scholarly journals A fluorescent residualizing label for studies on protein uptake and catabolism in vivo and in vitro

1990 ◽  
Vol 267 (1) ◽  
pp. 155-162 ◽  
Author(s):  
J L Maxwell ◽  
L Terracio ◽  
T K Borg ◽  
J W Baynes ◽  
S R Thorpe
Keyword(s):  
Half Life ◽  
Normal Tissue ◽  
Degradation Products ◽  
Rat Serum ◽  
Peripheral Tissues ◽  
Protein Uptake ◽  
Kinetics Of ◽  
Rat Serum Albumin

Residualizing labels are tracers which remain in lysosomes after uptake and catabolism of the carrier protein and have been especially useful for studies on the sites of plasma protein degradation. Thus far these labels have contained radioactive reporters such as 3H or 125I. In the present paper we describe a fluorescent residualizing label, NN-dilactitol-N′-fluoresceinylethylenediamine (DLF). Modification of asialofetuin (ASF) or rat serum albumin (RSA) with DLF affected neither their normal kinetics of clearance from the rat circulation nor their normal tissue sites of uptake and degradation. After injection of DLF-ASF, fluorescent degradation products were recovered nearly quantitatively in liver and retained with a half-life of about 2 days. Fluorescent degradation products from DLF-RSA were recovered in skin and muscle, and were localized in fibroblasts by fluorescence microscopy. These results confirm previous studies with radioactive residualizing labels in which fibroblasts in peripheral tissues were identified as primary sites of albumin degradation. Fluorescent catabolites also accumulated in fibroblasts incubated with DLF-RSA in vitro, and residualized with a half-life of about 2 days. Overall, the data establish that DLF functions efficiently as a fluorescent residualizing label both in vivo and in vitro. The advantages of fluorescent, compared with radioactive, residualizing labels should make them valuable tools for studies on protein uptake and catabolism in biological systems.

Kinetics of Various 99mTc-Sn-Pyrophosphate Compounds in the Rat

1977 ◽  
Vol 16 (04) ◽  
pp. 157-162 ◽  
Author(s):  
C. Schümichen ◽  
B. Mackenbrock ◽  
G. Hoffmann
Keyword(s):  
Normal Saline ◽  
Half Life ◽  
Compound A ◽  
Short Half Life ◽  
Low Concentrations ◽  
The Stability ◽  
Kinetics Of ◽  
In Vitro Stability

SummaryThe bone-seeking 99mTc-Sn-pyrophosphate compound (compound A) was diluted both in vitro and in vivo and proved to be unstable both in vitro and in vivo. However, stability was much better in vivo than in vitro and thus the in vitro stability of compound A after dilution in various mediums could be followed up by a consecutive evaluation of the in vivo distribution in the rat. After dilution in neutral normal saline compound A is metastable and after a short half-life it is transformed into the other 99mTc-Sn-pyrophosphate compound A is metastable and after a short half-life in bone but in the kidneys. After dilution in normal saline of low pH and in buffering solutions the stability of compound A is increased. In human plasma compound A is relatively stable but not in plasma water. When compound B is formed in a buffering solution, uptake in the kidneys and excretion in urine is lowered and blood concentration increased.It is assumed that the association of protons to compound A will increase its stability at low concentrations while that to compound B will lead to a strong protein bond in plasma. It is concluded that compound A will not be stable in vivo because of a lack of stability in the extravascular space, and that the protein bond in plasma will be a measure of its in vivo stability.

scholarly journals The pharmacokinetics of diminazene aceturate after intramuscular administration in healthy dogs

2005 ◽  
Vol 76 (3) ◽  
Author(s):  
D.M. Miller ◽  
G.E. Swan ◽  
R.G. Lobetti ◽  
L.S. Jacobson
Keyword(s):  
Half Life ◽  
Plasma Concentrations ◽  
Hplc Method ◽  
Terminal Phase ◽  
Diminazene Aceturate ◽  
German Shepherd ◽  
Peripheral Tissues ◽  
Plasma Fraction

The pharmacokinetics of diminazene aceturate following intramuscular (i.m.) administration at 4.2 mg/kg was evaluated in 8 healthy German Shepherd dogs. Blood samples were collected at 19 intervals over a period of 21 days. Diminazene plasma concentrations were measured using a validated HPLC method with UV detection and a sensitivity of 25 ng/m . The in vitro and in vivo binding of diminazene to blood elements was additionally determined. Diminazene pharmacokinetics showed a large inter-individual variation after i.m. administration. It had a short absorption half-life (K01-HL of 0.11 + 0.18 h), resulting in a Cmax of 1849 + 268.7 ng/mℓ at Tmax of 0.37 h and a mean overall elimination half-life (T1/2ß) of 5.31 + 3.89 h. A terminal half-life of 27.5 + 25.0 h was measured. At 1 h after i.m. injection, 75% of the diminazene in whole blood was in the plasma fraction. The results of this study indicate that diminazene is rapidly distributed and sequestered into the liver, followed by a slower terminal phase during which diminazene is both redistributed to the peripheral tissues and/or renally excreted. It is recommended that diminazene administered i.m. at 4.2 mg/kg should not be repeated within a 21-day period.

