A
solution of 0.30 g Na2CO3 in 8 mL of boiling water was placed in a
crucible. Then, salep phosphate (0.50 g) was added to the solution.
The moist paste was dried at 100 C prior to ashing in a muffle furnace
at 550 C for 7 h. After cooling to room temperature, 12 mL HCl
(4%) in 10 mL distilled water was added. Then, the solution was
transferred into a beaker and filled up to a volume of 50 mL with
distilled water. The filtrate was diluted to 250 mL with distilled
water and 5 mL thereof was mixed with 2 mL of vanadate-molybdate
reagent. The sulfuric ammonium molybdate (NH4)2MoO4 rapidly
reacts with orthophosphates to form molybdophosphoric acid
H3[P(Mo3O10)4]. In the presence of vanadium, an intensive/stable
yellow color [vanadomolybdophosphoric acid (H3+nPVnMo12nO40)]
forms proportionally to the phosphate concentration. This color
complex was quantified photometric. Its absorption maximum is
located at 360 nm and was measured by an UV–vis spectrometer
(Shimadzu UV–visible 1650 PC) 45 min after adding the vanadate–
molybdate reagent to the samples.