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Tacrine derivatives containing iodobenzoic acid were developed as a novel multitarget-directed ligand and find potential application in the treatment of Alzheimer's disease. The aim of this study is to perform a physicochemical profile of this series. Experimental log P and pK a values were determined and compared with those already calculated. The results indicated better values of the tested compounds than the values predicted using computer software. The stability report was obtained using the developed HPLC method. The stability assay in different environment conditions provided information about the photosensitivity of these compounds and a proper method for the storage of this series of compounds., (© 2020 John Wiley & Sons, Ltd.)

Serum albumin (SA) is the most abundant protein in plasma and is the main transporter of molecules in the circulatory system of all vertebrates, with applications in medicine, the pharmaceutical industry and molecular biology. It is known that albumins from different organisms vary in sequence; thus, it is important to know the impact of the amino-acid sequence on the three-dimensional structure and ligand-binding properties. Here, crystal structures of ovine (OSA) and caprine (CSA) serum albumins, isolated from sheep and goat blood, are described, as well those of their complexes with 3,5-diiodosalicylic acid (DIS): OSA-DIS (2.20 Å resolution) and CSA-DIS (1.78 Å resolution). The ligand-free OSA structure was determined in the trigonal space group P3 2 21 at 2.30 Å resolution, while that of CSA in the orthorhombic space group P2 1 2 1 2 1 was determined at 1.94 Å resolution. Both albumins are also capable of crystallizing in the triclinic space group P1, giving isostructural crystals that diffract to around 2.5 Å resolution. A comparison of OSA and CSA with the closely related bovine serum albumin (BSA) shows both similarities and differences in the distribution of DIS binding sites. The investigated serum albumins from domesticated ruminants in their complexes with DIS are also compared with the analogous structures of equine and human serum albumins (ESA-DIS and HSA-DIS). Surprisingly, despite 98% sequence similarity, OSA binds only two molecules of DIS, whereas CSA binds six molecules of this ligand. Moreover, the binding of DIS to OSA and CSA introduced changes in the overall architecture of the proteins, causing not only different conformations of the amino-acid side chains in the binding pockets, but also a significant shift of the whole helices, changing the volume of the binding cavities.

The mechanism for the chirogenesis in the photocyclodimerization of 2-anthracenecarboxylate (AC) bound to human serum albumin (HSA) was investigated using time-resolved fluorescence measurements in the presence of HSA inhibitors and/or an AC singlet excited state quencher. The photophysical studies were correlated with product studies to explain the high enantiomeric excess (ee) observed for the chiral photoproducts. AC binds to HSA in five different binding sites with decreasing affinities. AC bound to the sites with the highest affinity (sites 1 and 2) is unreactive, and the AC can be displaced from these sites by the use of known inhibitors. Time-resolved fluorescence studies isolated a singlet excited state AC bound to a site which exhibited moderate protection from interactions with species in the aqueous phase. This site was assigned to binding site 3, where the chiral photoproducts are formed with a high ee based on the correlation of the photophysical studies with product studies in the presence of a quencher. These results show that the use of inhibitors for multiple binding site proteins is useful to uncover the properties of binding sites for which guest binding has only moderate affinity and where the photophysical characterization of these binding sites is not possible in the absence of inhibitors.

Background: The alpha-emitting nuclide 211At is of great interest for radionuclide therapy when coupled to a tumor-targeting biomolecule, e.g. epidermal growth factor (EGF) the receptors of which are overexpressed in many malignancies. However, almost no information concerning the cellular processing of astatinated targeting agents is available., Materials and Methods: We indirectly astatinated EGF ([211At]-benzoate-EGF) and studied its cellular processing in A-431 carcinoma cells in comparison with data concerning [125I]-benzoate-EGF., Results: The biological half-life of astatine (3.5 h) was longer than the half-life of the iodine label (1.5 h). The increase of the half-life was due to longer retention of the internalised astatine radioactivity. The maximum accumulation for the astatine label occurred later (4-6h) than that for the iodine label (2-4h), indicating a slower excretion of astatine that was confirmed in experiment with 211At/1251-benzoate-EGF., Conclusion: The long retention of astatine might be advantageous for radionuclide therapy.

