Development of a novel intestinal and vascular access port (IVAP) rabbit model to study regiospecific oral absorption pharmacokinetics.

BACKGROUND AND PURPOSE: The limited availability and cost of many drugs prohibits routine use of the previously developed intestinal and vascular access port (IVAP) canine model by our group. A lower animal species model such as the rabbit is suitable for implanting intestinal and vascular access ports for investigating regiospecific intestinal absorption and hepatic elimination while requiring significantly lower doses of drugs. In addition, expression of certain cytochrome P450 enzymes and apical secretory and absorptive transporters in rabbit intestine is similar to that in humans making the rabbit a suitable model. METHODS: Individual 5-F Silastic catheters were placed in the proximal or distal portion of the small intestine or the colon of subject animals, while a 5-F Heparin Coated Polyurethane (HCP) catheter was implanted in the portal vein of each subject. The catheters were tunneled out of the abdomen and attached to separate subcutaneous access ports along the spine. The animals were allowed a two-week minimum recovery period prior to being used in pharmacokinetic studies. RESULTS AND DISCUSSION: After some initial difficulties, rabbits with IVAP implants proved to be an efficient and dependable model for investigating intestinal and hepatic extraction of drugs. Fluoroscopic visualization of intestinal and portal venous catheters indicated that surgically implanted catheters did not interfere with gastrointestinal motility or blood flow into the liver, respectively. Acute pH studies in the proximal portion of the small intestine were consistent with normal GI motility patterns.

Differentiation of gut and hepatic first-pass effect of drugs: 1. Studies of verapamil in ported dogs.

PURPOSE: To investigate the relative contributions of the gut and liver to the first-pass loss of verapamil (VL) using an in vivo intestinal-vascular access port (IVAP) dog model. METHODS: Basic pharmacokinetics of VL were determined after intravenous (IV: 0.5 mg/kg), portal venous (PV: 2 mg/kg), and duodenal (ID: 2 mg/kg) administration in IVAP dogs. Serial blood samples were collected for 8 h after dosing, and plasma was analyzed for unchanged drug by a high-performance liquid chromatography-fluorescence method. Extraction ratios in the liver and intestinal tract were determined from the area under the concentration-time curves for ID, PV, and IV administration. The functional role of CYP450 or secretory transporters such as P-gp on the gut and liver first-pass loss of VL was further studied using ritonavir, a known substrate or inhibitor of these processes. RESULTS: The liver had a high intrinsic capacity for clearing VL because the absolute bioavailability (BA) of VL was 21.7% after PV administration. The BA of VL after ID administration was 23.5%; therefore, intestinal absorption was complete and intestinal extraction was negligible (ER(GI) approximately 0). The BA of VL increased from 23.5% to 66.2% in the presence of ritonavir primarily due to a reduction in hepatic extraction. CONCLUSIONS: Although the liver had a high intrinsic capacity for extracting VL, the contribution of gut to the first-pass loss of VL was negligible. Because of the additive effects of intestinal CYP3A-mediated metabolism and secretory transport, a significant gut first-pass effect was expected, but not observed in dogs. These studies demonstrate the utility of the in vivo IVAP dog model for evaluating the relative contribution of the gut and liver to the first-pass loss of drugs and for characterizing the functional role that CYP450 metabolism and/or secretory transporters play in drug-drug interactions and reduced oral bioavailability.

Regional oral absorption, hepatic first-pass effect, and non-linear disposition of salmon calcitonin in beagle dogs.

The dose-dependent disposition, first pass hepatic elimination, and absorption pharmacokinetics (PK) of salmon calcitonin (sCT) were investigated in a canine Intestinal Vascular Access Port (IVAP) model. The PK of sCT were determined after intravenous (IV), subcutaneous (SC), portal venous (PV), and oral (PO) administration of sCT. Regional oral absorption of unformulated sCT was also evaluated by direct administration into the duodenum (ID), ileum (IL), and colon (IC) by means of surgically implanted, chronic catheters. Plasma samples were collected and analyzed by radioimmunoassay (RIA). Salmon calcitonin PK were evaluated using 2-compartmental and model independent methods. Intravenous sCT PK were non-linear over the dose range studied. High dose groups (100-1000 microg) demonstrated higher total plasma clearance (CL) and V(dss) than the low dose groups (1-25 microg). However, the MRT did not change for doses ranging from 10 to 1000 microg. After SC administration, the absorption of sCT was rapid with bioavailability (BA) varying from 21.4 to 52.9%. However, the BA of sCT was low after ID, IL, and IC administration (0.039, 0.064, and 0.021%, respectively). The role of hepatic first-pass elimination was negligible. The results of these studies demonstrate that the elimination of sCT is rapid but does not occur in the liver. Enhanced sCT clearance at higher doses was indicated by increasing V(dss) values, and it is hypothesized that increased renal blood flow and/or saturated plasma protein binding may contribute to the non-linear behavior. The IVAP canine model was found to have utility for probing the absorption and disposition PK of sCT. The combination of high oral bioavailability variability and non-linear disposition of sCT may produce highly variable therapeutic effects. The practical impact of the non-linear disposition of sCT remains to be determined. Based on the current results it appears that the rate-limiting step to the successful oral administration of sCT is its delivery into the portal vein since hepatic metabolism was negligible.

