A simple, rapid, and stability-indicating RPTLC method has been established and validated for quantitative analysis of two oral antidiabetic agents, the sulfonylureas gliclazide and glipizide, in tablets. Analysis was performed on RP-18 plates with 60% (
) acetonitrile in pH 2.3 phosphate buffer as mobile phase. Compact spots were obtained for gliclazide (
0.38 ± 0.02, mean ± SD) and glipizide (
0.51 ± 0.03). Detection and quantification were performed by classical densitometry at 215 nm, the wavelength of maximum absorption of gliclazide and glipizide. Calibration plots were constructed in the range 0.8–1.8 μg per 10 μL (per spot) for both drugs and were linear with good correlation coefficients (
= 0.998 ± 0.001, mean ± SD, for gliclazide and
= 0.993 ± 0.002 for glipizide). The limits of detection and quantification were 50 and 200 ng per spot, respectively, for gliclazide and 60 and 300 ng for glipizide. The method was validated for robustness, precision, accuracy, stability, and specificity. The drugs were subjected to acidic and alkaline hydrolysis, oxidation, photo-degradation, and heating and were found to be susceptible to some stress conditions. However, the degradation products were well resolved from the pure drugs with significantly different
Authors:Anna Gumieniczek, Tadeusz Inglot and Anna Kończak
Classical densitometry and videoscanning were compared for a new TLC method of quantitative analysis of two AT1 receptor antagonists, candesartan and losartan, in pharmaceuticals. Chromatography was performed on silica gel with 1,4-dioxane-hexane-99% formic acid 5:5:0.1 (ν/ν) as mobile phase. Classical densitometry was performed at the wavelengths of maximum absorption of candesartan (258 nm) and losartan (243 nm) whereas videoscanning was performed at 254 nm for both drugs. Compact spots were obtained for candesartan (RF 0.47 ± 0.01, mean ± SD) and losartan (RF 0.35 ± 0.01). Calibration plots were constructed in the range 0.2–1.4 μg per band for the both drugs and were linear with good correlation coefficients — 0.9997 and 0.9981 for candesartan, and 0.9986 and 0.9982 for losartan, for densitometry and videoscanning, respectively. The methods were validated for robustness, precision, accuracy, and specificity in relation to excipients present in the respective formulations. Finally, the methods were compared statistically in respect of robustness, precision and accuracy.
Authors:Anna Berecka, Anna Gumieniczek and Hanna Hopkała
The thin-layer chromatographic behavior of three of the newest oral antidiabetic agents, pioglitazone, rosiglitazone, and repaglinide, has been studied in reversed-phase systems. Chromatography was performed on RP-8 as adsorbent with buffer-organic modifier binary mobile phases of widely different composition. Phosphate buffers of pH 2.4, 4.4, 6.0 and 7.9 were used with three organic mobile-phase modifiers, acetonitrile, 2-propanol, and methanol; the concentration of organic modifier was varied between 20 and 80% (
). Plates were developed in horizontal chambers, visualized by UV illumination at
= 254 nm, and scanned with a densitometer. The effect of the mobile phase on retention was studied. The selectivity of the chromatographic systems is discussed. The linearity of relationships between
and modifier volume fraction, molar fraction, and logarithm of the molar fraction was calculated.
Authors:Anna Gumieniczek, Anna Berecka and Hanna Hopkała
A simple, rapid, and stability-indicating thin-layer chromatographic method has been developed for quantitative determination of repaglinide in tablets. Analysis was performed on RP-8 TLC plates with acetonitrile-pH 6.0 phosphate buffer, 60 + 40 (%
), as mobile phase. Detection and quantification were performed by classical densitometry at the wavelength of maximum absorption of repaglinide, 225 nm. A calibration plot constructed in the range 0.6–3.6 μg/10 μL was linear with a good correlation coefficient (
= 0.998 ± 0.001, mean ±
= 5). Limits of quantitation and detection of repaglinide were 0.27 μg/10 μL and 0.08 μg/10 μL, respectively. Instrumental precision established at three concentrations of the drug ranged from 3.92 to 0.97% for the lowest and highest concentrations of repaglinide, respectively. The mean intra-day and inter-day variability, including three concentrations of repaglinide, were 1.93 and 2.25% (
= 9), respectively. Recovery from model mixtures, at three levels of addition, ranged from 103.06 to 102.49% for the lowest and highest levels, respectively. Total mean ±
recovery was 102.71 ± 2.04% (
= 15). The mean ±
recovery from commercially available tablets was 101.85 ± 1.83% (
= 10). The effect of pH, temperature, and UV light on degradation of repaglinide was also investigated. The analytical method presented was found to be simple, reliable, and convenient for routine pharmaceutical analysis. Its analytical performance fulfilled acceptance criteria established for TLC methods in the official literature.
