Fig 1.
Color reaction of Trp by oxidative condensation with aldehydes.
(a) Schematic illustration of Trp coloring reaction with aldehyde based on the Hopkins-Cole reaction. (b) Color reaction of Trp with different reaction solutions: (I) reagent blank (0 mg/L of Trp with 200 g/L of glyoxylic acid monohydrate, 10% HCl, and 3% NaOCl·5H2O), (II) 100 mg/L of Trp with 200 g/L of glyoxylic acid monohydrate and 10% HCl, (III) 100 mg/L of Trp with 10% HCl and 3% NaOCl·5H2O, and (IV) 100 mg/L of Trp with 200 g/L of glyoxylic acid monohydrate, 10% HCl, and 3% NaOCl·5H2O.
Fig 2.
Color changes and absorption spectra of Trp in water with MSG.
(a) Absorption spectra of Trp color reaction with different reaction solutions: (I) reagent blank (0 mg/L of Trp with 200 g/L of MSG, 10% HCl and 3% NaOCl·5H2O), (II) 100 mg/L of Trp with 200 g/L of MSG and 10% HCl, (III) 100 mg/L of Trp added with10% HCl and 3% NaOCl·5H2O, and (IV) 100 mg/L of Trp with 200 g/L of MSG, 10% HCl and 3% NaOCl·5H2O. (b) Color reaction for 100 mg/L of 20 standard amino acids with 200 g/L of MSG, 10% HCl, and 3% NaOCl·5H2O. From upper left to right: Trp, Ser, Gly, Ala, Leu, Arg, Met, His, Thr, and Ile. From lower left to right: Pro, Phe, Lys, Val, Cys, Gln, Glu, Asn, Asp, and Tyr.
Fig 3.
Optical absorption spectra versus concentration of additive reagent.
(a) Absorption spectra for 50 mg/L of Trp and different concentrations of MSG (100, 200, 300, 400, and 500 g/L) with the addition of 10% HCl and 3% NaOCl·5H2O. Reagent blank (0 mg/L of Trp with 400 g/L of MSG, 10% HCl, and 3% NaOCl·5H2O). (b) Absorption spectra for 50 mg/L of Trp with 400g/L of MSG, 10% HCl, and several concentrations of NaOCl·5H2O (1, 3, 5, 10, and 20%). Reagent blank (0 mg/L of Trp with 400 g/L of MSG, 10% HCl, and 3% NaOCl·5H2O). (c) Absorption spectra for various concentrations of Trp (0, 25, 50, and 100 mg/L) after adding 400 g/L of MSG, 10% HCl, and 5% NaOCl·5H2O. (d) Change in the absorption intensity (525 nm) of Trp solution (50 mg/L) over time (1, 3, 5, 7, 10, 15, 20, and 30 min), after adding 400 g/L of MSG, 10% HCl, and 5% NaOCl·5H2O.
Fig 4.
Effects of temperature and pH on absorbance values of Trp.
Change in the absorption intensity (525 nm) of Trp solution (50 mg/L) (a) at different pH and (b) different temperatures, after the addition of 400 g/L of MSG, 10% HCl, and 5% NaOCl·5H2O. Results are the mean ± S.D. of three experiments.
Fig 5.
Effects of interference substances on absorbance values of Trp.
Change of the absorption intensity (525 nm) of Trp solution (50 mg/L) containing several concentrations of (a) AA, (b) glucose, (c) lysozyme, and (d) glyoxylic acid, after the addition of 400 g/L of MSG, 10% HCl, and 5% NaOCl·5H2O.
Fig 6.
Correlation between Trp concentration (10 to 100 mg/L) and absorbance (525 nm) under optimized conditions. Results are the mean ± S.D. of three experiments.
Table 1.
Spectrophotometric determination of Trp in pool specimens (n = 3).