograms

AbstractThe principal methods for the direct photometric evaluation of thin layer chromatograms are discussed with the aid of examples. Since light is strongly scattered by TLC adsorbents, absorption measurements in reflection are preferable to transmission measurements. In the reflection measurements, an approximation method for the Kubelka‐Munk function leads to satisfactory straight calibration lines. The working range for quantitative reflection measurements is in the μg range, and the detection limit is of the order of 10−8g. The standard deviation for spots of equal concentration is less than ±1%. The signal‐to‐background ratio is improved by simultaneous measurement of reflection and transmission. The measurement of diminution in fluorescence (“fluorescence quenching”) is found to be less sensitive than the reflection at the absorption maximum. However, combined measurement of the reflection and the diminution of fluorescence gives an increase in sensitivity as compared to pure reflection measurements. As in solution photometry, fluorescence measurements are generally much more sensitive than absorption measurements. The working range is normally in the ng region, and the detection limit is around 10−11g. All the investigations described in this paper were carried out on the Zeiss chromatogram spectrophotometer.

Microphotometric determination of 1-dimethylaminonaphthalene-5-sulfonylamino acids with glycine as an example

1. A method for the quantitative evaluation of thin-layer chromatograms of dansylglycine by photographing them and microphotometering the negative obtained has been proposed.