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C190-E204










            Technical                    Improving Method Transfer

            Report                       by Adjustment of Gradient Delay




                                         Akihiro Kunisawa , Daiki Fujimura , Yusuke Osaka 1
                                                     1
                                                                 1

            Abstract:

            In facilities that have number of high performance liquid chromatography (HPLC) systems, an existing method that gives proper result by
            one  HPLC  system  is  often  applied  to  other  HPLC  systems  (method  transfer).  However,  due  to  the  difference  in  system  volume,  pump
            characteristics, and liquid delivery mechanisms among systems, method transfer can yield different results even though the same method
            is used.
            In this report, we ÿrst  explain the gradient delay due to the system volume difference and the effect of the delay on separation. We then
            show  an  example  of  method  transfer  with  adjustment  of  gradient  delay  using  alkylphenones  and  UV  absorbents  as  samples.  We  also
            describe the integrated liquid chromatograph (LC) system "i-Series" that supports to optimize method transfer and the newly designed
            function of "Analytical Conditions Transfer and Optimization (ACTO) " equipped in the latest version of LabSolutions software.

            Keywords: HPLC, method transfer, ACTO, i-Series




                         ound
            1. Background
            1. Backgr
            HPLC is used for the analysis of target compounds and their related                       System A
            impurities in a variety of applications including pharmaceutical and
            food products. Facilities that use HPLC systems create methods using
            their own original analytical conditions and/or specified testing regu-
            lations. The validated methods are then used with a number of other
            HPLC  systems  in  many  cases.  In  such  situations,  reproducibility
            (compatibility) among systems is an important factor as well as re-
            peatability of measurements. Even when using the same method, dif-
            ferent HPLC systems can give different chromatograms (Fig. 1). Par-
            ticularly in gradient elution, retention times, resolution and other fac-
            tors will be largely affected as a result of method transfer. For exam-  0.00  0.50  1.00  1.50  2.00  2.50  3.00  3.50  4.00 min
            ple,  while  an  existing  method  may  succeed  in  separating  a  target
            compound  from  co-existing  impurities  in  one  system,  the  same
            method may not succeed in separating these compounds in other                  Method Transfer
            systems. So it is often required to optimize analytical conditions for
            each individual system, which is an extremely time-consuming pro-
            cess. Such variations in retention time and separation are caused by
            difference in system volume and pump performance among systems
            (see section 2 for details). Especially in ultrahigh-speed analysis, even   Unable to obtain the same  System B
            small difference in system volume can cause great difference in analy-  chromatogram
            sis results due to small volume of dedicated column.
            Further,  in  pharmaceuticals,  food  and  other  fields  where  the  test
            methods are specified by regulations, changes in analytical conditions
            are not permitted, which may be an issue.




                                                                 0.00  0.50  1.00  1.50  2.00  2.50  3.00  3.50  4.00 min
                                                                            Fig. 1  Problems in Method Transfer


            1 Analytical & Measuring Instruments Division                                                            1
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