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Imipenem
Molecular formula: C12H17N3O4S Molecular weight: 317.4 CAS Registry No.: 64221-86-9, 74431-23-5 (monohydrate)

SAMPLE Matrix: aqueous humor, blood Sample preparation: 1 mL Plasma or aqueous humor + 1 mL solvent, remove 0.5 mL mixture and add it to 0.5 mL MeOH, mix for 15 min, centrifuge at 4 at 4000 g for 10 min, inject a 50 jxL aliquot of the supernatant. HPLCVARIABLES Guard column: 37-53 [xm Pellicular ODS Column: 250 X 4.6 Viosfer octadecyl (Violet) Mobile phase: MeOH: 100 mM pH 7.2 borate buffer 10:90 Flow rate: 1.5 Injection volume: 50 Detector: UV 313 CHROMATOGRAM Retention time: 5 Limit of detection: 400 (plasma), 150 (aqueous humor) ng/mL KEYWORDS plasma REFERENCE
Carlucci, G.; Biordi, L.; Bologna, M. Human plasma and aqueous humor determination of imipenem by liquid chromatography with ultraviolet detection. J.Liq.Chromatogr., 1993, 16, 23472358

SAMPLE Matrix: blood, tissue, urine Sample preparation: Serum, plasma. Dilute serum or plasma 1:2 to 1:10 with buffer, centrifuge, inject a 20 JJLL aliquot of supernatant. Urine. Dilute urine 1:10 to 1:100 with buffer, centrifuge, inject a 20 JULL aliquot of supernatant. Tissue (lung, gut). Cut tissue with a scalpel, homogenize with 1-3 mL buffer, centrifuge at 9600 g for 5 min three times, inject a 20 JJLL aliquot. Tissue (chondral). Cut tissue with a scalpel, homogenize with 3-6 mL buffer in an ice bath for 2-3 min, centrifuge at 9600 g for 5 min four or five times, inject a 100 JJLL aliquot. Pleural. Dilute human pleural samples with buffer, centrifuge, inject a 20 ^L aliquot. (Buffer was 66.6 mM K2HPO4 adjusted to pH 7.40 with KH2PO4.) HPLC VARIABLES Column: 200 X 4 5 \xm Nucleosil C18 Mobile phase: 15 mM Phosphoric acid adjusted to pH 7.0 with tetrabutylammonium hydroxide Flow rate: 1 Injection volume: 20-100 Detector: UV 313 CHROMATOGRAM Retention time: 4 Limit of detection: 300 ng/mL

OTHER SUBSTANCES Simultaneous: metabolites

KEYWORDS

serum; plasma; lung; gut; pleural; chondral

REFERENCE
Knoller, J.; Konig, W.; Schonfeld, W.; Bremm, K.D.; Roller, M. Application of high-performance liquid chromatography of some antibiotics in clinical microbiology. J.Chromatogr., 1988, 427, 257-267

SAMPLE

Matrix: blood, urine Sample preparation: Add a stabilizing solution of 4-morpholineethanesulfonic acid buffer :ethylene glycol 1:1, prepare ultranltrate using an Amicon Centrifree micropartition system in a centrifuge, inject an aliquot.
HPLCVARIABLES

Column: ixBondapak C18 Mobile phase: MeOH: 100 mM sodium acetate 2:98, pH adjusted to 6.0 with acetic acid Flow rate: 1 Detector: UV 298
CHROMATOGRAM

Retention time: 7.4 Limit of detection: 600 ng/mL

KEYWORDS

plasma
REFERENCE
Paradis, D.; Vallee, R; Allard, S.; Bisson, C; Daviau, N.; Drapeau, C; Auger, R; LeBeI, M. Comparative study of pharmacokinetics and serum bactericidal activities of cefpirome, ceftazidime, ceftriaxone, imipenem, and ciprofloxacin. Antimicrob.Agents Chemother., 1992, 36, 2085-2092

SAMPLE

Matrix: enzyme incubations Sample preparation: Add 2 volumes of MeOH, mix well, centrifuge at 3000 g for 15 min, inject a 20 JJLL aliquot of the supernatant.
HPLCVARIABLES

Column: TSKgel ODS-80Tm (Tosoh) Mobile phase: MeOH: 100 mM pH 7.0 phosphate buffer 4:100 Flow rate: 0.75 Injection volume: 20 Detector: UV 290
CHROMATOGRAM

Limit of quantitation: 1 |xg/mL

REFERENCE
Hikida, M.; Kawashima, K.; Yoshida, M.; Mitsuhashi, S. Inactivation of new carbapenem antibiotics by dehydropeptidase-I from porcine and human renal cortex. J.Antimicrob.Chemother., 1992, 30, 129 134

