Calculating optimal modulation periods to maximize the peak capacity in two-dimensional HPLC.
Theoretical calculations are presented to optimize modulation period for maximum total peak capacity in comprehensive two-dimensional HPLC (2D-HPLC) taking into account the effect of modulation on the apparent peak capacity of the first-dimension (1D) separation. Results indicate that modulation periods are most favorable when they are adjusted to approximately 2.2-4 times the standard deviation of a 1D peak in order to avoid excessively short run times at the second dimension (2D). Data are presented that effective peak capacities of several thousand in 60 min can be expected for practical 2D-HPLC conditions, utilizing 1D gradient elution followed by 2D isocratic elution, that remain at approximately 50-70% of the theoretical maximum peak capacity. This work suggests that lower modulation frequencies and longer 2D separation times than previously proposed are favorable under realistic chromatographic conditions, alleviating some practical problems associated with 2D-HPLC.
Horie
Kanta
K
Department of Biomolecular Engineering, Kyoto Institute of Technology, Matsugasaki, Sakyo-ku, Kyoto 606-8585, Japan.
Kimura
Hiroshi
H
Ikegami
Tohru
T
Iwatsuka
Akira
A
Saad
Nabil
N
Fiehn
Oliver
O
Tanaka
Nobuo
N
D002851:Chromatography, High Pressure Liquid / Q000379:methods* / Q000592:standards; D008962:Models, Theoretical