Analysis of Anticoagulant Drugs by HPLC Manual
Contents
1. D Analysis of Anticoagulant Drugs by HPLC Udo Huber Pharmaceutical Abstract Warfarin is used in human medicine as anticoagulant to decrease the rate of blood clotting This helps to prevent thrombosis after operations In contrast warfarin is also used as a rodenticide against rats making use of the same effect here the animals die because blood clotting is prevented Figure 1 shows the chromatogram of warfarin using gradient analysis on a reversed phase column and UV detection To avoid decomposition of samples the autosampler temperature was Set to 4 C Absorbance ae Warfarin Column 4 6 x 75 mm Zorbax SB C18 3 5 um 200 Mobile phase A 0 025 M KH PO in water 150 pH 3 with H SO wi B acetonitrile Flow rate 1 0 ml min 50 Gradient 20 B to 80 B in 10 min Column wash 0 80 B to 20 B in 2 min 0 a 4 6 8 10 UV detector Time min variable wavelength detector 204 nm standard cell Figure 1 fluorescence detector Analysis of wafarin using a variable wavelength detector 272 355 nm standard cell Column compartment temperature De Stop time 12 min Post time 5 min Injection volume 5 l Agilent Technologies Innovating the HP Way The performance of the HPLC method is shown in the table below Compound LOD forS N 2 PrecisionofRT Precision of Area mg I RSD of 10 runs RSD of 10 runs 100 mg I 100 mg I Waferin 0 1 0 02 0 46 Injection volume 5 ul The analysis can also be done us2. art number 866953 902 e Recommended Guard cartridges Zorbax SB C18 5 um 4 6 x 12 5 mm Agilent part number 820950 920 4 pk Udo Huber is application chemist at Agilent Technologies Waldbronn Germany For more information on our products and services visit our worldwide website at http w ww agilent com chem Copyright 1998 Agilent Technologies Released 08 98 Publication Number 5968 1082E Agilent Technologies Innovating the HP Way
3. ing the Agilent 1100 Series fluorescence detector figure 2 The method presented here shows an easy but reliable and precise analysis of the anticoagulant drug warfarin The values for LOD precision of RT and precision of area show the good performance of the analysis Emission LU Excitation 272 nm Warfarin 1 2 3 4 5 6 7 8 9 Time min Figure 2 Analysis of wafarin using a fluorescence detector column Zorbax SB C18 2 1 x 50 mm 5 um Note Since the method was specifically developed on the Agilent 1100 Series system you might not be able to reproduce this analysis on an older system or even on a new system with lower performance To avoid sample decomposition it is necessary to use a cooled auto Sampler for example the Agilent 1100 Series thermostatted autosampler The analysis can be performed with either an Agilent 1100 Series variable wavelength detector or an Agilent 1100 Series fluorescence detector Agilent 1100 Series Quaternary pump includes vacuum degasser Thermostatted autosampler Thermostatted column compartment Variable wavelength detector standard flow cell 10 mm path length 13 ul cell volume Alternative Vacuum degasser Binary pump Diode array detector Standard flow cell 10 mm path length 13 yl cell volume e Fluorescence detector standard flow cell 8 ul cell volume e Agilent ChemStation 3D software Columns e Zorbax SB C18 3 5 um 4 6 x 75 mm Agilent p
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