Improved longevity and functionality of a canine model providing portal vein and multi-site intestinal access.

BACKGROUND AND PURPOSE: The canine intestinal and venous access port (IVAP) model is valuable for investigating hepatic elimination and region-specific intestinal absorption of pharmaceuticals. Previously, long-term functionality of this preparation has been variable. METHODS: Catheters of different construction were placed in the proximal and distal portions of the small intestine, colon, and portal vein of subject animals and were attached to separate subcutaneous access ports. Intraoperative, postoperative, and long-term maintenance techniques were developed, modified, and analyzed. RESULTS: Intestinal catheter infections and access site failures were associated with breakdown at the intestinal insertion site. The ileal catheter was prone to obstruction with ingesta. A modified Witzel technique, specialized port-catheter systems, scheduled port-flushing methods, and venous port infection treatment protocols improved the model's longevity. CONCLUSIONS: The canine IVAP model is a powerful tool for investigation of regional differences in intestinal absorption and hepatic elimination of drugs. Other researchers can derive increased longevity with the IVAP model by using the technical modifications detailed here: strict sterile technique, closed-end slit-valve catheters, GPV ports, the Witzel tunnel technique, routine portal vein infection surveillance, 50% dextrose intestinal catheter infusion, rapid removal of infected intestinal catheters, and critical appraisal of their results. Longevity of the model continues to be improved.

Regional differences in intestinal spreading and pH recovery and the impact on salmon calcitonin absorption in dogs.

PURPOSE: To investigate the regional influence of intestinal spreading and pH recovery on the performance of drug and excipient delivery systems and their impact on the oral absorption of a model peptide drug, salmon calcitonin (sCT), in conscious beagle dogs. METHODS: Male beagle dogs were surgically prepared with subdermal Intestinal Access Ports (IAP). The catheter from one port was placed in the duodenum and the other in the ileum. Fluoroscopy and Heidelberg pH capsule studies were performed to characterize intestinal spreading and pH recovery, respectively. Three treatments were performed: (1) a radiopaque dye and citric acid (CA) were infused into the intestinal segments, (2) a radiopaque powder capsule containing CA was given orally, and (3) capsules containing CA and sCT were given orally. Regular blood samples were collected and analyzed by radioimmunoassay (RIA) to determine the absorption characteristics of sCT. RESULTS: Since sCT is an excellent substrate for the pancreatic serine protease trypsin, the rate of degradation of sCT in the GI lumen is dependent upon the regional pH, activity of digestive enzymes and the concentration of sCT at the site of absorption. Fluoroscopy results clearly showed that when the radiopaque dye was infused into the duodenum and capsule disintegration occurred early, there was significant dilution and spreading of the excipients throughout a large section of the upper small intestine (USI). However, when the radiopaque dye was infused into the ileum and capsule disintegration occurred in the lower small intestine (LSI), the excipients moved along as a bolus (i.e., plug). The pH monitoring results were consistent with the fluoroscopy results. The pH dropped only momentarily and rose quickly in the USI consistent with well-stirred mixing kinetics. In the LSI, dilution and spreading were minimal and the drop in pH was greater and persisted for a longer period of time. Plasma levels of sCT were maximal when disintegration occurred in the LSI. CONCLUSIONS: Since significantly less dilution and spreading occurred in the LSI, the exposure of the intestine to pharmaceutical excipients and sCT was more concentrated resulting in a higher fraction of sCT absorbed. The results of this study demonstrate that intestinal mixing kinetics have a dramatic impact on the ability of pharmaceutical excipients to modulate the oral bioavailability of peptide drugs like sCT.

