Use In Quality Control Laboratories Research Articles

Development and validation of a sensitive and robust chiral method for (R)-propylene oxide: From chiral gas chromatography to chiral liquid chromatography with pre-column derivatization

Propylene oxide (PO) is an important chemical commodity and a chiral building block to prepare pharmaceutical compounds. It is crucial to develop a sensitive and robust chiral method that can be used to assess the enantiomeric purity of PO and safeguard the quality and safety of pharmaceutical products in commercial quality control (QC) laboratories. A chiral gas chromatography (GC) method based on direct enantioseparation of PO enantiomers on a 50-meter Chiraldex™ A-TA open capillary column was successfully developed for determination of enantiomeric impurity (S)-PO in (R)-PO. However, it was revealed that the chiral GC method lacked adequate robustness for routine use. Consequently, a chiral high-performance liquid chromatography (HPLC) method was developed through pre-column derivatization of PO with 2-naphthalenethiol followed by enantioseparation on a Lux Cellulose-1 (250 mm × 4.6 mm, 5 µm) column. The chiral HPLC method has been demonstrated excellent specificity, sensitivity, accuracy and robustness through method validation and robustness evaluation, thus establishing its suitability for routine use in QC laboratories.

Greenness-sustainability metrics for assessment smart-chemometric spectrophotometric strategy for evaluation of the combination of six gastric proton-pump inhibitors with two selected impurities

Green analytical approaches are employed for the determination of active pharmaceutical ingredients, in conjunction with their impurities. Smart chemometric spectrophotometric techniques, including orthogonal partial least square (OPLS), variable selection such as genetic algorithm (GA-OPLS), and interval selection (i-OPLS), were utilized. These chemometric models were implemented for assessing six proton-pump inhibitors Omeprazole, Esomeprazole, Lansoprazole, Pantoprazole, Rabeprazole, and Dexlansoprazole along with two selected official impurities, namely 4-Desmethoxy omeprazole impurity and Rabeprazole-impurity B. Experimental design was implemented to separate impurities, in the process of multivariate calibration, a five-level eight-factor calibration design consisting of 25 samples was selected. This design was deliberately selected to guarantee that the components were mutually orthogonal to assess the model's performance and reliability, a separate validation set of 15 samples was constructed. The best-performing of the proposed techniques were identified by considering the least favorable values of the Correlation Coefficient (R ≥ 0.9995), the Root Mean Square Error of Prediction (RMSEP) values between (0.0102–0.5622), and the Relative Error of Prediction (REP) values between (0.2961–1.1917). The proposed and reported methods' greenness-sustainability was quantitatively evaluated, and a comparative study of the greenness profile was established through a spider chart, the National Environmental Method Index tool, advanced and modified NEMI along with the Hexagon tool, and the whiteness qualities of the presented approaches were assessed by implementing the recently adopted Red-Green-Blue paradigm and White Analytical Chemistry tool. These approaches are well-suited for use in quality control laboratories due to their observed acceptance, long-term sustainability, simplicity, and affordability.

Formation and characterization of charge transfer complex between alectinib and 2,3-dichloro-5,6-dicyano-1,4-benzoquinone: Application to development of microwell spectrophotometric method

Alectinib (ALC) is a novel highly selective anaplastic lymphoma kinase (ALK) inhibitor that is effective in the treatment of patients with advanced ALK-positive non-small cell lung cancer and lymphomas. This study, describes, for the first time, the formation of a charge transfer complex (CTC) between ALC with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ). The CTC was characterized using different spectrophotometric and computational techniques. UV–visible spectrophotometry ascertained the formation of the CTC in methanol via the formation of a new broad absorption band with a maximum absorption peak (λmax) at 505 nm. The molar absorptivity (ε) of the complex was 2.72 × 103 L mol−1 cm−1 and its band gap energy was 1.94 eV. The stoichiometric ratio of ALC:DDQ was found to be 1:1. The association constant of the complex was 9.6 × 102 L mol−1, and its standard free energy was −16.98 J mol−1. Computational calculations were conducted using different calculation tools. The computed geometrical parameters, specifically bond lengths, exhibited variations in the CTC compared to the individual reactants (ALC and DDQ). The Mulliken atomic charge analysis indicated increased values for DDQ (the acceptor) and decreased values for ALC (the donor) in the CTC, confirming the formation of a CTC. The reaction was employed as a basis for a novel microwell spectrophotometric method for the quantification of ALC in its bulk form, capsule content, and uniformity testing. This greenness and high throughput of the method was documented. The method represents a valuable tool for routine use in quality control laboratories for analysis of ALC's bulk form and capsules.

