on dry ice and transported in frozen state to the laboratory for analy¬ sis. Aqueous humor samples were supplied to the laboratory identified only by patient number to permit a blind laboratory analy¬ sis for chloramphenicol. All samples were stored at -20 C until analyzed. After thawing at room tempera¬ ture, the chloramphenicol was extracted into ethyl acetate and spotted along with appropriate standards on scored (1-cm channels) silica gel thin-layer chromatography (TLC) plates within 18 hours after collection. The spotted plates were prewashed in hexane, dried, and chromatographed in chloroform/methanol/acetic acid (90:10:5). The dried plates were quantitated at 270 mm on a spectrodensitometer (Schoeffel). The active compound was analysed for identification in situ on the TLC plate by two different procedures: (1) reduction of the p-nitrophenyl group to phenylamino followed by coupling with dimethylaminobenzaldehyde and (2) the use of iodoplatinate spray reagent in visualization of the migrating component." Procedures for lo¬ cation and identification of possible me¬ tabolites were done by measurement of the ratio of migration of the spot to the solvent front (Rf) and the reductive/coupling pro¬ cedure described above.
ately placed a
Chloramphenicol in Aqueous Humor After Topical Application Harold
Beasley, MD; Jack
J.
Boltralik, MS; Henry A. Baldwin
Chloramphenicol 0.5% ophthalmic solution was applied topically to patients at various times before cataract surgery. The aqueous humor was obtained at the time of surgery and analyzed for chloramphenicol content by thin-layer chromatography. Aqueous humor chloramphenicol levels ranged from 3.5\g=m\g/ml to 6.7\g=m\g/ml at the initial sampling period one to two hours following topical administration. Chloramphenicol was measurable in samples for up to five hours following administration. The compound measured in the aqueous humor samples was chemically identified as intact chloramphenicol, and no metabolites of chloramphenicol were shown.
Chloramphenicol
has been used for many years in the treatment of and as prophylaxis against bacterial endophthalmitis. In addition to a broad spectrum of antibacterial activ¬ ity, chloramphenicol has been shown to penetrate ocular tissues, includ¬ ing aqueous and vitreous humor, af¬ ter systemic administration in rab¬ bits1'4 and humans.^ " However, adverse reactions such as
aplastic anemia, granulocytopenia, hypoplastic anemia, and thrombocytopenia resulting from systemic ad¬
ministration of chloramphenicol in humans have been well documented.
Although one case of bone marrow hypoplasia has been reported after long-term topical application of 0.5% chloramphenicol,7 preoperative ad¬ ministration of chloramphenicol eye drops continues to be utilized as a popular method for preventing bacte¬ rial endophthalmitis. Unfortunately,
aqueous humor levels of chloram¬ after topical applications in humans has not been as well defined
phenicol
as drug levels after systemic adminis¬ tration. It was, therefore, the intent of this investigation to determine these drug levels after topical, presurgical administration of chloram¬
phenicol eye drops. Subjects and
Eighteen patients undergoing elective cataract extraction surgery were chosen for study. At various time intervals VÁ to 5 hours before surgery, 14 of the 18 patients received topically a commercially avail¬ able preparation of 0.5% chloramphenicol
(Econochlor). The remaining four patients received similar topical applications of gentamicin sulfate (Garamycin Ophthal¬ mic Solution) to serve as a negative control (blank) for the laboratory analysis for chloramphenicol. Patients received doses according to a computer-derived random¬ ization code; each patient received two drops (0.1 ml) of test medication every five minutes for a total of six doses (3 mg of topical chloramphenicol over a 30-minute period). All patients
Submitted for publication Dec 28, 1973. From the Department of Ophthalmology, Alcon Laboratories, Inc. (Mr. Boltralik and Baldwin) and St. Joseph Hospital (Dr. Beasley), Fort Worth, Tex. Reprint requests to 1212 W Presidio, Fort Worth, TX 76102 (Dr. Beasley).
