HPLC Method Development and Validation: A Review

Sabir, Azim Md; Mehrabani, Mitra; Bhasin, Parminder S.

doi:10.7897/2230-8407.04407

Cited by 46 publications

(25 citation statements)

References 9 publications

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“…The precision of the method was carried by performing six independent determinations.The average assay of six determinations of droxidopa was100.83 with RSD of 0.303. % RSD<2.0 indicates that the method is precise (Sabir et al, 2013). The ruggedness was veri ied by performing the chromatographic analyses of samples by various analysts on two different days.…”

Section: Methods Precision and Intermediate Precisionmentioning

confidence: 99%

Method development and validation of droxidopa by HPLC technique

Pandilla¹,

Chitra

Nalini³

et al. 2020

ijrps

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The purpose of this work is to develop and validate stability, indicating reverse phase High-performance liquid chromatography (HPLC) method for the rapid and precise determination of droxidopa in its pure form and formulations. A simple, fast, accurate and economical way has been developed and validated for the quantification of droxidopa by HPLC technique. The chromato graphic system was equipped with Shimpack columnC18((250x 4.6) mm, 5µ) as stationary phase and UV detector at 220 nm, in conjunction with a mobile phase of phosphate buffer pH 2.0:acetonitrile (60:40,% v/v) at a flow rate of 1.0 mL/min. The developed HPLC technique was found to be rapid as the retention time was 2.2 minutes for droxidopa peak to elute. The method was validated as per the International Conference on Harmonization (ICH) guidelines for specificity linearity, accuracy, precision. The developed method was selective with well-resolved peak. Linearity was observed over the concentration range of 25-150µg/mL for droxidopa. The recovery of Droxidopa was found to be 100.54% - 101.65%. Statistical techniques were employed for the validation of precision, linearity, accuracy, robustness and ruggedness and can be applied for routine analysis. Validation revealed that the developed method was specific, accurate, precise, reliable, robust, reproducible and suitable for the systematic quantitative review.

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Section: Methods Precision and Intermediate Precisionmentioning

confidence: 99%

Method development and validation of droxidopa by HPLC technique

Pandilla¹,

Chitra

Nalini³

et al. 2020

ijrps

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show abstract

“…The injection volume of 20µl with a run time of 10min. Prior to use the buffer was filtered through Millipore 0.45µm filter and degassed on bath sonicator (Sabir et al, 2016), (Kazakevich& LoBrutto, 2006).…”

Section: B Methods 1) Chromatographic System and Conditionsmentioning

confidence: 99%

Solubility Studies of Gefitinib by Validated High Pressure Liquid Chromatographic Method

Vijetha¹,

Reddy²

2020

JSR

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“…When Chernobrrovkin et al compared NAC, NAP, IBLC, and NMC as chiral thiols for OPA derivatization [33], they found that NAC and NMC provided better chiral resolution than NAP and IBLC. However, the resolution factors may depend on the column; mobile phase composition; flow rate; and gradient [38,39]. We previously found that optimal elution condition for OPA adducts was around pH 8 instead of the typical acidic condi-tions [40].…”

Section: Separation Of Enantiomersmentioning

confidence: 99%

Dimethylcysteine (DiCys)/o-Phthalaldehyde Derivatization for Chiral Metabolite Analyses: Cross-Comparison of Six Chiral Thiols

Lkhagva

Tai

2021

Molecules

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Metabolomics profiling using liquid chromatography-mass spectrometry (LC-MS) has become an important tool in biomedical research. However, resolving enantiomers still represents a significant challenge in the metabolomics study of complex samples. Here, we introduced N,N-dimethyl-l-cysteine (dimethylcysteine, DiCys), a chiral thiol, for the o-phthalaldehyde (OPA) derivatization of enantiomeric amine metabolites. We took interest in DiCys because of its potential for multiplex isotope-tagged quantification. Here, we characterized the usefulness of DiCys in reversed-phase LC-MS analyses of chiral metabolites, compared against five commonly used chiral thiols: N-acetyl-l-cysteine (NAC); N-acetyl-d-penicillamine (NAP); isobutyryl-l-cysteine (IBLC); N-(tert-butoxycarbonyl)-l-cysteine methyl ester (NBC); and N-(tert-butylthiocarbamoyl)-l-cysteine ethyl ester (BTCC). DiCys and IBLC showed the best overall performance in terms of chiral separation, fluorescence intensity, and ionization efficiency. For chiral separation of amino acids, DiCys/OPA also outperformed Marfey’s reagents: 1-fluoro-2-4-dinitrophenyl-5-l-valine amide (FDVA) and 1-fluoro-2-4-dinitrophenyl-5-l-alanine amide (FDAA). As proof of principle, we compared DiCys and IBLC for detecting chiral metabolites in aqueous extracts of rice. By LC–MS analyses, both methods detected twenty proteinogenic l-amino acids and seven d-amino acids (Ala, Arg, Lys, Phe, Ser, Tyr, and Val), but DiCys showed better analyte separation. We conclude that DiCys/OPA is an excellent amine-derivatization method for enantiomeric metabolite detection in LC-MS analyses.

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HPLC Method Development and Validation: A Review

Cited by 46 publications

References 9 publications

Method development and validation of droxidopa by HPLC technique

Method development and validation of droxidopa by HPLC technique

Solubility Studies of Gefitinib by Validated High Pressure Liquid Chromatographic Method

Dimethylcysteine (DiCys)/o-Phthalaldehyde Derivatization for Chiral Metabolite Analyses: Cross-Comparison of Six Chiral Thiols

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