Biopharmaceutical approaches for developing and assessing oral peptide delivery strategies and systems: in vitro permeability and in vivo oral absorption of salmon calcitonin (sCT).

PURPOSE: To evaluate a biopharmaceutical approach for selecting formulation additives and establishing the performance specifications of an oral peptide delivery system using sCT as a model peptide. METHODS: The effect of formulation additives on sCT effective permeability and transepithelial electrical resistance (TEER) was evaluated in side-by-side diffusion chambers using rat intestinal segments. Baseline regional oral absorption of sCT was evaluated in an Intestinal and Vascular Access Port (IVAP) dog model by administration directly into the duodenum, ileum, and colon by means of surgically implanted, chronic catheters. The effect of varying the input rate and volume of the administered solution on the extent of sCT absorption was also evaluated. Citric acid (CA) was utilized in all studies to cause a transient reduction in local pH. In vitro samples and plasma samples were analyzed by radioimmunoassay (RIA). Two oral delivery systems were prepared based on the results of the in vitro and IVAP studies, and evaluated in normal dogs. RESULTS: Maximal permeability enhancement of sCT was observed using taurodeoxycholate (TDC) or lauroyl carnitine (LC) in vitro. Ileal absorption of sCT was higher than in other regions of the intestine. Low volume and bolus input of solution formulations was selected as the optimal condition for the IVAP studies since larger volumes or slower input rates resulted in significantly lower sCT bioavailability (BA). Much lower BA of sCT was observed when CA was not used in the formulation. The absolute oral bioavailability (mean+/-SD) in dogs for the control (sCT + CA) and two proprietary sCT delivery systems was 0.30%+/-0.05%, 1.10+/-0.18%, and 1.31+/-0.56%, respectively. CONCLUSIONS: These studies demonstrate the utility of in vitro evaluation and controlled in vivo studies for developing oral peptide delivery strategies. Formulation additives were selected, the optimal intestinal region for delivery identified, and the optimal release kinetics of additives and actives from the delivery system were characterized. These methods were successfully used for devising delivery strategies and fabricating and evaluating oral sCT delivery systems in animals. Based on these studies, sCT delivery systems have been fabricated and tested in humans with favorable results.

Unstructured cases in case-based learning benefit students with primary care career preferences.

BACKGROUND: The impact of instructional method on students with opposing surgical career orientations was investigated in a prospective study. METHODS: Students were randomly assigned to structured or unstructured case-based discussions. Clinical reasoning (OSCE and a diagnosis exercise), subject knowledge (multiple choice test [MCT]), method preference, and pre-third year career preference were compared. RESULTS: Twenty-two students listed a surgical career high (Surgical) and 20 low (Primary). Surgical MCT scores were higher than Primary regardless of instructional method. Surgical diagnosis exercise scores were higher than Primary with the structured method (22.0+/-2.3 versus 15.1+/-3.0, P <0.08). Unstructured scores on this exercise were similar (19.7+/-1.8 Surgical versus 20.3+/-3.5 Primary). Analysis of variance suggested an interaction on the diagnosis exercise between method and career (P = 0.16). Students preferred the unstructured method. CONCLUSIONS: The improved diagnosis exercise performance implies that unstructured cases positively influence surgical domain specific reasoning for nonsurgical career students. These method effects increase our understanding of case-based methods in surgical education.

Oral absorption of anti-aids nucleoside analogues: 3. Regional absorption and in vivo permeability of 2',3'-dideoxyinosine in an intestinal-vascular access port (IVAP) dog model.

The absolute oral and regional intestinal bioavailabilities (BAs) and pharmacokinetics (PK) of 2',3'-dideoxyinosine (ddI), a nucleoside analog used in the treatment of human immunodeficiency virus (HIV) infection, were investigated in an in vivo intestinal-vascular access port (IVAP) dog model. The mean (+/- SD) absolute regional intestinal BAs of ddI were 49.6 +/- 8.8, 42.7 +/- 7.9, and 13.6 +/- 5.4% after the bolus administration of unbuffered solutions containing 250 mg ddI into the duodenum, ileum, and colon of IVAP beagle dogs, respectively. The BA of the orally administered Videx 250 mg buffered chewable tablets was 44.9 +/- 1.6%. ddI absorption and disposition PK were modeled by simultaneously fitting intravenous, oral, and intestinal plasma level versus time data using a physiologically based PK model. The region-specific apparent absorption rates followed the rank order duodenum > ileum > colon. Apparent regional in vivo intestinal permeabilities correlated well with previously determined regional permeabilities in rats. The intestinal pH was monitored using a radiotelemetric pH monitoring system since ddI is unstable in an acidic environment. While the pH was found to be lower in the duodenum and proximal jejunum (approximately pH 6) than in the ileum or colon (pH > or = 7.0), ddI is reasonably stable across the entire pH range of the dog small intestine. These studies demonstrate that the regional reduction in ddI BA is consistent with a reported distal reduction in intestinal permeability and appears to be a significant contributing factor to the high degree of absorption variability reported for ddI.