Environmentally benign first derivative synchronous spectrofluorimetry for the analysis of two binary mixtures containing duloxetine with avanafil or tadalafil in spiked plasma samples.

Antidepressants can cause sexual dysfunction side effects, necessitating the co-administration of phosphodiesterase type 5 inhibitors. The simultaneous determination of these drugs in biological fluids is critical for therapeutic drug monitoring. For the first time, two binary mixtures containing duloxetine with either avanafil or tadalafil were estimated utilizing simple green spectrofluorimetric methods without the need for a previous separation step. The study was based on first derivative synchronous spectrofluorimetry in ethanol using a change in wavelength difference (∆λ) of 20 and 25 nm for the first and second combinations, respectively. Duloxetine and avanafil were estimated at 297.7 and 331 nm in their binary mixture, while duloxetine and tadalafil were determined at 290.3 and 297.7nm, respectively. The linearity was achieved over the ranges of 0.1-1.5μg mL-1 for both duloxetine and avanafil and 0.01-0.40 μg mL-1 for tadalafil, with limits of detection of 0.013, 0.022, and 0.004 μg mL-1 for duloxetine, avanafil, and tadalafil, respectively. Successful application of the developed approaches was accomplished for the estimation of the two mixtures in dosage forms as well as human plasma with excellent percentage recoveries (96-103.75% in plasma), which supports their suitability for use in quality control laboratories and pharmacokinetic studies. Moreover, the adopted approaches' greenness was evidenced by applying three tools.

An effective ultra‐performance liquid chromatography and derivatization method for the quantification of potential genotoxic impurity Hydrazine in Gliclazide and its formulation – Robustness study by the design of experiments

AbstractThe current study presents an accurate, precise, and highly selective ultra‐performance liquid chromatography derivatization method for the quantification of Hydrazine in Gliclazide and its formulations. Gliclazide, derivatizing agent‐ 3,4,5‐trimethoxy benzaldehyde, and Hydrazine derivatives were separated on YMC Pack pro C18 column (50 × 3.0 mm id, 2 μm), column oven temperature maintained at 40°C, with flow rate 0.6 ml/min on ultra‐performance liquid chromatography system with gradient elution monitored at 333 nm. 0.1% trifluoroacetic acid and acetonitrile were used as mobile phase‐A and mobile phase‐B. Evaluated the robustness of the optimized method by employing a quality‐by‐design‐based design of experiments. The method was successfully validated as per the International Conference on Harmonization Q2 (R1) guideline and the United States Pharmacopeia < 1225 > general chapter. The United States Pharmacopeia resolution between the Gliclazide and Hydrazine derivative was 3.6. The limit of detection and limit of quantitation for Hydrazine derivative are 0.10 μg/ml and 0.30 μg/ml, respectively. The obtained regression coefficient (R2) was > 0.9998, and the %recoveries from the limit of quantitation to 150% level were 96.9%–100.8% from the linearity and accuracy studies, respectively. The method validation data and quality‐by‐design‐based robustness study indicate that the developed derivatization method is suitable for routine use in Quality Control laboratories.