Authors:Anna Gumieniczek, Anna Berecka, Dariusz Matosiuk and Hanna Hopkała
Five anti-diabetic thiazolidinediones have been chromatographed on C
TLC plates with binary mobile phases containing water and the organic modifiers acetone, 1,4-dioxane, or methanol. Linear relationships were obtained between the
values of the compounds and the concentration of organic modifier in the mobile phase. These
values enabled calculation of
values by extrapolation. Calibration equations were then obtained for nine standards of known lipophilicity in the range 0.83–6.04. From these equations the partition coefficients log
were calculated for the drugs. Lipophilicity values were also calculated by use of computational methods. Finally, interesting relationships were obtained between experimental and theoretical log
values and pharmacological data from the literature.
Authors:Anna Gumieniczek, Hanna Hopkała, Anna Berecka and Dorota Kowalczuk
The chromatographic behavior of seven oral antidiabetic drugs — chlorpropamide, tolbutamide, glibenclamide, metformin, pioglitazone, rosiglitazone, and repaglinide — has been investigated. Normal-phase chromatography was performed on silica gel and alumina layers with mixtures of chloroform, diethyl ether, and ethyl acetate as mobile phases. For more effective resolution aqueous ammonia or acetic acid was added to the mobile phases. Silica gel enabled better separation than alumina. Reversed-phase chromatography was performed on octadecyl-bonded silica gel (RP-18) with mixtures of acetonitrile or 2-propanol with phosphate buffer as mobile phases. The effect of pH on the separation of the drugs was also examined. For separation of these drugs reversed-phase chromatography was more effective than use of normal-phase mode.
Authors:Łukasz Komsta, Patrycja Smardz, Rafał Pietraś, Robert Skibiński and Anna Gumieniczek
The retention (RM) values of nine one-point adsorption model compounds: diphenylamine, indol, 2-naphtol, 1-naphtol, 1-naphtylamine, 4-toluidine, carbazole, 4-chloraniline, and thymol were investigated on silica gel using six modifiers: acetone, dioxane, hexane, isopropanol, methylethylketone, ethyl acetate, and tetrahydrofurane (in hexane). These compounds showed small but visible curvilinearity of dependence of RMvs. modifier concentration. This curvilinearity is very similar among the investigated compounds, so relative differences of extrapolated RM are almost the same (strictly intercorrelated) regardless of the regression technique used. We have compared several robust and weighted regression methods and investigated their impact on extrapolated values. It can be concluded that one should primarily consider weighted regression with 1/x weights during retention extrapolation. It seems to be a better alternative than classical regression (better extrapolation) and also better than polynomial approaches (better stability).
Authors:Łukasz Komsta, Rafał Pietraś, Ewelina Bartuzi, Robert Skibiński and Anna Gumieniczek
Thin-layer chromatography (TLC) on two adsorbents (RP18 and CN) and with six modifiers (acetonitrile, acetone, dioxane, propan 2-ol, methanol, and tetrahydrofurane), followed by classical RM value extrapolation (previous results), was chemometrically compared with new one-run gradient high-performance liquid chromatography (HPLC) (C18, C18e, CN, and DIOL columns, acetonitrile, and methanol as modifiers) and, additionally, with seven computational algorithms (ALOGPs, AClogP, ALOGP, MLOGP, KOWWIN, XLOGP2, and XLOGP3) as a lipophilicity assessment tool on 35 model compounds with known lipophilicity. The statistical significance of intercepts and slopes of Collander equation (log P — retention dependence) and their values were compared. Whole results data set was subjected to scaled principal component analysis, which allowed exploring two main trends in these data. It can be concluded that one-run gradient HPLC does not outperform TLC in lipophilicity determination. Very good correlations were obtained between real log P and computational approaches; however, this is not a surprise for such simple molecules.
Authors:Łukasz Komsta, Krzysztof Sztanke, Paulina Mączka, Marzena Ucherek, Robert Skibiński and Anna Gumieniczek
The lipophilicity, log
, of 27 novel imidazo[2,1-
][1,2,4]triazine derivatives with strong biological activity has been determined by reversed-phase thin-layer chromatography. The compounds were chromatographed on RP18 plates with methanol-water binary mobile phases containing different proportions of methanol.
values for pure water were then extrapolated from the linear
equation. Twelve compounds with known literature log
values acted as a reference calibration data set for computation of experimental log
values. The results obtained were compared, by principal-components analysis, with those obtained by use of nine computational methods.