Cr. White et al., COMPARATIVE-STUDY OF HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC METHODS FOR THE DETERMINATION OF TETRACYCLINE ANTIBIOTICS, Journal of liquid chromatography, 16(13), 1993, pp. 2873-2890
This work reviewed all the past HPLC methods utilized for the detectio
n of tetracyclines and created a model to study the most promising one
s. Three general mobile phases were investigated. They included Mobile
Phase I (0.05 M buffer (A)-acetonitrile (B) programmed from 85A:15B t
o 40A:60B), Mobile Phase II (0.05 M buffer (A)-acetonitrile (B)-methan
ol (C) programmed from 80A:OB:20C to 30A:50B:20C), and Mobile Phase II
I (0.05 M buffer (A)-acetonitrile (B)-dimethylformamide (C) programmed
from 80A:OB:20C to 30A:50B:20C). Buffers made from citrate, oxalate,
and phosphate, each prepared at pH 2.0, 4.5, and 7.0, were used with e
ach mobile phase. Ethylenediaminetetraacetic acid (EDTA) at 0.005 M wa
s added to the pH 4.5 and pH 7.0 buffers (it would not dissolve in pH
2.0 buffers). Tetramethylammonium chloride (TMA) was also added at 0.0
1 M to all of the buffers at each pH. The columns chosen for this stud
y were a C18 bonded to silica (Supelco LC-18), a ''deactivated'' C18 b
onded column (Supelco LC-18-DB), and a polymeric styrenedivinylbenzene
copolymer column (Polymer Laboratories PLRP-S). The columns and mobil
e phases were evaluated using a mixed standard of oxytetracycline, tet
racycline, and chlortetracycline. Comparisons were based on peak shape
, separations, and comparative recoveries (integrated areas) of standa
rds. The polymeric column was markedly superior in all respects. Gener
ally optimum results were obtained with Mobile Phase I in pH 2.0 buffe
rs. The oxalate buffer was best with the bonded columns while little d
ifference was noted with the polymeric column. TMA markedly improved p
eak shape on the bonded columns. EDTA was of little benefit. Recoverie
s, especially of chlortetracycline, were higher from the polymeric col
umn.