VISIBLE SPECTROPHOTOMETRIC METHODS FOR THE ESTIMATION OF MODAFINIL IN PHARMACEUTICAL FORMULATION

Two simple, sensitive, and fast visible spectrophotometric techniques for quantifying modafinil in marketed dosage form have been proposed. The method A was focussed on the complexation involving charge transfer with picric acid to give pale yellow colored chromogen which shows absorption maxima at 450 nm. In the concentration range of 5-40 μg mL-1, Beer’s law is obeyed with a correlation value of 0.999. The method B depends on the oxidative coupling process with MBTH in the presence of FeCl3 in acidic pH to give green colour chromogen which shows absorption maxima at 657 nm. In the concentration range of 10-50 μg mL-1, Beer’s law was obeyed with a correlation value of 0.999. The developed methodologies have been statistically verified for use in quality control laboratories for pharmaceuticals

A New Strategy for the Software-Assisted LC Separations of Ketoconazole and Its Impurities.

Analytical quality by design and the use of dissimilar chromatographic systems can be employed to accelerate chromatographic separations. Herein, a software (S-Matrix)-assisted platform was used to proficiently screen, optimize and select the optimal parameters for the chromatographic separation of ketoconazole and its related impurities. This approach evaluated the various chromatographic parameters in a stepwise manner based on the statistical tools and provided an in-depth understanding of the critical parameters influencing the peak selectivities and separations. It was demonstrated that dissimilar conditions, such as different stationary phases, mobile phase pH and organic modifiers (i.e., critical method variables), can improve the peak resolution, while the critical quality attributes can provide conditions appropriate for quantification purposes via a quality target analytical method. Furthermore, an orthogonal method was established to support the primary method. The orthogonality between the two methods was defined by the correlation matrix between the two systems using the Pearson correlation coefficient and was found to be 0.12. Using the optimized method, the primary method was validated as per International Council for Harmonization in the range of 0.05-1.0% for impurities and 80.0-120.0% for ketoconazole, thereby indicating the suitability of the method for use in quality control laboratories.

RP-HPLC Method Development and Validation for the Estimation of Sacubitril and Valsartan in Pharmaceutical Dosage Form

A new, economical, simple, accurate, and precise RP-HPLC method was developed for simultaneous assay and content uniformity determination of Sacubitril and Valsartan in bulk and pharmaceutical dosage form. The separation of Sacubitril and Valsartan was achieved within 6 minutes on Phenomenex Luna C18 250 mm x 4.6mm and 5µm Particle Size, column using Acetonitrile: Methanol: Water (30:55:15% v/v/v) as the mobile phase. Detection was carried out at 250 nm wavelength. The retention time of Sacubitril and Valsartan was found to be 2.361 and 3.304 min, respectively. The validation of the developed method was performed in terms of specificity, accuracy, precision, linearity, the limit of detection, the limit of quantification as mentioned in International Conference on Harmonization (ICH) guidelines. The method showed adequate sensitivity concerning linearity, accuracy, and precision over the range 12-36 μg/ml and 13-39 μg/ml for Sacubitril and Valsartan, respectively. The percentage recoveries obtained for Sacubitril and Valsartan were found to be in the range of 98.00 – 102.00 %. The proposed method is suitable for use in quality-control laboratories for quantitative analysis.

Stability indicating HPLC-Fluorescence detection method for the simultaneous determination of linagliptin and empagliflozin in their combined pharmaceutical preparation

