Table 1.
HPLC gradient program.
Fig 1.
Normalized emission spectra of spent dialysate (solid line) and 4-pyridoxic acid (dotted line) in NaHCO3/acetate buffer pH 7,6.
Excitation at 320 nm.
Fig 2.
Example of an averaged Ex320/Em430 nm chromatogram of the spent dialysate (N = 10).
Samples were collected 7–10 min after the start of the dialysis. Compound 2 was identified as 4-pyridoxic acid. Raised chromatogram is of a reference 4-pyridoxic acid solution (1 μM).
Fig 3.
Comparison of mass spectra (negative ionization) of A) biggest fluorescent peak (no 2, Fig 2) and B) 4-pyridoxic acid reference.
Table 2.
Mean contribution (Mean ± SD, N = 10) values in percentage for main fluorescent peaks in the spent dialysate samples (Ex320/Em430 nm).
Fig 4.
Dependence of the correlation between fluorescence intensity of spent dialysate and 4-pyridoxic acid concentration on the excitation-emission wavelength conditions.
Fig 5.
Correlation between fluorescence intensity of spent dialysate and 4-pyridoxic acid concentration at Ex310 nm (A) and at Em460 (B).
The best correlation was found with Ex310/Em460 nm (Rmax value of 0.95, N = 195).
Fig 6.
An example of the regression line between concentration of 4-PA in spent dialysate and fluorescence intensity (Ex310/Em460 nm, p <0.0001, N = 195).