Prostatic acid phosphatase assay with self-indicating substrate 2,6-dichloro-4-acetylphenyl phosphate.
We characterized six self-indicating substrates, synthesized as the derivative compounds of acetylphenyl phosphate, for serum prostatic acid phosphatase (PAP) activity. One of the substrates, 2,6-dichloro-4-acetylphenyl phosphate (DCAPP), is superior to others in terms of stability, affinity, and low Km for PAP. The hydrolyzed product, 2,6-dichloro-4-acetylphenol (DCAP), has a maximum absorption at 334.2 nm, a pKa of 4.15, and a molar absorptivity at 340 nm of 21,490 L.mol-1.cm-1 in citrate-HCl buffer, pH 5.4. PAP activity was assessed by subtracting tartaric acid-inhibited acid phosphatase activity from total acid phosphatase activity. Our assay system involving DCAPP is a unique kinetic method that shows good reproducibility, wide analytical dynamic range, and high specificity for PAP. Moreover, it is easily adaptable to automated analyzers because the product, DCAP, can be monitored at 340 nm.
Osawa
S
S
Department of Laboratory Medicine, Chiba University Hospital, Japan.
Iida
S
S
Yonemitsu
H
H
Kuroiwa
K
K
Katayama
K
K
Nagasawa
T
T
D000135:Acid Phosphatase / Q000097:blood*; D002021:Buffers; D004355:Drug Stability; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008297:Male; D010755:Organophosphates / Q000378:metabolism*; D011467:Prostate / Q000201:enzymology*; D012680:Sensitivity and Specificity; D013053:Spectrophotometry; D013379:Substrate Specificity
C092308:2,6-dichloro-4-acetylphenyl phosphate; D002021:Buffers; D010755:Organophosphates; D000135:Acid Phosphatase