SAMPLE Matrix: formulations Sample preparation: Dilute 1:100 with mobile phase, inject a 30 jxL aliquot. HPLCVARIABLES Column: 200 X 4.6 RP-8 (Hewlett-Packard) Mobile phase: MeCN: MeOH:buffer 0.4:0.5:99.1, adjusted to pH 7.00 with NaOH (Buffer was 4 mM 3-[N-morpholino]propanesulfonic acid (MOPS) containing 2 g/L sodium hexane sulfate.) Flow rate: 1.8 Injection volume: 30 Detector: UV 250 CHROMATOGRAM Retention time: 5.5 OTHER SUBSTANCES Extracted: cilastatin KEYWORDS injections; total parenteral nutrition; stability-indicating REFERENCE
Zaccardelli, D.S.; Krcmarik, CS.; WoIk, R.; Khalidi, N. Stability of imipenem and cilastatin sodium in total parenteral nutrient solution. J.Parenter.Enteral.Nutr., 1990, 14, 306-309

SAMPLE Matrix: solutions Sample preparation: Inject a 10 jxL aliquot of a 400 |xg/mL solution. HPLCVARIABLES Column: 150 X 4.6 Microsorb C8 80-315 Mobile phase: 1 mM KH2PO4 adjusted to pH 6.8 with 500 mM NaOH Flow rate: 1.5 Injection volume: 10 Detector: UV 300 REFERENCE
Connolly, M.; Debenedetti, P.G.; Tung, H.-H. Freeze crystallization of imipenem. J.Pharm.ScL, 1996,85, 174-177

SAMPLE Matrix: solutions Sample preparation: Prepare a 2.5-5 jxg/mL solution, inject a 20 jxL aliquot. HPLCVARIABLES Column: 80 X 4.6 3.65 jim Zorbax Rx-SIL (similar to Zorbax SB-C8 (Mac-Mod Analytical)) Mobile phase: MeCN: buffer 0.5:99.5 (Buffer was 0.1% acetic acid adjusted to pH 7 with ammonium hydroxide.) Flow rate: 1 Injection volume: 20 Detector: UV 296 CHROMATOGRAM Retention time: k' 3.5

REFERENCE
Kirkland, K.M.; McCombs, D.A.; Kirkland, J.J. Rapid, high-resolution high-performance liquid chromatographic analysis of antibiotics. J.Chromatogr.A, 1994, 660, 327-337

SAMPLE

Matrix: solutions
HPLCVARIABLES

Column: 150 X 4.6 PLRP-S styrene-divinylbenzene copolymer (Polymer Labs) Mobile phase: Gradient. MeCN: 20 mM pH 7.2 KH2PO4-NaOH buffer 3:97 to 7:93 in 22 min. Column temperature: 40 Flow rate: 1.2 Detector: UV 295
CHROMATOGRAM

Retention time: 7
OTHER SUBSTANCES

Simultaneous: degradation products

KEYWORDS pH >4 buffer REFERENCE
Smith, G.B.; Dezeny, G.C.; Douglas, A.W. Stability and kinetics of degradation of imipenem in aqueous solution. J.Pharm.ScL, 1990, 79, 732-740

SAMPLE

Matrix: solutions
HPLCVARIABLES

Column: 250 X 4.6 Partisil PXS 5/25 PAC Mobile phase: Gradient. MeCN: water from 25:75 to 50:50 in 30 min. Flow rate: 2 Detector: UV 320
CHROMATOGRAM

Retention time: 6
OTHER SUBSTANCES

Simultaneous: degradation products

KEYWORDS pH 4 buffer REFERENCE
Smith, G.B.; Dezeny, G.C.; Douglas, A.W. Stability and kinetics of degradation of imipenem in aqueous solution. J.Pharm.ScL, 1990, 79, 732-740

SAMPLE

Matrix: solutions Sample preparation: Inject a 25 fxL aliquot.