scholarly journals Comparison of kinetic study of the photochemical changes of (ZZ)-bilirubin IXα bound to human serum albumin with that bound to rat serum albumin

1985 ◽  
Vol 230 (3) ◽  
pp. 561-567 ◽  
Author(s):  
S Onishi ◽  
S Itoh ◽  
T Yamakawa ◽  
K Isobe ◽  
M Manabe ◽  
...  
Keyword(s):  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
Rate Constants ◽  
Relative Rate ◽  
Rat Serum ◽  
Relative Rate Constants ◽  
Rat Serum Albumin

It has been stated by McDonagh, Palma & Lightner [(1982) J. Am. Chem. Soc. 104, 6867-6871] that complexing of bilirubin with serum albumin has a marked species-dependent influence on bilirubin photoisomerization in vitro and in vivo. Therefore the kinetics for the quantitatively important reaction: (Formula: see text) of the photochemical interconversion between bilirubin and its photoisomers bound to human or rat serum albumin in aqueous solution, assayed by h.p.l.c., was used to elucidate the observed species-dependent difference. The relative rate constants for bilirubin bound to human serum albumin, except for k4, the rate of interconversion from (ZZ)-bilirubin into (EZ)-bilirubin, proved to be considerably larger than those for bilirubin bound to rat serum albumin. In accordance with these rate constants, the formation of photoisomers of bilirubin bound to human serum albumin, except for (EZ)-bilirubin, is very rapid and much greater than that for bilirubin bound to rat serum albumin.

scholarly journals In vivo and in vitro degradation of peptide YY3–36 to inactive peptide YY3–34 in humans

2016 ◽  
Vol 310 (9) ◽  
pp. R866-R874 ◽  
Author(s):  
Signe Toräng ◽  
Kirstine Nyvold Bojsen-Møller ◽  
Maria Saur Svane ◽  
Bolette Hartmann ◽  
Mette Marie Rosenkilde ◽  
...  
Keyword(s):  
Half Life ◽  
Peptide Yy ◽  
Degradation Products ◽  
In Vivo Studies ◽  
Metabolic Clearance ◽  
In Vitro Degradation ◽  
Healthy Men ◽  
Amino Acid Peptide

Peptide YY (PYY) is a 36-amino-acid peptide released from enteroendocrine cells upon food intake. The NH2 terminally truncated metabolite, PYY3–36, exerts anorexic effects and has received considerable attention as a possible antiobesity drug target. The kinetics and degradation products of PYY metabolism are not well described. A related peptide, neuropeptide Y, may be degraded from the COOH terminus, and in vivo studies in pigs revealed significant COOH-terminal degradation of PYY. We therefore investigated PYY metabolism in vitro after incubation in human blood and plasma and in vivo after infusion of PYY1–36 and PYY3–36 in eight young, healthy men. A metabolite, corresponding to PYY3–34, was formed after incubation in plasma and blood and during the infusion of PYY. PYY3–34 exhibited no agonistic or antagonistic effects on the Y2 receptor. PYY1–36 infused with and without coadministration of sitagliptin was eliminated with half-lives of 10.1 ± 0.5 and 9.4 ± 0.8 min (means ± SE) and metabolic clearance rates of 15.7 ± 1.5 and 14.1 ± 1.1 ml·kg−1·min−1 after infusion, whereas PYY3–36 was eliminated with a significantly longer half-life of 14.9 ± 1.3 min and a metabolic clearance rate of 9.4 ± 0.6 ml·kg−1·min−1. We conclude that, upon intravenous infusion in healthy men, PYY is inactivated by cleavage of the two COOH-terminal amino acids. In healthy men, PYY3–36 has a longer half-life than PYY1–36.

