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Observational Study
. 2015 Apr;94(14):e640.
doi: 10.1097/MD.0000000000000640.

Comparison of UHPLC and HPLC in benzodiazepines analysis of postmortem samples: a case-control study

Behnam Behnoush  1 , Ardeshir Sheikhazadi, Elham Bazmi, Akbar Fattahi, Elham Sheikhazadi, Seyed Hossein Saberi Anary
Affiliations
Observational Study

Comparison of UHPLC and HPLC in benzodiazepines analysis of postmortem samples: a case-control study

Behnam Behnoush et al. Medicine (Baltimore). 2015 Apr.

Abstract

The aim of this study was to compare system efficiency and analysis duration regarding the solvent consumption and system maintenance in high-pressure liquid chromatography (HPLC) and ultra high-pressure liquid chromatography (UHPLC). In a case-control study, standard solutions of 7 benzodiazepines (BZs) and 73 biological samples such as urine, tissue, stomach content, and bile that screened positive for BZs were analyzed by HPLC and UHPLC in laboratory of forensic toxicology during 2012 to 2013. HPLC analysis was performed using a Knauer by 100-5 C-18 column (250 mm × 4.6 mm) and Knauer photodiode array detector (PAD). UHPLC analysis was performed using Knauer PAD detector with cooling autosampler and Eurospher II 100-3 C-18 column (100 mm × 3 mm) and also 2 pumps. The mean retention time, standard deviation, flow rate, and repeatability of analytical results were compared by using 2 methods. Routine runtimes in HPLC and UHPLC took 40 and 15 minutes, respectively. Changes in mobile phase composition of the 2 methods were not required. Flow rate and solvent consumption in UHPLC decreased. Diazepam and flurazepam were detected more frequently in biological samples. In UHPLC, small particle size and short length of column cause effective separation of BZs in a very short time. Reduced flow rate, solvent consumption, and injection volume cause more efficiency and less analysis costs. Thus, in the detection of BZs, UHPLC is an accurate, sensitive, and fast method with less cost of analysis.

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Conflict of interest statement

The authors have no funding and conflicts of interest to disclose.

Figures

FIGURE 1
FIGURE 1
Benzodiazepine metabolites pathway.
FIGURE 2
FIGURE 2
Comparison of chromatograms of clonazepam in HPLC and UHPLC, left and right panels, respectively.
FIGURE 3
FIGURE 3
Comparison of chromatograms of flurazepam in HPLC and UHPLC, left and right panels, respectively.
FIGURE 4
FIGURE 4
Comparison of chromatograms of oxazepam in HPLC and UHPLC, left and right panels, respectively.
FIGURE 5
FIGURE 5
Comparison of chromatograms of alprazolam in HPLC and UHPLC, left and right panels, respectively.
FIGURE 6
FIGURE 6
Comparison of chromatograms of diazepam in HPLC and UHPLC, left and right panels, respectively.
FIGURE 7
FIGURE 7
Comparison of chromatograms of lorazepam in HPLC and UHPLC, left and right panels, respectively.
FIGURE 8
FIGURE 8
Comparison of chromatograms of chlordiazepoxide in HPLC and UHPLC, left and right panels, respectively.
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