Analytical Techniques in Pharmaceutical Analysis for Samples Separation, Characterization, Determination and its Handling

Separations are key aspects of many modern analytical methods. Real world samples contain many analytes. Many analytical methods do not offer sufficient selectivity to be able to speciate all the analytes that might be present. Separation isolates analytes. Most separation methods involve separation of the analytes into distinct chemical species, followed by detection. Instrumental methods may be used to separate samples using chromatography, electrophoresis or field flow fractionation. The development of the pharmaceuticals brought a revolution in human health. These pharmaceutical would serve their intended only if they are free from impurities and administered in appropriate amount. To make drugs serve their purpose various chemical and instrumentation method were developed at regular intervals which are involved in the estimation of drugs. For this analytical instrumentation and methods plays an important role. This review highlights the role of the analytical instrumentation and analytical method in assessing the quality of the drug. The review highlights variety of analytical techniques such as chromatographic, spectroscopic, electrophoretic, titrimetric, microscopic and electrochemical and their corresponding methods that has been applied in the analysis of pharmaceuticals. Quality of pharmaceutical product largely depends upon the environment controls during its storage and handling. Each pharmaceutical product should be handled and stored under specified storage condition labelled on product information data sheet or product pack. Ion chromatography was based on the use of a low-capacity anion exchange separator column used with a basic eluent and a suppressor column. IC is a part of high-performance liquid chromatography used to separate and determine anions and cations. Keywords: Analytical techniques, Sample separation, Chromatography, Spectroscopy, Electrochemical method, Microscopically method, Sample handling, Sample determination Electrophoresis

Supercritical fluid chromatography coupled to mass spectrometry: A valuable tool in food analysis

Producción CientíficaSupercritical fluid chromatography (SFC), although known for several decades, has undergone a growing interest in the last few years fueled by the introduction of modern instruments with improved robustness, and hyphenation to mass spectrometry (MS). This allows the analysis of trace compounds in complex samples with high selectivity, high sensitivity and in a short time, which has contributed to its increased use in the food analysis area. This work reviews the principal applications of SFC-MS in food analysis, highlighting the most important achievements

Efikasnost superkritične fluidne hromatografije u kreiranju ekološki prihvatljivih rešenja u farmaceutskoj analizi

Initially employed primarily at a preparative scale for enantiomer separation of chiral drug candidates, Supercritical Fluid Chromatography (SFC) is nowadays extensively used in the analytical mode. Recent advances in SFC separation science have emphasized its potential for modern and environmentally friendly pharmaceutical analysis. The aim of this review is to provide a deeper insight into the main fundamental and practical aspects of the SFC technique in order to familiarize readers with its versatile nature and efficiency in creating sustainable chromatographic solutions. All considerations are made primarily in the context of the most widely used mode of operation - achiral SFC. In addition, recent applications of this promising technique are presented at the end of the article to further promote its use in pharmaceutical analytical practice.Na početku pretežno korišćena kao preparativna tehnika u enantioseparaciji hiralnih molekula kandidatâ za lek, superkritična fluidna hromatografija (eng. supercritical fluid chromatography, SFC) danas se široko koristi u analitičke svrhe. Noviji naučni napori ukazali su na značaj SFC tehnike u modernoj i ekološki prihvatljivoj farmaceutskoj analizi. Cilj ovog preglednog rada je pružanje dubljeg uvida u najvažnije fundamentalne i praktične aspekte SFC tehnike, kako bi se čitaocima približio njen svestrani karakter, te efikasnost u kreiranju održivih hromatografskih rešenja. Sva razmatranja prevashodno su data u kontekstu najzastupljenijeg režima rada - ahiralne SFC. Takođe, na kraju rada predstavljene su savremene primene ove obećavajuće tehnike kako bi se dodatno ohrabrilo njeno usvajanje u farmaceutsku analitičku praksu

Leveraging the power of supercritical fluid chromatography for eco-conscious solutions in pharmaceutical analysis

Initially employed primarily at a preparative scale for enantiomer separation of chiral drug candidates, Supercritical Fluid Chromatography (SFC) is nowadays extensively used in the analytical mode. Recent advances in SFC separation science have emphasized its potential for modern and environmentally friendly pharmaceutical analysis. The aim of this review is to provide a deeper insight into the main fundamental and practical aspects of the SFC technique in order to familiarize readers with its versatile nature and efficiency in creating sustainable chromatographic solutions. All considerations are made primarily in the context of the most widely used mode of operation - achiral SFC. In addition, recent applications of this promising technique are presented at the end of the article to further promote its use in pharmaceutical analytical practice

Sub/supercritical fluid chromatography purification of biologics.