Methods
were
prepared
as
usual for
surgery; a limbal groove was made, and su¬ tures were preplaced. Using a sterile 27gauge needle attached to a 1-ml disposable syringe, an anterior chamber paracentesis
accomplished by inserting the needle through the groove. The syringe contain¬ ing the aqueous humor was then immediwas
Downloaded From: http://archopht.jamanetwork.com/ by a Michigan State University User on 06/18/2015
Results
Patients utilized in this study in age from 54 to 88 years with 4 men and 14 women being en¬ rolled. Each patient was found to be free from concomitant ocular disease, and all cataracts were classified as
ranged
degenerative type. topical drug application sequence, no gross signs of ocular irritation were noted in either the chloramphenicol or con¬ being
of the
On examination after the
trol groups. The results of the labora¬ tory analysis for chloramphenicol are shown in the Figure. Depicted are the actual values of chloramphenicol for each sample analyzed along with the calculated least-squares line. The quantity of aqueous humor supplied to the laboratory was insufficient for analysis in two cases, and no chloram¬ phenicol was identified in four blind control samples. The active compound (intact chlo¬
ramphenicol) migrated on all TLC plates identically to the applied stan¬ dard, whereas control (blank) samples showed neither ultraviolet absorbing components in the vicinity of the
divided topical dose, maximum aqueous humor levels of chloram¬ phenicol (3ßg/ml to 6^g/ml) would be similar to those obtained after a single 1.0-gm oral dose. Our results also indicate that concentrations of 3µg/ml would be present in the aqueous humor for approximately two hours after the 3.0-mg topical dose. From the standpoint of sys¬ temic toxicity associated with oral ad¬ ministration, the advantage of topi¬ cal administration of chloramphenicol is obvious. Since the compound extracted from the aqeuous humor migrated on the TLC plates identically to the applied standard and no metabolites of chlor¬ amphenicol were identified, we have concluded that the major active com¬ pound measured in this study was in¬ tact chloramphenicol. However, small quantities of metabolites that were below the sensitivity limits of detec¬ tion may have been present and can¬ not be ruled out as a possibility.
Busby, Ms. Bette McCue, and Dr. R. supplied technical assistance for the laboratory analysis of chloramphenicol. Ms. E. Adamski
Informed consent
was
obtained from the sub¬
jects. Chloramphenicol 0.5% (Econochlor) was sup¬ plied by Alcon Laboratories, Inc., Fort Worth, Tex.
References Hours
Topical Administration
analysis for chloramphenicol in human aqueous humor after topical application (individual samples and least-squares line). Results of TLC
standard
nor
chemical reactions that
identify chloramphenicol. Evidence of enzymatic hydrolysis of the dichloroacetyl group to produce the chlor¬ amphenicol metabolite of D-threo-1-pnitrophenyl-2-amino-l, 3-propanediol was also absent on all plates. Comment
Following oral administration of chloramphenicol in humans, blood lev¬ els have been found to be roughly pro¬ portional to the size of the dose,
reaching a maximum in two to four hours. For example, peak serum levels averaged 4u.g/ml after a single 0.5gm dose, 9µg after 1.0 gm and l^g after 2.0 gm.3 Abraham and Burnett have reported that from 33% to 79% (mean, 52%) of the blood concentra¬ tion of chloramphenicol passes into the primary aqueous humor after each oral administration.6 Correlation of these blood-aqueous levels with the results of the present experiment would indicate that, utilizing a 3.0-mg
1. Leopold IH, Nichols AC, Vogel AW: Penetration of chloramphenicol U.S.P. (Chloromycetin) into the eye. Arch Ophthalmol 44:22-36,1950. 2. Sorsby A, Ungar J, Crick RP: Aureomycin, chloramphenicol, and Terramycin in ophthalmology. Br Med J 2:301-304, 1953. 3. Bleeker GM, Maas ER: The penetration of Aureomycin, Terramycin and chloramphenicol in the ocular tissues. Ophthalmologica 130:1-8,1955. 4. Langham M: Factors affecting the penetration of antibiotics into the aqueous humor. Br J Ophthalmol 35:614-620, 1951. 5. Leopold IH: Clinical trial with chloramphenicol in ocular infections. Arch Ophthalmol 45:44-52, 1951. 6. Abraham RK, Burnett HH: Tetracycline and chloramphenicol studies on rabbit and human eyes. Arch Ophthalmol 54:641-659, 1955. 7. Rosenthal RL, Blackmon A: Bone marrow hypoplasia following use of chloramphenicol eye drops. JAMA 191:136-137, 1965. 8. Dawson RMC, Elliott DC, Elliott WH, et al (eds): Data for Biochemical Research, ed 2. Oxford University Press, 1969, p 538. 9. Ley HL Jr, Smadel JE, Crocker TT: Administration of Chloromycetin to normal human subjects. Proc Soc Exp Biol Med 68:9-12, 1948.