Clearance of SP-C and recombinant SP-C in vivo and in vitro

1998 ◽  
Vol 274 (6) ◽  
pp. L933-L939 ◽  
Author(s):  
Machiko Ikegami ◽  
Ann D. Horowitz ◽  
Jeffrey A. Whitsett ◽  
Alan H. Jobe
Keyword(s):  
Amino Acid ◽  
Half Life ◽  
Surfactant Protein ◽  
Dipalmitoyl Phosphatidylcholine ◽  
Human Sequence ◽  
Clearance Kinetics ◽  
C Metabolism ◽  
Kinetics Of

Surfactant protein (SP) C metabolism was evaluated in vivo by measurements of the clearance of bovine native SP-C (nSP-C) and a recombinant SP-C (rSP-C) in rabbits and mice and in vitro by the uptake into MLE-12 cells. rSP-C is the 34-amino acid human sequence with phenylalanine instead of cysteine in positions 4 and 5 and isoleucine instead of methionine in position 32. Alveolar clearances of iodinated SP-C and rSP-C after tracheal instillation were similar and slower than those for dipalmitoyl phosphatidylcholine (DPC) in the rabbit. nSP-C and rSP-C were cleared from rabbit lungs similarly to DPC, each with a half-life ( t 1/2) of ∼11 h. In mice, the clearance of rSP-C from the lungs was slower ( t 1/2 28 h) than the clearance of DPC ( t 1/2 12 h). Liposome-associated dinitrophenyl-labeled rSP-C was taken up by MLE-12 cells, and the uptake was inhibited by excess nSP-C. The pattern of inhibition of dinitrophenyl-rSP-C uptake by SP-B, but not by SP-A, was similar to that previously reported for nSP-C. Clearance kinetics of nSP-C were similar to previous measurements of pulmonary clearance of SP-B in rabbits and mice. rSP-C has clearance kinetics and uptake by cells similar to those of nSP-C.

scholarly journals The reversible binding of vinblastine to platelets: implications for therapy

Blood ◽  
1981 ◽  
Vol 57 (3) ◽  
pp. 431-438 ◽  
Author(s):  
JG Kelton ◽  
JW McDonald ◽  
RM Barr ◽  
I Walker ◽  
W Nicholson ◽  
...  
Keyword(s):  
Half Life ◽  
Immune Thrombocytopenia ◽  
Suitable Model ◽  
Reversible Binding ◽  
In Vivo Binding ◽  
Rabbit Platelets ◽  
Kinetics Of ◽  
Plasma Half Life

Abstract The ability of platelets to adsorb vinblastine has been used to treat patients with immune thrombocytopenia. It is hypothesized that the drug- platelet complex is coated with antibody, taken up by macrophages which are then destroyed by the drug. We gave 16 courses of vinblastine- platelets to six patients with immune thrombocytopenia. Only one patient responded, and therefore we examined possible reasons for the lack of benefit. Using 3H-vinblastine, the kinetics of vinblastine binding to platelets was studied in vitro. The binding of vinblastine to both human and rabbit platelets was identical with maximal binding occurring within 10 min at 600 microgram/ml vinblastine. Similarly, the plasma half-life of vinblastine in rabbits was close to that reported for man, and therefore, in vivo binding of vinblastine to platelets in rabbits was considered a suitable model for man. Homologous donor rabbit platelets were labeled with 51Cr alone, 51Cr plus vinblastine, or 3H-vinblastine and infused into recipient rabbits. Vinblastine had no effect on 51Cr survival, but all measureable vinblastine had left the platelets within 2 hr of the infusion. These observations suggest that delivery of the vinblastine to the macrophages depends on the platelets being phagtocytized before the drug leaves the platelets. This would be likely to occur only in those patients with severe immune thrombocytopenia. Further investigations into this treatment should be directed at methods to maintain the drug within the platelet.

scholarly journals The reversible binding of vinblastine to platelets: implications for therapy

Blood ◽  
1981 ◽  
Vol 57 (3) ◽  
pp. 431-438
Author(s):  
JG Kelton ◽  
JW McDonald ◽  
RM Barr ◽  
I Walker ◽  
W Nicholson ◽  
...  
Keyword(s):  
Half Life ◽  
Immune Thrombocytopenia ◽  
Suitable Model ◽  
Reversible Binding ◽  
In Vivo Binding ◽  
Rabbit Platelets ◽  
Kinetics Of ◽  
Plasma Half Life

The ability of platelets to adsorb vinblastine has been used to treat patients with immune thrombocytopenia. It is hypothesized that the drug- platelet complex is coated with antibody, taken up by macrophages which are then destroyed by the drug. We gave 16 courses of vinblastine- platelets to six patients with immune thrombocytopenia. Only one patient responded, and therefore we examined possible reasons for the lack of benefit. Using 3H-vinblastine, the kinetics of vinblastine binding to platelets was studied in vitro. The binding of vinblastine to both human and rabbit platelets was identical with maximal binding occurring within 10 min at 600 microgram/ml vinblastine. Similarly, the plasma half-life of vinblastine in rabbits was close to that reported for man, and therefore, in vivo binding of vinblastine to platelets in rabbits was considered a suitable model for man. Homologous donor rabbit platelets were labeled with 51Cr alone, 51Cr plus vinblastine, or 3H-vinblastine and infused into recipient rabbits. Vinblastine had no effect on 51Cr survival, but all measureable vinblastine had left the platelets within 2 hr of the infusion. These observations suggest that delivery of the vinblastine to the macrophages depends on the platelets being phagtocytized before the drug leaves the platelets. This would be likely to occur only in those patients with severe immune thrombocytopenia. Further investigations into this treatment should be directed at methods to maintain the drug within the platelet.

