Hydroxypropyl-β-cyclodextrin effect on the fluorescence of auxin and skatole and on the simultaneous determination of binary mixtures of indole compounds in urine by first derivative spectrofluorimetry

The effect of hydroxypropyl-β-cyclodextrin (HPβCD, β-cyclodextrin (βCD) derivative) was determined on the fluorescence emission spectra of skatole (3-methylindole, MI) and auxin (indole 3-acetic acid, IA). The experiments were conducted at excitation wavelength (λ_ex) = 280 nm, in aqueous solutions at different pH values, with and without HPβCD as receptor. The enhanced fluorescence in the presence of the receptor showed a 1:1 host–guest interaction and the values of the association constants K_A were between 100 and 200 mol⁻¹ L. The effects of methanol and propanol with and without receptor were also studied. The limit of detection (L_D) for the HPβCD enhanced fluorimetric method at pH = 7.00 were 0.279 and 0.765 ng mL⁻¹ for MI and IA, respectively. For the determination of indole compounds with closely overlapping spectral bands a zero-crossing first derivative spectrofluorimetric method with or without HPβCD is described. Binary mixtures of MI or IA with melatonin (N-acetyl-5-methoxytryptamine, M) or 5-methoxytryptamine (5-methoxy-3-(2-aminoethyl)indole, 5M) were analysed in the presence of HPβCD. In the absence of receptor, binary mixtures of MI with IA and M with 5M were determined. The matrix effect was evaluated in urine samples by the method of standard addition. Good apparent recoveries were found for each indolic compound by the direct method (98–105% with R.S.D. 0.5–5%) and for mixtures of them by the first derivative method (90–105% with R.S.D. 1–5%) indicating the applicability of them with the advantage of their simplicity, low cost in materials, no time consuming and no requirement of the use of a sophisticate calibration program.