A sensitive, accurate, and precise liquid chromatographic method has been developed and validated for the determination of Linagliptin (LNG) and Empagliflozin (EMP) in their combined tablets. Chromatographic separation was carried out on ODS-3 Inertsil® C18 column (150×4.6 mm, 5 µm). The mobile phase A (consisting of 0.30% Triethyl amine buffer (TEA) at pH = 4.5, adjusted using ortho-phosphoric acid); the mobile phase B (consisting of acetonitrile) was pumped through the column whose temperature was maintained at 40 °C, with a flow rate 1.7 mL/min, using gradient elution from 0-3 min A:B (75:25, v:v), then from 3-6 min the ratio changed to be A:B (60:40, v:v). Fluorescence detection (FLD) was performed at 410 nm after excitation at 239 nm. Acceptable linearity, accuracy and precision values of the proposed method were found over the concentration ranges of 0.5-15 µg/mL for LNG and 1.0-30 µg/mL for EMP with correlation coefficients of 0.9997 and 0.9998 in the case of LNG and EMP, respectively. The recoveries and relative standard deviations percentages were found in the following ranges: 98.56-101.85 and 0.53-1.52% for LNG and 98.00-101.95 and 0.31-1.05% for EMP. The detection and quantification limits were 0.15 and 0.45 µg/mL for LNG and 0.22 and 0.67 µg/mL for EMP. The optimized method was validated and proved to be specific, robust, accurate and reliable for the determination of the drugs in pure form or in their combined pharmaceutical preparations. No significant difference was found regarding accuracy and precision upon statistical comparison between the obtained results of the proposed method and those of the reported method. Furthermore, the proposed method is proved to be a stability-indicating assay after exposure of the studied drugs to variable forced degradation parameters, such as acidic, alkaline and oxidative conditions, according to the recommendations of the International Conference on Harmonization guidelines. The simplicity and selectivity of the proposed method allows its use in quality control laboratories.

The first spectrofluorimetric approach for quantification of colistin sulfate and its prodrug colistimethate sodium in pharmaceutical dosage form and human plasma.

A new, accurate, nonextractive, and sensitive fluorimetric approach was proposed and validated for the first time estimation of colistin sulfate and its inactive prodrug colistimethate sodium in its bulk form, pharmaceutical formulations, and human plasma. The approach relied on condensation between acetylacetone/formaldehyde and the primary amino moiety of nonfluorescent colistin in Teorell and Stenhagen buffer (pH 2.8) by the Hantzsch reaction to form a highly fluorescent dihydropyridine derivative. The fluorescent product was measured at 460 nm (λex = 402 nm). A plot of relative fluorescence intensity (RFI) versus concentration was rectilinear over the range 200-4000 ng ml-1 with excellent correlation (r) and determination (r2 ) coefficients of 0.9999 and 0.9998, respectively. The limit of detection (LOD) and limit of quantitation (LOQ) were 40.91 and 123.99 ng ml-1 , respectively. The present procedure was useful for determination of colistin sulfate either in powder form for suspension or in its parenteral prodrug colistimethate sodium in vial formulation. The investigated approach was applied for in vitro quantification of this drug in spiked human plasma, with a per cent mean recovery of 98.24 ± 1.34. The proposed method is reliable, selective, and does not require tedious sample pretreatment steps, expensive instrumentation, or harmful reagents, all of which make it ideally suited for use in quality control laboratories.

An Improved Reversed-Phase High-Performance Liquid Chromatography Method for the Analysis of Related Substances of Prednisolone in Active Ingredient.

Prednisolone, an important active pharmaceutical ingredient, is a synthetic glucocorticoid used for the preparation of various pharmaceutical products with anti-inflammatory and immunosuppressive properties. It is a challenge in high-performance liquid chromatography (HPLC) to separate the prednisolone peak and its structurally related substance (hydrocortisone), which only differs in a double bond at the C-1 position. Successful application of the HPLC method according to the European Pharmacopoeia monograph for related substances of prednisolone is very often limited to the chromatographic system available. This is due to the nonbaseline separation of the prednisolone and hydrocortisone peaks, which is strongly influenced by the instrument parameters and the chosen C18 column. First, an adjusted European Pharmacopoeia method for related substances of prednisolone was developed within the allowable adjustments. Next, an improved stability-indicating reversed-phase HPLC method for related substances of prednisolone was developed and validated for use in quality control laboratories for routine analysis. The optimized separation was performed on a Phenomenex Gemini C18 column (150 mm × 4.6 mm, 3 μm) using a gradient mobile-phase system consisting of acetonitrile/tetrahydrofuran/water (15:10:75 v/v/v), acetonitrile/water (80:20 v/v), and ultraviolet detection at 254 nm. A baseline separation was achieved, and stability indicating capability was demonstrated by a forced degradation study. A full validation procedure was performed in accordance with International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use guidelines.