Downloaded From: http://archopht.jamanetwork.com/ by a Michigan State University User on 06/18/2015
ately placed a
Chloramphenicol in Aqueous Humor After Topical Application Harold
Beasley, MD; Jack
J.
Boltralik, MS; Henry A. Baldwin
Chloramphenicol 0.5% ophthalmic solution was applied topically to patients at various times before cataract surgery. The aqueous humor was obtained at the time of surgery and analyzed for chloramphenicol content by thin-layer chromatography. Aqueous humor chloramphenicol levels ranged from 3.5\g=m\g/ml to 6.7\g=m\g/ml at the initial sampling period one to two hours following topical administration. Chloramphenicol was measurable in samples for up to five hours following administration. The compound measured in the aqueous humor samples was chemically identified as intact chloramphenicol, and no metabolites of chloramphenicol were shown.
Chloramphenicol
has been used for many years in the treatment of and as prophylaxis against bacterial endophthalmitis. In addition to a broad spectrum of antibacterial activ¬ ity, chloramphenicol has been shown to penetrate ocular tissues, includ¬ ing aqueous and vitreous humor, af¬ ter systemic administration in rab¬ bits1'4 and humans.^ " However, adverse reactions such as
aplastic anemia, granulocytopenia, hypoplastic anemia, and thrombocytopenia resulting from systemic ad¬
ministration of chloramphenicol in humans have been well documented.
Although one case of bone marrow hypoplasia has been reported after long-term topical application of 0.5% chloramphenicol,7 preoperative ad¬ ministration of chloramphenicol eye drops continues to be utilized as a popular method for preventing bacte¬ rial endophthalmitis. Unfortunately,
aqueous humor levels of chloram¬ after topical applications in humans has not been as well defined
phenicol
as drug levels after systemic adminis¬ tration. It was, therefore, the intent of this investigation to determine these drug levels after topical, presurgical administration of chloram¬
phenicol eye drops. Subjects and
Eighteen patients undergoing elective cataract extraction surgery were chosen for study. At various time intervals VÁ to 5 hours before surgery, 14 of the 18 patients received topically a commercially avail¬ able preparation of 0.5% chloramphenicol
(Econochlor). The remaining four patients received similar topical applications of gentamicin sulfate (Garamycin Ophthal¬ mic Solution) to serve as a negative control (blank) for the laboratory analysis for chloramphenicol. Patients received doses according to a computer-derived random¬ ization code; each patient received two drops (0.1 ml) of test medication every five minutes for a total of six doses (3 mg of topical chloramphenicol over a 30-minute period). All patients
Submitted for publication Dec 28, 1973. From the Department of Ophthalmology, Alcon Laboratories, Inc. (Mr. Boltralik and Baldwin) and St. Joseph Hospital (Dr. Beasley), Fort Worth, Tex. Reprint requests to 1212 W Presidio, Fort Worth, TX 76102 (Dr. Beasley).