Air bags alone compared with the combination of mechanical restraints and air bags: implications for the emergency evaluation of crash victims.

BACKGROUND: Air bags (ABs) may be perceived by the public and physicians as protection for thoracoabdominal injuries. This study compares injury patterns when air bags are used alone with injury patterns when air bags plus mechanical restraints (MRs) are used. METHODS: Patients treated over a 4-year period with emergency medical services-documented AB deployment alone (n = 16) or AB plus MR (n = 22) were identified by trauma registry query. Medical records were reviewed and injuries recorded. RESULTS: Air bag-alone users had more severe overall (injury severity score > or = 15:9 vs 5), chest (abbreviated injury score [AIS] > or = 3:5 vs 1), and abdominal injuries (AIS > or = 3:6 vs 0). They required more tube thoracostomies (5 vs 0) and laparotomies (6 vs 0), longer hospitalizations (11.9 +/- 3.2 vs 5.3 +/- 1.4 days), and more intensive care unit admissions (8 vs 1). Craniofacial injuries (AIS > or = 3:6 vs 6) and fractures were similar. More victims using air bags alone required impatient rehabilitation and some patients died (6 vs 1). CONCLUSIONS: Crash victims using air bags alone (vs AB plus MR) had increased injury severity, hospitalizations, thoracoabdominal procedures, and rehabilitation. Physicians must be aware of the incomplete protection by air bags alone.

Cognitive trauma care is undervalued: adult splenic injury as a paradigm.

Nonoperative management (NOM) of adult splenic injury is evolving. Economic aspects of NOM have not been examined. We hypothesize that NOM reduces hospital and professional charges. Surgeon, radiologist, and hospital charges and reimbursements, and clinical outcome were obtained for 77 consecutive adult splenic injury patients (> or = 15 years old) over a 3-year period. NOM succeeded in 30 of 31 patients. NOM was associated with lower surgeon fee ($1,148 vs $4,452; P < 0.0001), surgeon reimbursement ($587 vs $2,773; P = 0.0001), and hospital charge ($18,982 vs $48,790; P = 0.001) relative to operative management. Radiologist fee ($1,776 vs $2,285) and reimbursement ($1,069 vs $1,537) were not significantly affected. No significant difference existed between surgeon (primary care provider) and radiologist reimbursement for NOM. ISS poorly correlated with economic variables. We conclude that cost reductions are another potential advantage of NOM. Surgeon reimbursement for the cognitive skills involved in NOM is minimal. Future health finance policy should recognize the cognitive aspects of trauma care.

Role of case structure and prior experience in a case-based surgical clerkship.

BACKGROUND: As case-based methods replaced lectures in a surgical clerkship, the influences of case structure and prior experience on learning were investigated. METHODS: Early and late third-year students randomly received different cases. "Structured" cases had data presented and summarized. "Unstructured" cases required questions to faculty for information. Multiple choice tests and differential diagnosis activities were administered. An attitudinal questionnaire gauged student perceptions. RESULTS: In both multiple choice and differential diagnosis activities, the late rotation, "unstructured" group scored higher than the "structured" group. Conversely, the early rotation, "unstructured" group scored lower than the "structured" group. Combined, rotation, and structure significantly affected both multiple choice and differential diagnosis activities (ANOVA, P < or = 0.02). Early rotation, "unstructured" students described a more enjoyable experience, despite lower evaluation scores. CONCLUSIONS: Surgical clerkship case-based learning is profoundly affected by case structure and prior clinical experience. Case-based curriculum should be tailored to accommodate these interactions.

Bacterial translocation after mesenteric ligation in dogs.

