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Effect of Storage Temperature and Ph on the Stability of Antimicrobial Agents in MIC Trays JONZ-MEI DEBRA HWANG, TONI E

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Effect of Storage Temperature and Ph on the Stability of Antimicrobial Agents in MIC Trays JONZ-MEI DEBRA HWANG, TONI E JOURNAL OF CLINICAL MICROBIOIOGY, May 1986. p. 959-961 Vol. 23, No. S 0095-1137/86/050959-03$02.00/0 Copyright © 1986. American Society for Microbiology Effect of Storage Temperature and pH on the Stability of Antimicrobial Agents in MIC Trays JONZ-MEI DEBRA HWANG, TONI E. PICCININI, CLAUDIA J. LAMMEL, W. KEITH HADLEY, AND GEO F. BROOKS* Departnent of Laboratorv Medicine, University of Califo)r-nia, Sani Friancisco, Califorzia 94143 Received 25 November 1985/Accepted 15 January 1986 Twelve antimicrobial agents, ampicillin, aztreonam, cefamandole, cefazolin, cefonicid, ceforanide, ceftazi- dime, ceftizoxime, ceftriaxone, cefuroxime, ciprofloxacin, and norfloxacin, were prepared at pH 6.80 and 7.31 in microdilution trays for storage at 4, -10, -25, and -70°C and for weekly susceptibility testing. All 12 drugs had stable biological activity when stored at -70°C for I year. All but ampicillin and aztreonam were stable at -25°C. Storage at -10°C was least satisfactory. Desiccation occurred at 40C, but short-term storage at this temperature is possible since the antimicrobial agents are stable for up to several months. The routine use of microdilution trays for antimicrobial aireius ATCC 29213, Escherichia (oli ATCC 25922. and susceptibility testing requires a knowledge of the stability of Pseuidomizonas aeruigin.osa ATCC 27853, were the same as in the antimicrobial agents when stored at different tempera- the previous study (1). These bacteria had at least one and tures. A previous report from our laboratory showed data for usually two endpoints within the range of drug concentra- 11 beta-lactams prepared at two pHs and stored at four tions selected. MICs were determined by the methods pre- temperatures over 1 year (1). The data indicated that the viously described (1). drugs were stable when stored at -70°C; some of the To estimate desiccation in the microdilution tray wells, the antimicrobial agents were stable when stored at -25°C; 4°C broth in the first three wells containing cefazolin (the tray was suitable for short-term storage; and the antimicrobial used for Pseiudounionias testing) was pooled, and sodium agents were most unstable when stored at -10°C. In this concentrations were tested weekly by the ion-selective elec- paper, we report data from a second study of the stability of trode method in a Beckman Astra 8 (Beckman Instruments, antimicrobial agents in microdilution trays. For comparison. Inc., Brea, Calif.). The results are shown in Table 2. The three drugs were included in both studies; nine drugs were sodium concentrations remained stable in the trays stored at studied for the first time. -25 and -70°C but increased by 7 and 17%, respectively, at The drugs, manufacturers, and dilutions tested were as -10 and 4°C. The trays stored at 4°C were tested for 26 follows: cefamandole (Eli Lilly & Co., Indianapolis. Ind.), weeks because of the desiccation. cefonicid (Smith Kline & French Laboratories, Philadelphia, The MIC data were recorded and entered into a spread Pa.), ceforanide (Bristol Laboratories, Syracuse, N.Y.), and sheet program (Lotus Development Corp., Cambridge, aztreonam (E. R. Squibb & Sons, Princeton. N.J.), each at Mass.) by using an International Business Machines (Boca 0.125 to 16 p±g/ml; cefazolin (Sigma Chemical Co., St. Louis. Raton, Fla.) PC-XT computer. The data were plotted, and Mo.), 0.125 to 8 ,ug/ml; cefuroxime (Glaxo Inc., Research the half-lives of biological activity in weeks were determined Triangle Park, N.C.), ceftizoxime (Smith Kline & French). by the least-squares fit of the log, plots of antimicrobial ceftriaxone (Hoffmann-La Roche Inc., Nutley, N.J.), and concentration versus time (1). ceftazidime (Glaxo), each at 0.25 to 32 jig/ml; ampicillin The data showing the stability of the 12 antimicrobial (Wyeth Laboratories. Philadelphia, Pa.), 0.25 to 16 ,ug/ml; agents prepared at final pHs of 6.80 and 7.31 and stored at and ciprofloxacin (Miles Laboratories, Inc., Elkhart, Ind.) four temperatures are shown in Table 3. The three drugs and norfloxacin (Merck Sharp & Dohme, West Point, Pa.), included in the previous study (1), ampicillin, cefazolin, and each at 0.06 to 8 p.g/ml. The drugs were tested at twofold ceftriaxone, yielded similar results in the current study, serial dilutions. except that ampicillin at pH 6.80 and stored at -10°C had a The