scholarly journals Kinetics of biliary excretion of the main two bilirubin photoproducts after injection into Gunn rats

1981 ◽  
Vol 198 (1) ◽  
pp. 107-112 ◽  
Author(s):  
S Onishi ◽  
N Kawade ◽  
S Itoh ◽  
K Isobe ◽  
S Sugiyama ◽  
...  
Keyword(s):  
Biliary Excretion ◽  
Half Life ◽  
Gunn Rats ◽  
Kinetics Of

The kinetics of biliary excretion of the main two photoproducts after injection into Gunn rats were examined. The photoproducts that are obtained from experiments in vitro consist of unknown pigment, photobilirubin IXa and a small amount of (ZZ)-bilirubin IXa. It was confirmed previously that the first two photoproducts are identical with the main two photoproducts obtained in vivo. In experiments on four animals, the average of total biliary recoveries of unknown pigment was 81.4%, and that of photobilirubin IXa in the bile estimated by the Sigma-minus method was 29.8 min and that for unknown pigment was 4.3 min. The rate of thermal reversion of photobilirubin IXa to (ZZ)-bilirubin IXa in the bile at 37 degrees C was very rapid, i.e. its half-life was 6.2 min.

FRACTIONATION OF 3–8S IODOCOMPOUNDS OF THE RAT THYROID

1971 ◽  
Vol 67 (2) ◽  
pp. 225-240 ◽  
Author(s):  
O. Spira ◽  
A. Gordon ◽  
J. Gross
Keyword(s):  
Serum Albumin ◽  
Electrophoretic Mobility ◽  
Half Life ◽  
Polyacrylamide Gel ◽  
Pool Size ◽  
Rat Serum ◽  
Rat Thyroid ◽  
Rat Serum Albumin

ABSTRACT The 3–8S iodocompounds of the rat thyroid were separated by means of electrodialysis on polyacrylamide gel columns into a dialyzable fraction (D) and non-dialyzable fraction (ND). The non-dialyzable fraction consisted of 4 peaks, one of which had the electrophoretic mobility of rat serum albumin. All 4 iodoproteins of the non-dialyzable fraction were shown in vitro to incorporate 3H-Leucine. The dialyzable fraction consisted of 6 iodine containing peaks, all of which incorporated 3H-Leucine. The half life and relative iodine pool size of each compound were determined and compared to the 19S thyroglobulin fraction.

scholarly journals Biosynthesis of Rat serum albumin

1971 ◽  
Vol 123 (4) ◽  
pp. 649-655 ◽  
Author(s):  
J. D. Judah ◽  
Marion R. Nicholls
Keyword(s):  
Amino Acid ◽  
Serum Albumin ◽  
Time Course ◽  
Time Lag ◽  
Rat Serum ◽  
Albumin Content ◽  
Double Label ◽  
Rat Serum Albumin

1. The labelling of intracellular and extracellular serum albumin was studied in liver slices and in whole rats by using new methods for the purification of the protein. 2. The results suggest that a polypeptide precursor is formed that is converted relatively slowly into serum albumin. 3. The effect of liver cell K+has been examined by a double-label method and it is shown that K+accelerates the rate of conversion of ‘precursor’ into albumin. The rate of transit of albumin across the cell membrane appears to be unrelated to the concentration of K+within the cell. 4. The time-course of incorporation of radioactive amino acid into albumin follows a sigmoidal mode. There is a pronounced time-lag before label starts to appear in intracellular albumin, and a further time-lag before it appears in extracellular albumin. 5. In slices the sum of intra- and extra-cellular label rises steadily from 30min after the start of labelling with a pulse of labelled leucine or valine and continues to rise for at least another 60min. This occurs whether labelling is stopped by addition of excess of carrier amino acid or with cycloheximide (100μm) or both. 6. The intracellular albumin content remains constant whether slices are maintained with low or normal intracellular K+concentrations. 7. Specific radioactivities of intracellular albumin (and fractions thereof) and of extracellular albumin were determined in vitro and in vivo. The results show that the intracellular albumin cannot be a precursor of extracellular albumin, unless a very small compartment is turning over much more rapidly than the bulk of the liver albumin or even of the microsomal albumin.

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