ICP-MS, HPLC-ICP-MS and HPLC-ICP-MS/ESI-MS have been applied to determine the disposition and metabolic fate of 2-, 3- and 4-iodobenzoic acids following intraperitoneal administration at 50 mg kg(-1) to male bile duct cannulated rats. Quantitative excretion balance studies based on the determination of the total iodine content of urine and bile showed that all three iodobenzoic acids were rapidly excreted. Recoveries ranging from 95 to 105% of the administered doses were achieved within 24 h of administration. Metabolite profiles for urine and bile showed extensive metabolism with unchanged iodobenzoic acids forming a minor part of the total. A combination of alkaline hydrolysis and MS enabled the identification of the major metabolites of all three iodobenzoic acids as glycine and ester glucuronide conjugates with very little if any of the parent compounds excreted unchanged., (Copyright 2004 Elsevier B.V.)

In familial amyloidotic polyneuropathy, TTR (transthyretin) variants are deposited as amyloid fibrils. It is thought that this process involves TTR tetramer dissociation, which leads to partially unfolded monomers that aggregate and polymerize into amyloid fibrils. This process can be counteracted by stabilization of the tetramer. Several small compounds, such as diclofenac, diflunisal and flufenamic acid, have been reported to bind to TTR in vitro, in the T4 (thyroxine) binding channel that runs through the TTR tetramer, and consequently are considered to stabilize TTR. However, if these agents bind plasma proteins other than TTR, decreased drug availability will occur, compromising their use as therapeutic agents for TTR amyloidosis. In the present work, we compared the action of these compounds and of new derivatives designed to increase both selectivity of binding to TTR and inhibitory potency in relation to TTR amyloid fibril formation. We found two diflunisal derivatives that, in contrast with diclofenac, flufenamic acid and diflunisal, displaced T4 from TTR in plasma preferentially over binding to albumin and thyroxine binding globulin. The same diflunisal derivatives also had a stabilizing effect on TTR tetramers in plasma, as studied by isoelectric focusing of whole plasma under semi-denaturing conditions. In addition, by transmission electron microscopy, we demonstrated that, in contrast with other proposed TTR stabilizers (namely diclofenac, flufenamic acid and diflunisal), one of the diflunisal derivatives tested efficiently inhibited TTR aggregation. Taken together, our ex vivo and in vitro studies present evidence for the selectivity and efficiency of novel diflunisal derivates as TTR stabilizers and as inhibitors of fibril formation.

Renal elimination pathways of three positional isomers of iodobenzoic acid (2-iodobenzoate, 3-iodobenzoate and 4-iodobenzoate radiolabelled with 125I) were compared using the perfused rat kidney in-situ. All agents were eliminated both in a parent form (involving all renal elimination mechanisms i.e. glomerular filtration, tubular secretion, and tubular reabsorption) and also metabolized to a large extent in the kidney. After 3-iodobenzoate and 4-iodobenzoate administration, the major fractions of radioactivity found in urine were in the form of their metabolites, whereas 2-iodobenzoate was eliminated into urine mostly as the parent compound. Proportions of the individual metabolites in the urine of the perfused rat kidney were similar to those in intact rats for all agents. The results suggest that the kidney is the major organ for both the excretion and metabolism of iodobenzoates in rats. The principal renal metabolic reaction for all compounds under study was conjugation with glycine to produce the corresponding hippuric acid derivatives.

Halogenated organic substances are among the main environmental concerns. A number of micro-organisms are able to dehalogenate these compounds. However, the methods for the assessment of micro-organismal ability to dehalogenate are expensive and require complex instrumentation. Here, an easy colorimetric assay for the screening and assessment of the ability of bacterial cultures to deiodinate, and potentially dehalogenate, chemical substances is proposed. The method is based on the oxidation of iodide, released due to biotransformation, to iodine followed by a subsequent detection of iodine by a classical reaction with starch.