These experiments were designed to determine the relationship between translocation of Escherichia coli and viability of ischemic small bowel. Twenty beagles were gavaged with 14C-labeled E. coli at two time intervals (3 and 24 h) prior to ligation of the blood supply to a 40-cm segment of ileum. Mesenteric lymph node (MLN) biopsies and bacterial cultures of the peritoneal fluid, peripheral arterial blood, and splanchnic venous blood were taken immediately prior to ligation and 24 h later both before and after the ischemic bowel was resected and anastomosed. Biopsies of each resection margin were taken to measure translocation of E. coli into the bowel wall. Several hemodynamic hemodynamic parameters were also measured before and 24 h after ligation. Seven of the 20 dogs died of further bowel necrosis. In survivors A-alpha DO2 was significantly decreased 24 h after mesenteric ligation vs. preligation, whereas in dogs that died DO2 was significantly increased after ligation vs. preligation. The incidence of mesenteric venous cultures positive for E. coli was significantly higher 24 h after ligation vs. preligation. However, there was no correlation between survival and the incidence of positive E. coli cultures in the blood or peritoneal fluid. Mean MLN counts were significantly higher in dogs gavaged at 3 h vs. those gavaged 24 h prior to laparotomy. However, there was no correlation between survival and translocation into either the bowel wall or MLN at either time interval. Viability of ischemic small bowel in this canine model was not affected by translocation of E. coli. Hemodynamic parameters that are altered during the course of sepsis also did not correlate with survival.

Antibiotic patterns associated with fungal colonization in critically ill surgical patients.

Fungal infections (FI) in surgical patients are increasing; mortality approaches 50 per cent. Prior studies identified factors predicting fungal colonization (FC) including broad spectrum antibiotics (BSA). This study investigates antibiotic patterns predicting FC. Other risk factors and outcomes are analyzed. A total of 72 consecutive SICU patients receiving > or = 7 days BSA were followed. None received prophylactic antifungals. Input data: Age, APACHE II, surgical procedure, lines, ulcer prophylaxis, TPN duration, antibiotic/antifungal regimen. Outcome data: FC, FI, length of SICU and hospital stay, mortality. A total of 32 patients (44%) developed FC; five (16%) developed FI (P = 0.08). All infected patients died (P = 0.0002). FC of GU (25%), respiratory (19%), and GI (19%) tracts were common. Multiple site colonization occurred in 25 per cent of colonized patients. Metronidazole and duration of ventilation predicted FC. APACHE II and TPN duration predicted mortality. Mortality occurred exclusively among patients requiring systemic antifungals. Among BSA, only metronidazole independently predisposed to FC. Other predictors of colonization and mortality agree with prior studies. The high mortality among patients requiring systemic antifungals implies that a more aggressive approach to prophylaxis may be warranted.

"Adult" trauma surgeons with pediatric commitment: a logical solution to the pediatric trauma manpower problem.

Pediatric trauma care by "adult" surgeons is debated, despite the paucity of pediatric trauma surgeons; 424 patients < or = 17 admitted to a Level I Center run by "adult" surgeons were analyzed. Demographics mirrored NPTR (mean ISS 11.5; mean age 10). "Adult" critical care surgeons treated MTOS-comparable patients with outcomes comparable to MTOS. Among other specialists, only neurosurgeons saw a MTOS-comparable population. Nonoperative protocols for solid organ injury were used appropriately. Z for all patients was +0.17 with M 0.908. Ps was 0.951; acute survival was 0.958 with 18 deaths (mean Ps 0.158). There were two unexpected survivors and one unexpected death; 73% of survivors had age-appropriate locomotion. Pediatric trauma outcomes by "adult" surgeons compare favorably to national standards. The recommendation that pediatric trauma care be directed by pediatric surgeons should be qualified in view of such outcomes and the paucity of pediatric trauma surgeons.

Computed tomography is inaccurate in estimating the severity of adult splenic injury.

Computed tomography (CT) is increasingly utilized in evaluation of adult splenic injury (SI). CT correlation with operative findings, CT relationship to successful nonoperative (NO) management, and CT reading reproducibility were examined. Records of patients > or = 15 years old admitted over a 3-year period were reviewed. Computed tomography scans were graded by two radiologists blinded to clinical results. Computed tomography scans were performed on 49 of 77 patients with SI. Eighteen underwent initial operation (OR) and 31 initial NO. Operative patients had higher Injury Severity Scores and Abdominal Abbreviated Injury Scale scores (p < 0.0001). Grade II readings predominated in the NO group (55%). Nonoperative management was successful for 9 grade III and 3 grade IV readings. Computed tomography matched OR grade in 10 readings, underestimated it in 18, and overestimated it in 6. Computed tomography missed SI in five patients. Radiologists disagreed on 9 of 45 (20%) scans. Computed tomography poorly predicted operative findings. Interobserver variability was common. SI management should not be based solely on CT severity.

