Drug Substance and Excipient Characterization
Dilip M. Parikh in Handbook of Pharmaceutical Granulation Technology, 2021
In addition to its application in the separation and identification of materials, chromatography is also employed to detect potential interactions between materials. Both thin-layer chromatography and liquid chromatography are commonly employed for this purpose. In thin-layer chromatography, the stationary phase consists of a powder adhered onto a glass, plastic, or metal plate. The powders commonly used are silica, alumina, polyamides, celluloses, and ion-exchange resins. Solutions of the drug, excipient, and drug–excipient mixture are prepared and spotted on the same baseline at one end of the plate. The plate is then placed upright in a closed chamber containing the solvent, which constitutes the mobile phase. As the solvent moves up the plate, it carries with it the materials. Those materials that have a stronger affinity for the stationary phase will move at a slower rate. The material is identified by its Rf value, which is defined as the ratio of the distance traveled by the material to the distance traveled by the solvent front. The position of the material on the plate is indicated by spraying the plate with certain reagents or exposing the plate to ultraviolet radiation. If there is no interaction between the drug and excipient, the mixture will produce two spots whose Rf values are identical to those of the individual drug and excipient. If there is interaction, the complex formed will produce a spot whose Rf value is different from those of the individual components.
Gas Chromatography
Joseph Chamberlain in The Analysis of Drugs in Biological Fluids, 2018
The type of stationary phase has a bearing on the separations obtained for various classes of compounds. The stationary phases can be divided into two classes: selective and nonselective. The nonselective phases separate the analytes more by virtue of molecular size and shape rather than by chemical properties. Thus, very closely related compounds do not separate well on nonselective phases, but such phases are useful for the separation and characterization of homologs. Selective phases will separate according to the the selective retention of certain groups. For example, pentoxifylline and its primary metabolite (Figure 6.4) have virtually identical molecular weights and do not separate on phases such as OV-1, but separate on phases such as OV-101, which selectively retains oxo compounds compared with the corresponding hydroxy compound. Specific mention should also be made of the group of column packings designed for low molecular weight analogs (such as alcohol). These columns (of which the Porapaks and Chromosorbs are the most widely used) separate the analytes by molecular size exclusion rather than by a partitioning effect.
Basics Of Gas Chromatography Mass Spectrometry System
Raquel Cumeras, Xavier Correig in Volatile organic compound analysis in biomedical diagnosis applications, 2018
The polarity of the analytical GC column is a function of the stationary phase’s chemical composition, and the selection of the appropriate column for a particular application is dictated by both the polarity of the target compounds and the GC column’s stationary phase composition. If the stationary phase and the target compounds have comparable polarities, this will result in a strong retention interaction; higher column retention generally equates to greater chromatographic separation and higher resolution. The polarity of the stationary phase will also strongly influence column selectivity and separation. The selectivity factor (α) can be directly related to the stationary phase composition, giving rise to subsequent interaction with the target compounds via intermolecular forces such as hydrogen bonding and dipole–dipole interaction. The effect of temperature is also an important consideration as high polarity stationary phase generally has a lower maximum operating temperature limit (Agilent, 2016; Restek, 2016).
On the potential of micro-flow LC-MS/MS in proteomics
Published in Expert Review of Proteomics, 2022
Yangyang Bian, Chunli Gao, Bernhard Kuster
The stationary phase, i.e. the chromatographic column, is central to any LC separation. The physical dimensions of the column and material packed into the column determine the type of analyte and the quantities that can be separated (particle material, internal diameter (i.d.)) and the efficiency of the separation (length, particle size, flow rate, temperature etc.). As for other areas of chromatography, the trend toward smaller particle sizes and the concomitant necessity to separate at very high pressure (ultra-high performance liquid chromatography, UHPLC, >1,000 bar) has also been followed in proteomics [8–10]. The use of very small-diameter columns has dominated the field for many years because this offered the sensitivity required to support proteomic applications. As discussed below, this ethos is now beginning to change. For the purpose of the discussion in this review, LC columns are broadly classified into four groups that are based on the internal diameter (Figure 1). Analytical columns typically have 2.1–4.6 mm i.d. and are used at flow rates of >200 μL/min. Columns with 0.5–1 mm i.d. and a flow rate of 10–200 μL/min are referred to as micro-flow, and columns with 150–300 μm i.d. and a flow rate of 1–10 μL/min are known as capillary flow. Finally, columns with an i.d. <100 μm and used at a flow rate <1 μL/min are termed nano-flow [11,12].
