Abstract
We report problems encountered during preparation of tritium-labeled unconjugated bilirubin ( 3H-UCB) from precursor 3H-5-aminolevulinic acid ( 3H-ALA) in 2 dogs with external biliary drainage installed into the animals under general anesthesia. Under prolonged sedation, 12.9 or 14.0 mCl of 3H-ALA was administered intravenously in two divided doses, and bile was collected for 9 hours. In one animal, taurocholate (TC) infusion was needed to maintain bile flow. 3H-UCB was isolated from the bile and recrystallized with the improved method of Webster et al (Webster CC, Tiribelli C, Ostrow JD. J Lab Clin Med 2001;137:370-3). Based on radioactivity and pigment content, hourly bile collections were pooled to optimize specific activities. Surprisingly, in the first dog, only 2.9% of injected radioactivity was recovered in bile and only 14.1% in urine, and the specific activities of the crystalline 3H-UCB from the two pools were only 39.5 and 30.0 × 10 3 dpm/μg. High-performance liquid chromatography analysis revealed that only 4% of ALA degraded during 5 minutes in injection solution at pH 6.8. The low incorporation of 3H-ALA and low specific activity of 3H-UCB was apparently caused mainly by prior degradation and exchange of labile tritium of the 3H-ALA and probably by enhanced endogenous ALA synthesis caused by the anesthetic/sedative agents. Revised procedures in the second dog improved the incorporation of 3H-ALA to 11.9% excreted in bile and the specific activity of the crystalline 3H-UCB to 122.0 and 50.8 × 10 3 dpm/μg, while urinary excretion of tritium increased to 28.5%. These experiences emphasize possible pitfalls in preparing 3H-UCB by biosynthetic labeling from 3H-ALA administered to dogs.
| Original language | English |
|---|---|
| Pages (from-to) | 313-321 |
| Number of pages | 9 |
| Journal | The Journal of Laboratory and Clinical Medicine |
| Volume | 138 |
| Issue number | 5 |
| DOIs | |
| Publication status | Published - 2001 |
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ASJC Scopus subject areas
- Medicine(all)
- Pathology and Forensic Medicine
Cite this
Pitfalls in preparation of 3H-unconjugated bilirubin by biosynthetic labeling from precursor 3H-5-aminolevulinic acid in the dog. / Bayón, J. Enriqué; Pascolo, Lorella; Gonzalo-Orden, José M.; Altonaga, Jose R.; González-Gallego, Javier; Webster, Cecile; Haigh, W. Geoffrey; Stelzner, Matthias; Pekow, Cyndy; Tiribelli, Claudio; Ostrow, J. Donald.
In: The Journal of Laboratory and Clinical Medicine, Vol. 138, No. 5, 2001, p. 313-321.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Pitfalls in preparation of 3H-unconjugated bilirubin by biosynthetic labeling from precursor 3H-5-aminolevulinic acid in the dog
AU - Bayón, J. Enriqué
AU - Pascolo, Lorella
AU - Gonzalo-Orden, José M.
AU - Altonaga, Jose R.
AU - González-Gallego, Javier
AU - Webster, Cecile
AU - Haigh, W. Geoffrey
AU - Stelzner, Matthias
AU - Pekow, Cyndy
AU - Tiribelli, Claudio
AU - Ostrow, J. Donald
PY - 2001
Y1 - 2001
N2 - We report problems encountered during preparation of tritium-labeled unconjugated bilirubin ( 3H-UCB) from precursor 3H-5-aminolevulinic acid ( 3H-ALA) in 2 dogs with external biliary drainage installed into the animals under general anesthesia. Under prolonged sedation, 12.9 or 14.0 mCl of 3H-ALA was administered intravenously in two divided doses, and bile was collected for 9 hours. In one animal, taurocholate (TC) infusion was needed to maintain bile flow. 3H-UCB was isolated from the bile and recrystallized with the improved method of Webster et al (Webster CC, Tiribelli C, Ostrow JD. J Lab Clin Med 2001;137:370-3). Based on radioactivity and pigment content, hourly bile collections were pooled to optimize specific activities. Surprisingly, in the first dog, only 2.9% of injected radioactivity was recovered in bile and only 14.1% in urine, and the specific activities of the crystalline 3H-UCB from the two pools were only 39.5 and 30.0 × 10 3 dpm/μg. High-performance liquid chromatography analysis revealed that only 4% of ALA degraded during 5 minutes in injection solution at pH 6.8. The low incorporation of 3H-ALA and low specific activity of 3H-UCB was apparently caused mainly by prior degradation and exchange of labile tritium of the 3H-ALA and probably by enhanced endogenous ALA synthesis caused by the anesthetic/sedative agents. Revised procedures in the second dog improved the incorporation of 3H-ALA to 11.9% excreted in bile and the specific activity of the crystalline 3H-UCB to 122.0 and 50.8 × 10 3 dpm/μg, while urinary excretion of tritium increased to 28.5%. These experiences emphasize possible pitfalls in preparing 3H-UCB by biosynthetic labeling from 3H-ALA administered to dogs.
AB - We report problems encountered during preparation of tritium-labeled unconjugated bilirubin ( 3H-UCB) from precursor 3H-5-aminolevulinic acid ( 3H-ALA) in 2 dogs with external biliary drainage installed into the animals under general anesthesia. Under prolonged sedation, 12.9 or 14.0 mCl of 3H-ALA was administered intravenously in two divided doses, and bile was collected for 9 hours. In one animal, taurocholate (TC) infusion was needed to maintain bile flow. 3H-UCB was isolated from the bile and recrystallized with the improved method of Webster et al (Webster CC, Tiribelli C, Ostrow JD. J Lab Clin Med 2001;137:370-3). Based on radioactivity and pigment content, hourly bile collections were pooled to optimize specific activities. Surprisingly, in the first dog, only 2.9% of injected radioactivity was recovered in bile and only 14.1% in urine, and the specific activities of the crystalline 3H-UCB from the two pools were only 39.5 and 30.0 × 10 3 dpm/μg. High-performance liquid chromatography analysis revealed that only 4% of ALA degraded during 5 minutes in injection solution at pH 6.8. The low incorporation of 3H-ALA and low specific activity of 3H-UCB was apparently caused mainly by prior degradation and exchange of labile tritium of the 3H-ALA and probably by enhanced endogenous ALA synthesis caused by the anesthetic/sedative agents. Revised procedures in the second dog improved the incorporation of 3H-ALA to 11.9% excreted in bile and the specific activity of the crystalline 3H-UCB to 122.0 and 50.8 × 10 3 dpm/μg, while urinary excretion of tritium increased to 28.5%. These experiences emphasize possible pitfalls in preparing 3H-UCB by biosynthetic labeling from 3H-ALA administered to dogs.
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U2 - 10.1067/mlc.2001.118746
DO - 10.1067/mlc.2001.118746
M3 - Article
VL - 138
SP - 313
EP - 321
JO - Journal of Laboratory and Clinical Medicine
JF - Journal of Laboratory and Clinical Medicine
SN - 0022-2143
IS - 5
ER -