Direct injection of solid‐phase extraction eluents onto silica columns for the analysis of polar compounds isoniazid and cetirizine in plasma using hydrophilic interaction chromatography with tandem mass spectrometry

Abstract

Isoniazid and cetirizine do not retain well on reversed-phase columns due to their high polarity. Silica columns, when operated under hydrophilic interaction conditions, do provide excellent retention of these compounds. We have developed simple and proof of concept analytical methods for the analysis of isoniazid and cetirizine in animal and human plasma, respectively. Both methods employed the approach of direct injection of solid-phase extraction (SPE) organic eluents onto silica columns for analysis, thus eliminating evaporation and reconstitution steps that are typically needed for reversed-phase liquid chromatographic analysis. Isoniazid was extracted from animal plasma samples using a Waters Oasis HLB 96-well plate and then eluted with acetonitrile, while cetirizine was extracted from human plasma with a Waters MCX mu-Elute plate and then eluted with acetonitrile containing 5% concentrated ammonium hydroxide. The direct injection of the SPE eluent onto the analytical column was necessary since significant loss of isoniazid was found during the evaporation and reconstitution steps. The method for isoniazid also enabled ultra-fast analysis due to the relatively low back-pressure exhibited by silica columns even under high flow conditions. Both methods show good linearity, accuracy and precision covering the range of 10-2000 ng/mL of isoniazid, and 1-1000 ng/mL of cetirizine in plasma. Substantial time savings were realized as a result of both the elimination of the evaporation and reconstitution steps and the fast chromatographic analysis.

A Sensitive HPLC Method for Determination of Isoniazid in Rat Plasma, Brain, Liver and Kidney

“…Intra and inter-day variability's (RSD) for extraction of INH from plasma and other tissue homogenates were less than 5% and accuracy was within ± 5% (Table 1) and were comparable with other reported methods of isoniazid [15,16,23].The absolute recovery of INH was calculated for replicate spiked QC samples (200, 1000 and10,000 ng/ mL) ( …”

“…Poor retention of polar compounds has been reported earlier on non polar reverse phase HPLC columns earlier [15], however, numerous methods for determination of polar compounds using reverse phase columns specific to isoniazid exists in the literature [16][17][18][19]. Establishment of a reproducible analytical method is always prerogative step before the development of a pharmaceutical product.…”

Critical Review of Development, Validation, and Transfer for High Throughput Bioanalytical LC-MS/MS Methods

“…In many cases, an evaporation step followed by reconstitution with mobile phase compatible solvent is necessary. Further improvement on throughput for another 50% was made when the organic extracts from SPE [51,52] or LLE [53,54] were injected directly onto the silica column with low aqueous -high organic mobile phases. While the commonly used SPE or LLE extraction solvents are stronger elution solvents (not compatible) than a mobile phase on typical reversed-phase chromatography, they are weaker elution solvents (compatible) on the silica column with low aqueous -high organic mobile phases operated under hydrophilic interaction chromatography (HILIC).…”

“…(Fig. 4) shows a fast gradient elution of nicotinic acid and its six metabolites within one minute [77]. Nicotinic acid and metabolites are extremely polar compounds and have no retention at all on a reversed-phase monolithic column.…”

“…Matrix effects has been defined as the analyte ionization suppression or enhancement at the presence of the matrix components that could originated from the endogenous compounds, metabolites, and co-administered drugs [52]. More recently, matrix effects from internal standards [93], dosing vehicles [94][95][96], and, mobile phase additives [97], and plastic tubes [98] were also reported.…”