High reproducibility of fluorescence detector with temperature controlled cell

October 22, 2020


Indole-3-acetic acid known as auxin, a growth inhibitor which play a role in morphosis in plants. Artificially synthesized auxin is widely used as a herbicide and a fruit growth promoter, and is expected to make a significant contribution in increasing research into increasing crop yields and biomass.

Generally, the intensity of fluorescence in a compound is temperature dependent. Indole-3-acetic acid is a compound whose fluorescence intensity is decreased with a rise of temperature.

In this article, indole-3-acetic acid was measured using an HPLC with FP-4020 fluorescence detector with a temperature controlled cell, and the effects of fluctuation in room temperature and the benefits of cell temperature control were reported.

FP-4020 Fluorescence detector


Measurement conditions
Column: CrestPak C18S (4.6 mmI.D. x 150 mmL, 5 µm)
Eluent: 0.3% acetic acid in methanol/water (42/58)
Flow rate: 1.0 mL/min
Column temp.: 40 ̊C
Wave length: Ex 280 nm, Em 360 nm, Gain x100
Cell temp.: 30 ̊C and Off
Injection volume: 5 µL
Standard: 100 ng/mL Indole-3-acetic acid

Structure of Indole-3-acetic acid


Figure 1 shows peak height change of indole-3-acetic acid with rise in room temperature.

Figure 1. Change of peak height of indole-3-acetic acid with a change in room temperature

Figure 2  shows chromatograms of indole-3-acetic acid with and without cell temperature control.

Figure 2 Chromatograms of indole-3-acetic acid with and without cell temperature control (A: Cell temperature control: ON, B: Cell temperature control; OFF)

Table 1 shows change rate of peak height and the relative standard deviation with and without cell temperature control. As a result of using cell temperature control, the peak intensity is stabilized minimizing the effect of fluctuations in room temperature providing improved reproducibility.

Table 1. The rate of change in peak height and the relative standard deviation (with and without cell temperature control)

Temperature change (ºC) Rate of change (%) %RSD
Cell temperature control: ON (30ºC) 6.6 0.95 0.75
Cell temperature control; OFF 6.7 -19.49 8.77

*Rate of change: The change rate of peak height. The peak height at minimum room temperature is normalized as ”1”, as the reference value.
*%RSD: The relative standard deviation of the peak height at six repeat measurement.

About the Author

Chromatography Group