A denitrifying bacterium was isolated from a river sediment after enrichment on 3-chlorobenzoate under anoxic, denitrifying conditions. The bacterium, designated strain 3CB-1, degraded 3-chlorobenzoate, 3-bromobenzoate, and 3-iodobenzoate with stoichiometric release of halide under conditions supporting anaerobic growth by denitrification. The 3-halobenzoates and 3-hydroxybenzoate were used as growth substrates with nitrate as the terminal electron acceptor. The doubling time when growing on 3-halobenzoates ranged from 18 to 25 h. On agar plates with 1 mM 3-chlorobenzoate as the sole carbon source and 30 mM nitrate as the electron acceptor, strain 3CB-1 formed small colonies (1-2 mm in diameter) in 2 to 3 weeks. Anaerobic degradation of both 3-chlorobenzoate and 3-hydroxybenzoate was dependent on nitrate as an electron acceptor and resulted in nitrate reduction corresponding to the stoichiometric values for complete oxidation of the substrate to CO2. 3-Chlorobenzoate was not degraded in the presence of oxygen. 3-Bromobenzoate and 3-iodobenzoate were also degraded under denitrifying conditions with stoichiometric release of halide, but 3-fluorobenzoate was not utilized by the bacterium. Utilization of 3-chlorobenzoate was inducible, while synthesis of enzymes for 3-hydroxybenzoate degradation was constitutively low, but inducible. Degradation was specific to the positive of the halogen substituent, and strain 3CB-1 did not utilize 2- or 4-chlorobenzoate.

The cellular uptake of 11 radioiodinated aminoalkyl-iodobenzamides (ABA) was studied using cultivated murine melanoma cells (B16/C3). All derivatives showed a high uptake (up to about 80%) of radioactivity in melanotic melanoma cells; hence, accumulation of all compounds radioiodinated in the ortho position was reduced by approximately 30%. Using the compound para-[131I]iododiethyl-aminoethylbenzamide (p-131I-ABA-2-2) a close correlation of the cellular melanin content with the tracer uptake (R2 = 0.95) was verified. The presence of extracellular melanin, however, had no effect on the cellular tracer uptake. Because the accumulation was independent of the specific activity of p-131I-ABA-2-2, a significant contribution to the uptake process by binding to receptor sites could be excluded.

We identified a structural analog of thyroid hormone, methyl-3,5-diiodo-4-(4'-methoxyphenoxy) benzoate (Oncocidin A1), that inhibits human carcinoma cell proliferation and the growth of human breast (MDA MB-231) and ovarian (OVCAR-3) carcinoma xenografts in nude mice. This novel antitumor agent is orally bioavailable and well tolerated by animals. Exposure of MCF-7 and MDA MB-231 breast carcinoma cells to Oncocidin A1 in vitro caused a cell-cycle arrest in prometaphase (a G2/M arrest) and apoptosis, suggesting a cytotoxic mechanism involving mitotic spindle function. The interaction of Oncocidin A1 with microtubules was demonstrated by: 1) immunofluorescence studies of microtubule assembly in the presence of the drug in cell-free and in cellular assays; and 2) in vitro binding inhibition studies involving radiolabeled Oncocidin A1 or colchicine and tubulin monomers. Taken together, these experiments indicate that Oncocidin A1 perturbs cellular microtubule assembly, possibly by binding to the colchicine site on tubulin. Three-dimensional structural modelling of Oncocidin A1 revealed that it can adopt a twisted conformation similar to that of combretastatin A-4, which binds to the colchicine site of tubulin. The novel structural features of Oncocidin A1 could guide the design of a new class of microtubule-binding antitumor agents having substantially reduced normal tissue toxicity upon oral administration.