HPLCVARIABLES

Column: 100 X 4 5 \xm ODS-Hypersil Mobile phase: MeCN: 10 mM ammonium acetate 20:80 Flow rate: 2 Injection volume: 25 Detector: UV 300
OTHER SUBSTANCES

Also analyzed: penicillin G (UV 227)

REFERENCE
Eley, A.; Greenwood, D. Beta-lactamases of type culture strains of the Bacteroides fragilis group and of strains that hydrolse cefoxitin, latamoxef and imipenem. J.Med.MicrobioL, 1986, 21, 49-57

ANNOTATED BIBLIOGRAPHY

Jenke, D.R. Drug binding by reservoirs in elastomeric infusion devices. Pharm.Res., 1994, 11, 984-989 [formulations; saline; 5% dextrose; simultaneous cilastatin] Ebey, W.J.; Boucher, B.A.; Pieper, J.A. A rapid HPLC method for determination of imipenem in plasma. J.Liq.Chromatogr., 1988, 11, 3471-3481 [plasma; LOD 1000 ng/mL] Krausse, R.; Ullmann, U. Determination of imipenem and cilastatin in serum and tissue by high-pressure liquid chromatography. Infection, 1986, 14, 243245 Gravallese, D.A.; Musson, D.G.; Pauliukonis, L.T.; Bayne, W.F. Determination of imipenem (N-formimidoyl thienamycin) in human plasma and urine by high-performance liquid chromatography, comparison with microbiological methodology and stability. J.Chromatogn, 1984, 310, 7184 [plasma; urine; serotonin (IS); ultrafiltrate; LOD 300 ng/mL (plasma); LOD 1 |xg/mL (urine); non-interfering cilastatin; pharmacokinetics] Myers, CM.; Blumer, J.L. Determination of imipenem and cilastatin in serum by high-pressure liquid chromatography. Antimicrob.Agents Chemother., 1984, 26, 78-81

Indapamide
Molecular formula: C16H16CIN3O3S Molecular weight: 365.8 CAS Registry No.: 26807-65-8

SAMPLE Matrix: blood Sample preparation: 1 mL Whole blood + 1 mL 4 jmg/mL glipizide in 50 mM pH 6.6 KH2PO4, vortex, add 6 mL diethyl ether, shake at 150 20 oscillations/min on a reciprocating shaker for 15 min, centrifuge at 1500 g for 10 min. Remove the organic layer and evaporate it to dryness at 37 under a stream of nitrogen. Dissolve residue in 200 JJIL mobile phase, centrifuge at 1000 g for 3 min, inject a 40 |xL aliquot of the supernatant. HPLCVARIABLES Column: 150 x 4.6 5 |xm Nucleosil C18 Mobile phase: MeCN:isopropanol:buffer 30:5:65 (Buffer was 80 mM ammonium acetate adjusted to pH 3.5 with concentrated HCl.) Flow rate: 1 Injection volume: 40 Detector: UV 241 CHROMATOGRAM Retention time: 5.2 Internal standard: glipizide (5.9) Limit of quantitation: 10 ng/mL OTHER SUBSTANCES Noninterfering: acetaminophen, aspirin, caffeine, dextromethorphan, ibuprofen, nicotine, phenylpropanolamine, theophylline KEYWORDS whole blood REFERENCE
Miller, R.B.; Dadgar, D.; Lalande, M. High-performance liquid chromatographic method for the determination of indapamide in human whole blood. J.Chromatogr., 1993, 614, 293-298

SAMPLE Matrix: blood Sample preparation: Condition a Sep-Pak C18 SPE cartridge with 5 mL MeOH and 5 mL 10 mM HCl. 2 mL Plasma + 2 mL 2 |ig/mL IS in water + 10 mL 10 mM HCl, add to the SPE cartridge, wash with 4 mL 10 mM HCl, elute with 1 mL MeOH, inject a 20 \xL aliquot of the eluate. HPLCVARIABLES Column: 250 X 4 10 [xm LiChrosorb SI 60 ODS Mobile phase: MeCN: 10 mM pH 3.5 sodium phosphate 30:70 Flow rate: 1 Injection volume: 20 Detector: UV 241 CHROMATOGRAM Retention time: 9

Internal standard: sulfadimethoxime (6.2) Limit of detection: 10 ng/mL KEYWORDS plasma; SPE; see Anal.Abs. 1986, 48, 12D80 REFERENCE
Gaetani, E.; Laureri, CR; Vitto, M.; Borghi, L.; Elia, G.F.; Novarini, A. Determinazione deirindapamide nel plasma mediante HPLC [Determination of indapamide in plasma by HPLC]. Boll.Chim.Farm., 1986, 125, 35-37