Methods
were
prepared
as
usual for
surgery; a limbal groove was made, and su¬ tures were preplaced. Using a sterile 27gauge needle attached to a 1-ml disposable syringe, an anterior chamber paracentesis
accomplished by inserting the needle through the groove. The syringe contain¬ ing the aqueous humor was then immediwas
Downloaded From: http://archopht.jamanetwork.com/ by a Michigan State University User on 06/18/2015
Results
Patients utilized in this study in age from 54 to 88 years with 4 men and 14 women being en¬ rolled. Each patient was found to be free from concomitant ocular disease, and all cataracts were classified as
ranged
degenerative type. topical drug application sequence, no gross signs of ocular irritation were noted in either the chloramphenicol or con¬ being
of the
On examination after the
trol groups. The results of the labora¬ tory analysis for chloramphenicol are shown in the Figure. Depicted are the actual values of chloramphenicol for each sample analyzed along with the calculated least-squares line. The quantity of aqueous humor supplied to the laboratory was insufficient for analysis in two cases, and no chloram¬ phenicol was identified in four blind control samples. The active compound (intact chlo¬
ramphenicol) migrated on all TLC plates identically to the applied stan¬ dard, whereas control (blank) samples showed neither ultraviolet absorbing components in the vicinity of the
divided topical dose, maximum aqueous humor levels of chloram¬ phenicol (3ßg/ml to 6^g/ml) would be similar to those obtained after a single 1.0-gm oral dose. Our results also indicate that concentrations of 3µg/ml would be present in the aqueous humor for approximately two hours after the 3.0-mg topical dose. From the standpoint of sys¬ temic toxicity associated with oral ad¬ ministration, the advantage of topi¬ cal administration of chloramphenicol is obvious. Since the compound extracted from the aqeuous humor migrated on the TLC plates identically to the applied standard and no metabolites of chlor¬ amphenicol were identified, we have concluded that the major active com¬ pound measured in this study was in¬ tact chloramphenicol. However, small quantities of metabolites that were below the sensitivity limits of detec¬ tion may have been present and can¬ not be ruled out as a possibility.
Busby, Ms. Bette McCue, and Dr. R. supplied technical assistance for the laboratory analysis of chloramphenicol. Ms. E. Adamski
Informed consent
was
obtained from the sub¬
jects. Chloramphenicol 0.5% (Econochlor) was sup¬ plied by Alcon Laboratories, Inc., Fort Worth, Tex.
References Hours
Topical Administration
analysis for chloramphenicol in human aqueous humor after topical application (individual samples and least-squares line). Results of TLC
standard
nor
chemical reactions that
identify chloramphenicol. Evidence of enzymatic hydrolysis of the dichloroacetyl group to produce the chlor¬ amphenicol metabolite of D-threo-1-pnitrophenyl-2-amino-l, 3-propanediol was also absent on all plates. Comment
Following oral administration of chloramphenicol in humans, blood lev¬ els have been found to be roughly pro¬ portional to the size of the dose,
reaching a maximum in two to four hours. For example, peak serum levels averaged 4u.g/ml after a single 0.5gm dose, 9µg after 1.0 gm and l^g after 2.0 gm.3 Abraham and Burnett have reported that from 33% to 79% (mean, 52%) of the blood concentra¬ tion of chloramphenicol passes into the primary aqueous humor after each oral administration.6 Correlation of these blood-aqueous levels with the results of the present experiment would indicate that, utilizing a 3.0-mg
1. Leopold IH, Nichols AC, Vogel AW: Penetration of chloramphenicol U.S.P. (Chloromycetin) into the eye. Arch Ophthalmol 44:22-36,1950. 2. Sorsby A, Ungar J, Crick RP: Aureomycin, chloramphenicol, and Terramycin in ophthalmology. Br Med J 2:301-304, 1953. 3. Bleeker GM, Maas ER: The penetration of Aureomycin, Terramycin and chloramphenicol in the ocular tissues. Ophthalmologica 130:1-8,1955. 4. Langham M: Factors affecting the penetration of antibiotics into the aqueous humor. Br J Ophthalmol 35:614-620, 1951. 5. Leopold IH: Clinical trial with chloramphenicol in ocular infections. Arch Ophthalmol 45:44-52, 1951. 6. Abraham RK, Burnett HH: Tetracycline and chloramphenicol studies on rabbit and human eyes. Arch Ophthalmol 54:641-659, 1955. 7. Rosenthal RL, Blackmon A: Bone marrow hypoplasia following use of chloramphenicol eye drops. JAMA 191:136-137, 1965. 8. Dawson RMC, Elliott DC, Elliott WH, et al (eds): Data for Biochemical Research, ed 2. Oxford University Press, 1969, p 538. 9. Ley HL Jr, Smadel JE, Crocker TT: Administration of Chloromycetin to normal human subjects. Proc Soc Exp Biol Med 68:9-12, 1948.
Downloaded From: http://archopht.jamanetwork.com/ by a Michigan State University User on 06/18/2015
