Calculating optimal modulation periods to maximize the peak capacity in two-dimensional HPLC

Kanta Horie, Hiroshi Kimura, Tohru Ikegami, Akira Iwatsuka, Nabil Saad, Oliver Fiehn, Nobuo Tanaka

Research output: Contribution to journalArticlepeer-review

99 Scopus citations


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 ∼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 ∼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.

Original languageEnglish (US)
Pages (from-to)3764-3770
Number of pages7
JournalAnalytical Chemistry
Issue number10
StatePublished - May 15 2007

ASJC Scopus subject areas

  • Analytical Chemistry


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