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Introduction to Modern Liquid Chromatography, Third Edition part 8

Introduction to Modern Liquid Chromatography, Third Edition part 8. High-performance liquid chromatography (HPLC) is today the leading technique for chemical analysis and related applications, with an ability to separate, analyze, and/or purify virtually any sample. Snyder and Kirkland's Introduction to Modern Liquid Chromatography has long represented the premier reference to HPLC. This Third Edition, with John Dolan as added coauthor, addresses important improvements in columns and equipment, as well as major advances in our understanding of HPLC separation, our ability to solve problems that were troublesome in the past, and the application of HPLC for new kinds of samples. . | 26 BASIC CONCEPTS AND THE CONTROL OF SEPARATION where K Cs C t is the equilibrium constant for Equation 2.2 and fl Vs V is the phase ratio the ratio of stationary and mobile-phase volumes within the column. We will see that k is a very important property of each peak in the chromatogram values of k can help us interpret and improve the quality of a separation. A solute molecule must be present in either the mobile or stationary phase so that if die frattion of moleculesinthemobiie phaseis t the fraction in the stationfryphasemustbe 1 mR thprpfofefrpmEqbation 2.3i wahave bi i 2.3ai h or R TTk 2 3b Thp rptpntion timp tR of X can be defined as distance divided by speed or band ph E tR 2.4 Similarly theretentioniime of ihesolaentmaaP h t0 u 2-4a where u is the averaga moPiie-paasnaerocitp. Elimrnatrng tf between Equations 2.4 and 2.41 gives tR 2.4b 111 . 11 which wichO. s ue pnq. 2.1 and Equation 2.3b then gives ÎR to l k 2.5 Equation 2.5 can also be expressed in terms of retention volume Vr IrF where F is the mobfle-phosa flow rate mL min VR Vm l k 2.5a Here Vm isehc epiumnrfeba-co pwe ewhal io taF Pseetho iuriiKirliiruisioii of Wm and Eq. ISa balow . Equation 2.5 can be rearranged to give k - 2-6 m 2.3 RETENTION 27 which allows the calculation of values of k for each peak in the chromatogram. Visual estimates of k from the chromatogram based on Eq. 2.6 are often used in practice because exact values of k are seldom needed for developing a separation method development or during routine analysis. Thus k is equal to the corrected retention time t to measured in units of to or m hj_i 2.6a Se As illustratedm 6igu_2.3A whic_2orrespondstot6e chromatogram of Fig. 2.3c the distance to can be used to mark off approximate values of k beginning at time to thus k equals 1 2 and 4 respectively for compounds X Y and Z. We will see in Section 2.4.1 that values of k between about 1 and 10 are preferred for various reasons. Therefore it is important to be able to estimate or .