Infrared analyzers for the measurement of breastmilk macronutrient content in the clinical setting
Published in Expert Review of Molecular Diagnostics, 2020
Cristina Borràs-Novell, Ana Herranz Barbero, Victoria Aldecoa-Bilbao, Georgina Feixas Orellana, Carla Balcells Esponera, Erika Sánchez Ortiz, Oscar García-Algar, Isabel Iglesias Platas
Liquid chromatography is used to separate, identify, and quantify each component of a mixture based on their mass (HPLC, UPLC) or electric charge (HPAEC-PAD) (volume needed 0.1 ml) [25,282930]. The mixture is dissolved in a fluid called the mobile phase, which carries it through a structure holding another material called the stationary phase. The various constituents of the mixture, in this case lactose, travel at different speeds, causing them to separate and this is applied to their identification. It can discriminate between lactose and the oligosaccharides that are abundantly present in human milk [31]. In the lactose IK-Chloramine-T reaction, lactose is quantified according to its reducing properties, which are used to produce iodine from 0.04 M thiosulfate [5]. This procedure is completed in approximately 120 min and requires 10 ml [32]. The automated orcinol method analyzes the degradation products obtained during enzymatic hydrolysis of sugars in deproteinized samples. These two last methods will react with both lactose and oligosaccharides [3,5].
Antioxidant and cytoprotective properties of loganic acid isolated from seeds of Strychnos potatorum L. against heavy metal induced toxicity in PBMC model
Published in Drug and Chemical Toxicology, 2022
Alagarsamy Abirami, Simran Sinsinwar, Perumal Rajalakshmi, Pemaiah Brindha, Yamajala B. R. D. Rajesh, Vellingiri Vadivel
Among four fractions, LF2 with high antioxidant activity (98.24%) was selected for further purification by column chromatography (Cunha et al. 2017). The stationary phase consists of silica gel which was packed in a glass column (38.2 cm length × 2.2 cm dia) with hexane. The silica column was packed with hexane without air bubbles and the dried LF2 fraction was made into slurry with silica, so the adsorbed compounds on silica particles were eluted with hexane as the initial solvent. Compounds were eluted from column by using different ratios of hexane/ethyl acetate (100:0, 90:10, 80:20, 70:30, 60:40, 50:50, 40:60, 30:70, 20:80, 10:90, 0:100%). Similarly ethyl acetate/methanol combinations (100:0, 90:10, 80:20, 70:30, 60:40, 50:50, 40:60, 30:70, 20:80, 10:90, 0:100%) were also used to elute the compounds. Totally nine fractions were eluted such as CF1 (100% hexane), CF2 (100% hexane), CF3 (10:90% ethyl acetate: hexane), CF4 (20:80% ethyl acetate: hexane), CF5 (50:50% ethyl acetate: hexane), CF6 (100% ethyl acetate), CF7 (50:50% ethyl acetate: methanol), CF8 (100% methanol) and CF9 (100% methanol). All the fractions were evaporated and subjected to in vitro screening (DPPH assay). Among the nine fractions, CF7 was found to had high antioxidant activity (92.145%) when compared other column fractions. It also showed single spot in TLC with the solvent system of ethyl acetate: methanol (50:50%) (Gini and Jothi 2018) and hence CF7 was submitted for LC-MS/MS and NMR analysis.
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