Novel Analytical Study For The Charge-Transfer Reactions Of Omeprazole With 2,3-Dichloro-Naphthoquinone And 2,3,5,6-Tetrabromo- 1,4-Benzoquinone: Application For The Development Of Microwell Assay Of Omeprazole

Novel analytical study was performed in order to develop and validate new high-throughput microwell-based spectrophotometric assays for determination of omeprazole (OMZ) in its pharmaceutical formulations. The proposed assays were based on the charge-transfer (CT) reaction of OMZ with 2,3-dichloronaphthoquinone (DCNQ) and 2,3,5,6-tetrabromo-1,4-benzo-quinone (BROM). In the present study, the CT reactions was carried out in microwell plates as reaction vessels in order to increase the automation of the assays and the efficiency of its use in quality control laboratories (QCLs). All factors affecting the CT reactions were carefully studied, and the conditions were optimized. Kinetics and stoichiometry of the CT reactions were investigated, and the mechanism was postulated. Activation energy of the CT reactions was determined and found to be 13.87 and 16.27 Kcal mol−1 for the reaction of OMZ with DCNQ and BROM, respectively. The initial rate and fixed time methods were applied for generating the calibration graphs for determination of OMZ concentrations. Under the optimum conditions, the linear range was 0.145 – 1.45 x 10-4 and 1.45 – 7.25 x 10-4 M with LOD of 0.6 and 6.0 microgram ml-1 for DCNQ and BROM, respectively. Analytical performance of the proposed methods, in terms of accuracy and precision, was statistically validated and the results were satisfactory; RSD was <2.8% for both repeatability and reproducibility. The proposed methods were successfully applied to the analysis of OMZ in its dosage forms and the recovery results (98.64 – 100.6 ± 0.25 -2.74 %) were comparable with those of the reported method. The developed method may provide a safer and economic tool for the analysis of OMZ in QCLs.

<b>Development and validation of analytical methodology for quantification of propranolol hydrochloride in a multiparticulate biphasic system by UV-vis spectrophotometry

Hypertension is one of the most important mortality risks in the world and is considered the leading cause of deaths associated with cardiovascular disease. In an attempt to improve the treatments and apply them to a large number of disorders, a propranolol hydrochloride (Prop) based medicine it was developed in our laboratory, employing a multiparticulate system which operates in a biphasic manner, one portion of immediate and other of modified release. This study aimed at developing and validating a suitable method to quantify this new version of the drug using UV spectrophotometry, following what is recommended in RE 899/2003. The results showed that the method employed was linear, accurate, precise and robust in the range of 0.8 to 96.0 μg mL -1 , which allows it for routine use in quality control laboratories.

Rapid micellar HPLC analysis of loratadine and its major metabolite desloratadine in nano-concentration range using monolithic column and fluorometric detection: application to pharmaceuticals and biological fluids

BackgroundLoratadine is a commonly used selective non-sedating antihistaminic drug. Desloratadine is the active metabolite of loratadine and, in addition, a potential impurity in loratadine bulk powder stated by the United States Pharmacopeia as a related substance of loratadine. Published methods for the determination of both analytes suffer from limited throughput due to the time-consuming steps and tedious extraction procedures needed for the analysis of biological samples. Therefore, there is a strong demand to develop a simple rapid and sensitive analytical method that can detect and quantitate both analytes in pharmaceutical preparations and biological fluids without prior sample extraction steps.ResultsA highly-sensitive and time-saving micellar liquid chromatographic method is developed for the simultaneous determination of loratadine and desloratadine. The proposed method is the first analytical method for the determination of this mixture using a monolithic column with a mobile phase composed of 0.15 M sodium dodecyl sulfate, 10% n-Butanol and 0.3% triethylamine in 0.02 M phosphoric acid, adjusted to pH 3.5 and pumped at a flow rate of 1.2 mL/min. The eluted analytes are monitored with fluorescence detection at 440 nm after excitation at 280 nm. The developed method is linear over the concentration range of 20.0–200.0 ng/mL for both analytes. The method detection limits are 15.0 and 13.0 ng/mL and the limits of quantification are 20.0 and 18.0 ng/mL for loratadine and desloratadine, respectively. Validation of the developed method reveals an accuracy of higher than 97% and intra- and inter-day precisions with relative standard deviations not exceeding 2%.ConclusionsThe method can be successfully applied to the determination of both analytes in various matrices including pharmaceutical preparations, human urine, plasma and breast milk samples with a run-time of less than 5 min and without prior extraction procedures. The method is ideally suited for use in quality control laboratories. Moreover, it could be a simple time-saving alternative to the official pharmacopeial method for testing desloratadine as a potential impurity in loratadine bulk powder.Graphical abstractTypical chromatogram of loratadine and its major metabolite desloratadine using the proposed micellar HPLC method

