[Serum monitoring of methotrexate (MTX) and 7-hydroxymethotrexate concentrations in patients treated with MTX using high-pressure liquid chromatography (HPLC) and comparison of serum MTX levels between HPLC method and fluorescence polarization immunoassay (FPIA)].
In order to measure simultaneously the serum levels of methotrexate (MTX) and its major metabolite, 7-hydroxymethotrexate (7-OH-MTX), in samples obtained from patients treated with MTX, we have investigated the reversed-phase high-pressure liquid chromatographic assay using ion-pairing reagents. The mobile phase consisted of 77.5% solution of 0.005M tetrabutylammonium and 22.5% acetonitrile. SEP-PAK C18 Cartridges were used for the precolumn. The detectable range of MTX and 7-OH-MTX were 0.02-0.03 and 1.0 mumol/l respectively. A significant positive correlation was observed (r = 0.983) between FPIA and HPLC methods. Serum MTX levels with FPIA were significantly (p less than 0.05) higher than those of HPLC method. The serum 7-OH-MTX levels at 24 hr and 48 hr were 4.857 +/- 1.383 (n = 10) and 1.835 +/- 0.286 (n = 6) mumol/l respectively with the dosage of 400 mg/m2. The serum 7-OH-MTX levels at 48 hr were 6.254 +/- 3.053 mumol/l (n = 5) with the dosage of 3,000 mg. The serum half lives of MTX and 7-OH-MTX were 8.05 +/- 1.03 (n = 4) and 14.8 +/- 1.35 (n = 6) hours respectively between 24 hr and 48 hr after administration. The T1/2 7-OH-MTX/MTX ratio was 1.8. Percent cross-reactivity of 7-OH-MTX with concentrations ranging from 1-10 mumol/l were 0.6-2.0% by FPIA. However, patients' serum levels of 7-OH-MTX were 15-85 times (n = 21) higher than those of MTX. MTX levels of containing both MTX and 7-OH-MTX (7-OH-MTX/MTX ratio was 50/1) were significantly higher than those of containing MTX alone by FPIA.
Sakamoto
S
S
Dept. of Pharmacy, Shikoku Cancer Center Hospital.
Matsuhisa
T
T
Miyoshi
K
K
Naruse
R
R
Urayama
H
H
Shimokawa
T
T
Okabe
K
K
Toki
H
H
Motoi
M
M
Moriwaki
S
S
D000328:Adult; D000368:Aged; D002675:Child, Preschool; D002851:Chromatography, High Pressure Liquid; D005260:Female; D016231:Fluorescence Polarization Immunoassay; D006801:Humans; D008228:Lymphoma, Non-Hodgkin / Q000097:blood / Q000188:drug therapy*; D008297:Male; D008727:Methotrexate / Q000031:analogs & derivatives* / Q000097:blood* / Q000627:therapeutic use; D008875:Middle Aged; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000097:blood / Q000188:drug therapy*
C011864:7-hydroxymethotrexate; D008727:Methotrexate