SAMPLE Matrix: blood Sample preparation: Keep tubes in crushed ice except when being processed throughout this procedure. 2 mL Plasma + 100 jxL 10 |xg/mL sulfanilamide in MeCN + 8 mL diethyl ether, vortex for 2 min, centrifuge at 4. Remove ether layer and add it to 1 mL 100 mM NaOH, vortex for 2 min, centrifuge at 4. Remove aqueous layer and add it to 1 mL 100 mM HCl and 500 jxL 50 mM pH 7.4 sodium phosphate, add 8 mL ether, vortex for 2 min, centrifuge at 4. Evaporate ether layer to dryness, reconstitute in 200 |xL mobile phase, inject 50 fxL aliquot. HPLCVARIABLES Column: 250 X 4.6 5 jxm Zorbax ODS Mobile phase: MeCN: 100 mM pH 3.6 sodium acetate buffer 43:57 Column temperature: 54 Flow rate: 1 Injection volume: 50 Detector: UV 241 CHROMATOGRAM Retention time: 6.3 Internal standard: sulfanilamide (5.3) Limit of detection: 25 ng/mL KEYWORDS plasma REFERENCE
Choi, R.L.; Rosenberg, M.; Grebow, RE.; Huntley, T.E. High-performance liquid chromatographic analysis of indapamide (RHC 2555) in urine, plasma and blood. J.Chromatogr., 1982, 230, 181-187

SAMPLE Matrix: bulk, formulations, urine Sample preparation: Bulk, tablets. Weigh out amount containing 10 mg indapamide, dissolve in 10 mL 150 ixg/mL sulfisoxazole in MeOH, make up to 100 mL with MeOH, inject an aliquot. Urine. 1 mL Urine + 1 mL 150 |xg/mL sulfisoxazole in MeOH H - 10 mL ethyl acetate, vortex, centrifuge. Remove the organic layer and evaporate it to dryness under a stream of nitrogen, reconstitute the residue in 1 mL mobile phase, inject a 10 |xL aliquot. (If indapamide concentration is >100 ng/mL mix 1 mL urine, 1 mL 150 |xg/mL sulfisoxazole in MeOH, and 8 mL mobile phase, vortex, inject a 10 JJLL aliquot.) HPLCVARIABLES Guard column: 20 X 4 Bondapak C18 Corasil Column: 300 X 4 10 |xm jxBondapak C18 Mobile phase: MeCN:buffer 35:65 (Buffer was water adjusted to pH 2.8 with 10% phosphoric acid.)

Flow rate: 2 Injection volume: 10 Detector: UV 254 CHROMATOGRAM Retention time: 7 Internal standard: sulfisoxazole (4) Limit of detection: 25 ng/mL OTHER SUBSTANCES Extracted: metabolites KEYWORDS tablets REFERENCE
Pietta, P.; Calatroni, A.; Rava, A. High-performance liquid chromatographic assay for monitoring indapamide and its major metabolite in urine. J.Chromatogr., 1982, 228, 377-381

SAMPLE Matrix: solutions HPLC VARIABLES Column: Chiralcel OD-R Mobile phase: MeCN: water 40:60 Column temperature: 40 Flow rate: 1 Detector: UV 254 CHROMATOGRAM Retention time: 11.3, 15.8 (enantiomers) KEYWORDS chiral REFERENCE
Application Guide for Chiral Column Selection, Second Edition, Chiral Technologies Inc., Exton PA, 1995, p. 39

SAMPLE Matrix: solutions HPLCVARIABLES Column: 250 X 4.6 Chirex 3022 (Phenomenex) Mobile phase: Hexane: 1,2-dichloroethane: EtOH/trifluoroacetic trifluoroacetic acid was premixed 20:1.) Flow rate: 0.7-1 Injection volume: 20 Detector: UV 248 KEYWORDS chiral; a = 1.08 REFERENCE
Cleveland, T. Pirkle-concept chiral stationary phases for the HPLC separation of pharmaceutical racemates. J.Liq.Chromatogr., 1995, 18, 649-671

acid 58:35:7 (EtOH/

SAMPLE Matrix: solutions Sample preparation: Inject a 20 J U L L aliquot. HPLCVARIABLES Column: 250 X 4.6 5 |xm Lichrosorb RP-18 Mobile phase: MeOH.buffer 50:50 (Buffer was 1% aqueous acetic acid containing 0.2% triethylamine.) Flow rate: 1 Injection volume: 20 Detector: UV 250 CHROMATOGRAM Retention time: 11 Limit of detection: 500 ng/mL OTHER SUBSTANCES Simultaneous: degradation products KEYWORDS stability-indicating REFERENCE
Padval, M.V.; Bhargava, H.N. Liquid chromatographic determination of indapamide in the presence of its degradation products. J.Pharm.Biomed.Anal, 1993, 11, 1033-1036

ANNOTATED BIBLIOGRAPHY

Chen, D. [Determination of indapamide in human serum by high performance liquid chromatography]. Chung Kuo I Hsueh Ko Hsueh Yuan Hsueh Pao, 1990, 12, 286-289