A Validated High-Throughput Fluorometric Method for Determination of Omeprazole in Quality Control Laboratory via Charge Transfer Sensitized Fluorescence.

A high-throughput 96-microwell plate fluorometric method was developed and validated to determine omeprazole (OMZ) in its dosage forms. The method was based on the charge-transfer (CT) sensitized fluorescence reaction of OMZ with 2, 3-dichloro-5, 6-dicyano-1, 4-benzoquinone (DDQ). This fluorescence reaction provided a new approach for simple, sensitive and selective determinations of OMZ in pharmaceutical preparations. In the present method, the fluorescence reaction was carried out in 96-microwell plates as reaction vessels in order to increase the automation of the methodology and the efficiency of its use in quality control laboratories. All factors affecting the fluorescence reaction were carefully studied and the conditions were optimized. The stoichiometry of the fluorescence reaction between OMZ and DDQ was determined and the reaction mechanism was suggested. Under the optimum conditions, the linear range was 100-6000ng/ml with the lowest LOD of 33ng/ml. Analytical performance of the proposed assay, in terms of accuracy and precision, was statistically validated and the results were satisfactory; RSD was <2.6% and the accuracy was 98.6-101.6%. The method was successfully applied to the analysis of OMZ in its dosage forms; the recovery values were 98.26-99.60±0.95-2.22%. The developed methodology may provide a safer, automated and economic tool for the analysis of OMZ in quality control laboratories.

Routine Screening for the Presence of Adulteration in Raw Materials Using Automated Nuclear Magnetic Resonance Spectroscopy

In an effort to increase the security of the supply chain for raw materials used in the manufacture of human therapeutics, a routine screen to detect the presence of adulteration using fully automated nuclear magnetic resonance spectroscopy has been developed and qualified for use in quality control laboratories. The method involves the collection of one-dimensional (1)H and (13)C spectra, which are subsequently processed to identify and quantitate raw material constituents by comparison to a spectral database. The resulting method is an easy-to-use limit test that can automatically determine the integrity of incoming raw materials. The method is intended to be used in good manufacturing practice production facilities and is suitable for excipients and aqueous soluble raw materials used in biopharmaceutical processes. In an effort to increase the security of the supply chain for raw materials used in the manufacture of human therapeutics, a routine screen to detect the presence of adulteration using fully automated nuclear magnetic resonance (NMR) spectroscopy has been developed and qualified for use in quality control laboratories. The method involves the collection of NMR spectra, which are subsequently processed to identify and quantitate raw material constituents by comparison to a spectral database. The resulting method is an easy-to-use limit test that can automatically determine the integrity of incoming raw materials. The method is intended to be used in good manufacturing practice production facilities and is suitable for excipients and aqueous soluble raw materials used in biopharmaceutical processes.