antimicrobial agents were dissolved according to the slightly longer half-life in the present study, and ceftriaxone specifications of the manufacturer. Microdilution trays were stored at -10°C had a slightly longer half-life for both pH prepared and stored as previously reported (1). The storage 6.80 and 7.31 when tested against the PFseiudo,nonas strain. temperatures and manual defrost freezers and refrigerators The optimum temperature for storage of microdilution were as follows: -70°C, model ULT 1785 Revco Ultra-Low, MIC test is as Revco, Inc., West Columbia, S.C.; -25°C, model UC trays -70°C, indicated by these and our 25CW-FMS Kelvinator, Commercial Products, Inc., Manitowac, Wis.; -10°C, model 3551-1000 Lab-Line refrig- erator-freezer, Lab-Line Instruments, Melrose Park, Ill.; TABLE 1. Temperature stability of refrigerator and freezers and 4°C, model LA 77 LC refrigerator, General Electric Co., Storage No. of Mean temp (°C) + SD Schenectady, N.Y. The means of the maximum and mini- temp (°C) observations Minimum" Maximum" Observed mum temperatures for each week and the observed temper- atures are listed in Table 1. 4 26 1.9+ 1.4 6.9±3.2 3.0± 1.1 The three bacterial test strains used, Staphvlococcus -10 50 -16.3 + 3.8 -4.5 + 3.6 -11.0 + 2.7 -25 50 -31.0 + 2.2 -14.0 + 10.1 -26.2 ± 2.3 -70 49 -70.8 ± 4.7 * Corresponding author. " From temperatures measured by maximum-minimum thermometers. 959 960 NOTES J. CLIN. MICROBIOL. TABLE 2. Sodium concentrations in Mueller-Hinton broth TABLE 3-Continued medium from antimicrobial microdilution trays measured by the ion-selective electrode method Half-life (wk) at indicated test time Drug, pH, and and temp No. tested' Storage temp (OC) Mean sodium concn t % Changeb organism tested 26 wk, 52 wk, 52 wk, 52 wk, SD (mmol per liter) 4°C -10°C -250C -700C 26 4 140.0 ± 23.2 +21.5 52 -10 128.6 ± 24.5 -3.9 E. coli .13 .26 .26 2226 52 -25 120.1 ± 3.6 -4.8 pH 7.31 52 -70 120.0 ± 3.1 -5.0 S. aureus .-1L3 (26) .26 .26 .226 E. coli .:-1L3 .26 .26 .226 a Sodium concentrations were measured each week when the MIC testing was done. Fifty-two determinations were made for storage temperatures of -10, -25, and -70°C, and 26 determinations were made for 4°C. Cefonicid b Percent change from the mean of the first sodium concentration for all pH 6.80 temperatures to the mean of the concentrations of the last three determina- S. aureus .13 (23) .26 .26 .26 tions for the individual temperatures. E. coli .13 .26 .26 .26 pH 7.31 S. aureus .13 18 .26 .26 E. coli .13 22 .26 .26 previously published results (1). With the exception of ampicillin at pH 7.31 and aztreonam at both pH 6.80 and 7.31 Ceforanide -25°C is also suitable for storing all the antibiotics studied. pH 6.80 In our previous study, 6 of the 11 antibiotics, mezlocillin, S. aureus .13 .26 .26 226 piperacillin, ticarcillin, azlocillin, moxalactam, and ampicil- E. coli .13 .26 .26 .26 lin, showed decreased half-life activity at pH 7.31 when pH 7.31 S. aureus .12.5 .26 .26 .26 stored at -25°C. In the present study, that pattern was not E. coli .13 .26 .26 apparent with the nine additional antibiotics (Table 3). The .26 shorter half-life of some of the antimicrobial agents noted in Ceftazidime the previous study compared with that of the present study pH 6.80 may have been caused by freezer breakdown; the data, S. aureus .13 (25) .26 .26 226 however, did not show shifts in biological activity directly P. aeruginosa .13 (26) 23 .26 .26 related to freezer problems. There were no freezer break- pH 7.31 downs during the present study. S. aureus 11.5 6 .26 226 In the previous study (1), some of the drugs were more P. aeruginosa 12.5 8.5 .-26 .26 stable when made up at pH 6.80 than at 7.31. In this study, Ceftizoxime enhanced stability at pH 6.80 was again noted for ampicillin, pH 6.80, S. aureus .13 .26 .26 .26 aztreonam, cefonicid, ceftazidime, and cefuroxime. pH 7.31, S. aureus 212.5 .26 .26 .26 Norfloxacin, ciprofloxacin, ceftriaxone, ceftizoxime, cefa- mandole, cefazolin, and ceforanide were stable at both pH Ceftriaxone 6.80 and 7.31 and -10, -25, and -70°C. Cefonicid and pH 6.80 S. aureus .13 .:26 .26 .26 P. aeruginosa .13 .:26 .26 .26 TABLE 3. Half-lives of antimicrobial activity for 10 beta-lactams pH 7.31 and 2 quinolones at pHs 6.80 and 7.31 S. aureus 212.5 .:-26 .26 .26 Half-life (wk) at indicated test time P. aeruginosa .12.5 .-26 .26 .26 Drug, pH, and and temp Cefuroxime organism tested 26 wk, 52 wk, 52 wk, 52 wk, pH 6.80 4°C -10°C -25°C -700C S. aureus .-13 (23) .::26 .26 .26 Ampicillin E. coli .:13 (26) .::26 .26 .26 pH 6.80 pH 7.31 S. aureus 213 (26)a 17.5 226 226 S. aureus .-12.5 18 .:26 .26 E.
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