Tracheostomy and percutaneous endoscopic gastrostomy in the management of the head-injured trauma patient.

Forty-three trauma patients underwent tracheostomy (TRACH) and percutaneous endoscopic gastrostomy (PEG) over 21 months. Thirty-one patients had a head injury with Abbreviated Injury Scale > or = 3 associated with multi-trauma. This study was undertaken to analyze demographic and outcome variables with respect to timing of TRACH/PEG in this population. Patients were divided into EARLY (< or = 7 days) and LATE (> 7 days) groups and were analyzed for admission Glasgow Coma Scale, Apache II, Injury Severity Score, and [(A-a)DO2] at time of TRACH/PEG. Outcome variables were ICU length of stay (LOS), hospital LOS, days of mechanical ventilation (MV) post-TRACH/PEG, complications, and mortality. Esophagogastroduodenoscopy findings with PEG and days to full enteral nutrition were recorded. All demographic variables were statistically similar between the EARLY and LATE groups. The EARLY group had shorter hospital LOS (P < 0.05), total Intensive Care Unit LOS (P < 0.05), ICU LOS post-TRACH/PEG (P < 0.05), and fewer days of MV post-TRACH/PEG (P < 0.05). There were no procedure-related complications of TRACH/PEG in either group. Full Esophagogastroduodenoscopy performed at the time of PEG had a high diagnostic yield in both groups. We conclude that TRACH/PEG performed within the first 7 days of injury in the head trauma patient is the procedure of choice for long-term airway protection, mechanical ventilation, and enteral nutrition. Combined use of these procedures reduces ICU and hospital LOS and shortens the course of MV.

The identification and formation of 20-aldehyde leukotriene B4.

Microsomes of human polymorphonuclear leukocytes (PMN) in the presence of 100 microM NADPH converted 0.6 microM leukotriene B4 (LTB4) to 20-OH-LTB4 (retention time = 18.0 min) and to two additional compounds designated I (retention time = 16.8 min) and II (retention time = 9.6 min) as analyzed by reverse-phase high performance liquid chromatography (HPLC). Compounds I and II were also formed from the reaction of 1.0 microM 20-OH-LTB4, PMN microsomes, and 100 microM NADPH; the identity of compound II was confirmed as 20-COOH-LTB4 by gas chromatography-mass spectrometry. Equine alcohol dehydrogenase in the presence of 100 microM NAD+ in 0.2 M glycine buffer (pH 10.0) converted 20-OH-LTB4 to 20-aldehyde (CHO) LTB4, which coeluted with compound I on reverse-phase HPLC. In the presence of 100 microM NADH in 50 mM potassium phosphate buffer (pH 6.5), equine alcohol dehydrogenase reduced both 20-CHO-LTB4 and compound I to 20-OH-LTB4, indicating the identity of compound I as 20-CHO-LTB4. Gas chromatography-mass spectrometry of trideuterated O-methyl-oxime trimethylsilyl ether methyl ester derivative of 3H-labeled compound I definitively established compound I as 20-CHO-LTB4. Addition of immune IgG to cytochrome P-450 reductase or 1.0 mM SKF-525A completely inhibited the formation of 20-CHO-LTB4 from 20-OH-LTB4, indicating that the reaction was catalyzed by a cytochrome P-450. LTB5 (3.0 microM), a known substrate for cytochrome P-450LTB and a competitive inhibitor of LTB4 omega-oxidation, completely inhibited the omega-oxidation of 1.5 microM 20-OH-LTB4 to 20-CHO-LTB4, indicating that the cytochrome P-450 was P-450LTB. Conversion of 1.0 microM 20-CHO-LTB4 to 20-COOH-LTB4 by PMN microsomes was also dependent on NADPH and inhibited by antibody to cytochrome P-450 reductase, 1.0 mM SKF-525A, or 5.0 microM LTB5, indicating that this reaction was also catalyzed by cytochrome P-450LTB. These results identify the novel metabolite 20-CHO-LTB4 and indicate that cytochrome P-450LTB catalyzes three sequential omega-oxidations of LTB4 leading to the formation of 20-COOH-LTB4 via 20-OH-LTB4 and 20-CHO-LTB4 intermediates.