MAPREC assay for quantitation of mutants in a recombinant flavivirus vaccine strain using near-infrared fluorescent dyes

Mutant analysis by PCR and restriction enzyme cleavage (MAPREC) is a quantitative assay of revertants in batches of live viral vaccines. The assay is highly sensitive and reliable but requires radioactive isotopes, which complicates its use in quality control laboratories. To quantify mutants in the cDNA of the West Nile (WN)/Dengue 4 chimera that was proposed as a new candidate of live vaccine against West Nile disease, alternative MAPREC protocols using non-radioactive dyes were explored. To compare the utility of different fluorescent dyes for MAPREC, the G2337→C mutation that was revealed by microarray hybridization in WN/Dengue 4 chimera virus was used as a model. DNA fragments produced by restriction endonuclease digestion were visualized in polyacrylamide gels by visible-range fluorescent dyes including ethidium bromide (EtBr) and SYBR Green I as well as by near-infrared (NIR) dye SYTO 60 and NIR dyes 700 and 800. The MAPREC assay performed with SYTO 60 and SYBR Green I was more sensitive than with EtBr but less sensitive than with NIR dyes 700 or 800. The NIR dyes 700 and 800 exhibited a wide linear range that may enable the detection of 0.05% of mutants in viral stocks. The NIR-based MAPREC assay was validated by using World Health Organization (WHO) international references for poliovirus type 3 with known contents of mutants. Values of mutant content produced by the non-radioactive assay were similar to the values determined in a previous WHO international collaborative study. The modified MAPREC assay could be used as an alternative to the radioisotope-based standard protocol for quality control of live viral vaccines.

Chiral LC for Separation of the Enantiomers of Abacavir Sulfate

A chiral liquid chromatographic method has been developed for separation of the enantiomers of abacavir sulfate, a nucleoside reverse transcriptase inhibitor for treatment of HIV. The enantiomers were resolved in a Chiralcel OD column with n-hexane–ethanol–trifluoroacetic acid, 92:8:0.1 (v/v), as mobile phase. The resolution between the enantiomers was not less than 3.5. The trifluoroacetic acid had a very important effect on the separation of enantiomers. The method was extensively validated to prove its suitability and robustness. Solution stability and mobile-phase stability were studied for 48 h and found to be stable during this period. The method was found to be suitable for use in quality-control laboratories for analysis of the commercial drug substance.

Prediction of frozen storage time of Cape hake (Merluccius capensis and Merluccius paradoxus) by instrumental methods

AbstractAn instrumental method to predict frozen storage time of Cape hake was developed by evaluation of a combination of methods previously known to classify hake into different quality categories, such as apparent viscosity, Kramer and puncture tests. The results showed that with multiple linear regression analysis it was possible to obtain linear models for the prediction of storage time at − 20°C. Only apparent viscosity (ηapp) and Kramer shear resistance (Kramer) were included in the regression model (months = 15.003 − 0.001ηapp + 0.144 Kramer; $R_{\rm {adj}}^{2} = 0.91$). The model had a typical error of estimation of 2.0 months. The simplicity of the model, which requires only two parameters, and the easiness of their measurement make the method a good candidate for use in quality control laboratories of hake processing industries. Copyright © 2006 Society of Chemical Industry

Determination of Theanine in Commercial Tea by Liquid Chromatography with Fluorescence and Diode Array Ultraviolet Detection

Two liquid chromatographic methods that involve precolumn derivatization with o-phthaladehyde (OPA) and phenylisothiocyanate (PITC) with fluorescence and diode array UV detection for the determination of theanine have been developed. The chromatographic separations were achieved by reverse-phase high-performance liquid chromatography using octadecyl columns and gradient elution. The methods were applied to evaluate the theanine content of commercial tea leaves. The coefficient of variation of the peak area repeatability for within day (n = 8) and between day (n = 8 over 10 days) was lower than 3% for both of the methods. The estimated limit of detection (LOD) and limit of quantitation (LOQ) for the OPA method was 0.12 and 0.35 microg theanine, respectively. The PITC method was 500-fold more sensitive with LOD and LOQ values of 0.25 and 0.75 ng, respectively. The theanine content of the commercial tea samples varied from 2-5 mg/g leaf. The overall % recoveries for these methods ranged from 93-99.3. The sensitivity and simplicity of the method render them suitable for use in quality control laboratories.