Doctoral Degree. University of KwaZulu-Natal, Durban.Peptide and protein drugs are highly versatile with numerous therapeutic properties such as anti-cancer, anti-diabetic, anti-hypertensive, and anti-microbial; which are therefore ideal candidates for the development of next-generation drugs. This is exemplified using these drugs for the treatment of diseases such as diabetes. Diabetes is one of the most prevalent non-communicable diseases worldwide. The rapid increase in the number of diabetic patients globally places a burden on current insulin manufacturers. The traditional reversed-phase high-performance liquid chromatography (RP-HPLC) purification methods of insulin and peptides are problematic, tedious with long run times of approximately 50 minutes, low yields employ harsh solvents such as acetonitrile, which has a negative impact on the environment. There is a need for a greener process for the purification of insulin and peptides. Sub/supercritical fluid chromatography (SFC) can provide the solution since it utilises greener mobile phases such as carbon dioxide (CO2) and methanol, which can be recycled. However, there is a paucity of knowledge regarding the SFC purification of human insulin and peptides. Therefore, this research study aimed to provide an efficient, innovative approach for the biosynthesis of human insulin and the SFC purification of biosynthesised human insulin, as well as the extension into the SFC purification of peptides. The background of these topics is presented in Chapter One. Chapter two (manuscript one) presents the development of a novel and more efficient method of human insulin biosynthesis in Escherichia coli (E. coli). Several of the conventional steps were eliminated. The crude biosynthesised protein sequence was verified using protein sequencing, which had a 100% similarity to the human insulin sequence. The biological activity of the biosynthesised human insulin was tested in vitro using a 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay. The biosynthesis of human insulin was conducted on a laboratory-scale basis; future studies should investigate scaling up of this method. Chapter three (manuscript two) was based on the SFC purification of the commercially available standard sample of human insulin and a crude biosynthesised sample of human insulin. The SFC purified standard the biosynthesised human insulin samples were detected and quantified using liquid chromatographymass spectrometry (LC-MS) and protein sequencing techniques. SFC columns, i.e., silica, 2’ ethyl pyridine, diol-HILIC, and the pentafluoro phenyl (PFP), were evaluated to determine the ideal column. The PFP column gave the best results since it displayed good peak shapes, resolution, retention factors, retention times, and the least relative standard deviation in comparison to the other columns. Therefore, the aforementioned column was selected for further analysis using the biosynthesised human insulin, whereby a column efficiency test was conducted on a semi-preparative scale, yielding 84% recovery. Subsequently, the biological activities of the SFC purified standard sample of human insulin and biosynthesised version were tested in vitro using a MTT assay. The results indicated that the biological activities of the standard and biosynthesised human insulin derivatives were retained subsequent to SFC purification. The biological activities were highly significant, with a p-value of < 0.0001. From chapter three, band broadening and phase separation peaks were experienced during SFC purification of the commercially available standard sample of human insulin and biosynthesised human insulin. Therefore, in Chapter four (manuscript three), a SFC purification method was developed to purify peptides at an analytical scale. A tetrapeptide [insulin β chain peptide (15-18)], octapeptide [angiotensin II], nonapeptide [insulin β chain peptide (15-23)] were purified using four SFC columns, i.e., PFP, diolHILIC, silica, and 2’ ethyl pyridine. Subsequently, the 2’ ethyl pyridine column was selected for further analysis based on the reproducibility, peak shapes, efficient separations and retention factors. The three peptides were monitored using LC-MS analysis. The successful peptide recoveries ranged from 80-102%. Chapter five pertains to the summary and conclusion drawn from the study and reflects on possible future endeavours. The present study was successful in providing a more affordable and innovative approach for the biosynthesis of human insulin. The work also successfully developed a rapid, greener, and more efficient method of SFC purifying biosynthesised human insulin and peptides as opposed to conventional HPLC purification methods. As far as we are aware, this study is the first of its kind to purify biosynthesised human insulin and this combination of peptides using SFC purification techniques. Future research studies can focus on the SFC purification of larger protein molecules and consider the use of custom columns and or other modifiers for the improvement of the isolation of other highly sought after biologics within the pharmaceutical industry

Folia Pharmaceutica Universitatis Carolinae: LI

Abstracts from the 10th Postgraduate and Postdoc Conference of the Faculty of Pharmacy in Hradec Králové, Charles University, Hradec Králové, 22–23 January 2020

Resolución enantiomérica de compuestos volátiles quirales mediante técnicas multidimensionales: cromatografía en lecho móvil simulado con fluídos supercríticos

El primer objetivo de la presente Memoria es el estudio del comportamiento de la heptakis-(2,3,6-tri-O-metil)-β-ciclodextrina en el reconocimiento quiral de racematos volátiles mediante técnicas cromatográficas unidimensionales. La selección de fuentes naturales constituye el segundo objetivo de este trabajo y se plantea desde la perspectiva de establecer nuevas alternativas sostenibles para la obtención de enantiómeros puros. A este respecto, se propone el empleo de técnicas cromatográficas multidimensionales adecuadas para la necesaria determinación previa de los excesos enantioméricos de aquellos compuestos quirales presentes en las diversas muestras complejas que sean de interés en cada caso particular. El tercer objetivo es el diseño y construcción de una planta de cromatografía en lecho móvil simulado con fluidos supercríticos (Supercritical Fluid Simulated Moving Bed Chromatography, SF-SMB) para su posterior aplicación a la obtención de enantiómeros a escala preparativa, lo que supone el desarrollo y puesta a punto de una nueva tecnología que cumple con los requisitos de las actuales técnicas analíticas avanzadas