Two peptides of potential utility for targeting melanoma cells, alpha-melanocyte-stimulating hormone (alpha-MSH) and its more potent analogue [Nle4,D-Phe7]-alpha-MSH, were radioiodinated in 45-65% yield using N-succinimidyl 3-[125I]iodobenzoate (SIB). To determine whether this labeling method resulted in improved in vitro and in vivo characteristics, these peptides also were labeled with 131I by direct iodination with the iodogen method. For alpha-MSH, the rapid tissue clearance of both radionuclides in mice was consistent with rapid degradation of the peptide; however, significantly lower levels of 125I were observed in thyroid and stomach, reflecting a greater inertness to deiodination. More extensive comparisons were performed with [Nle4,D-Phe7]-alpha-MSH. The in vitro binding of [Nle4,D-Phe7,Lys11-(125I)IBA]-alpha-MSH (prepared using SIB) to the murine B-16 melanoma cell line, 34.1 +/- 4.7%, was more than twice as high as that for [Tyr2(131I),Nle4,D-Phe7]-alpha-MSH (15.0 +/- 0.1%), and its KD was more than 10-fold lower than that for conventionally labeled peptide (10 +/- 5 versus 140 +/- 14 pM). The normal tissue clearance of [Nle4,D-Phe7,Lys11-(125I)IBA]-alpha-MSH in mice was faster than that of [Tyr2(131I),-Nle4,D-Phe7]-alpha-MSH. The 19-40-fold lower activity concentrations of [Nle4,D-Phe7,Lys11-(125I)IBA]-alpha-MSH in tissues accumulating free iodide (thyroid and stomach) suggest a greater inertness of this peptide to deiodination. The primary urinary catabolite of [Nle4,D-Phe7, Lys11-(125I)IBA]-alpha-MSH was the lysine conjugate of iodobenzoic acid, whereas radioiodide was the chief catabolite generated from [Tyr2(131I),Nle4,D-Phe7]-alpha-MSH. We conclude that further evaluation of [Nle4,D-Phe7,Lys11-(125I)IBA]-alpha-MSH for targeting alpha-MSH receptors is warranted and that SIB may be a useful method for the radioiodination of peptides.

Two positional iodine derivatives of benzoic acid, i.e. ortho- (OIB) and para- (PIB), were used alone and in combination with salicylic acid (SA) to study the effects of plasma binding on their pharmacokinetics. Their lymphatic bioavailability (central lymph), their biotransformation and urinary excretion in rats were also studied. Plasma binding of the two benzoates is different, about 95% of PIB and approximately 50% of OIB are bound. The competitive inhibition effect of SA was shown by an increase in the amount of free drug in plasma in both benzoates. Lymphatic binding is lower compared to plasma binding, an effect of SA of the free faction of drug in lymph was shown only with PIB. Kinetic parameters of benzoates are influenced by plasma binding; significant differences were found mainly in total clearance and areas under concentration curves. Lymphatic bioavailability (FL) differs only slightly with different plasma binding; a significant change in FL was, however, found in PIB after SA premedication. Significantly higher urinary excretion of OIB as compared with PIB corresponds to plasma binding of drugs, SA premedication decreases total excretion of both benzoates. SA also changes the proportion of the individual fractions of metabolites of benzoates in urine.

In previous studies we have demonstrated that antibodies radioiodinated with N-succinimidyl 3-iodobenzoate (SIB) are less susceptible to loss of radioiodine in vivo than antibodies iodinated directly by electrophilic substitution on their tyrosine residues with Iodogen. Since the Bolton-Hunter reagent, N-succinimidyl 3-(4-hydroxy-3-iodophenyl)propionate, is identical with SIB except that it contains a hydroxyl group on the aromatic ring and a two-methylene spacer, a comparison of their coupling chemistry and in vivo behavior was performed to better understand the structural requirements for a useful iodinated acylation agent. Protein concentration and pH had a significant effect on the coupling efficiency of both SIB and the Bolton-Hunter reagent; however, protein-labeling yields with SIB were generally higher by a factor of 2. Paired-label biodistribution studies in mice demonstrated that thyroid uptake (a monitor of dehalogenation) of antibody labeled by the Bolton-Hunter method was twice that of antibody labeled with SIB but only 7% of that observed for antibody labeled with Iodogen. These results suggest that even minor differences in iodination site can profoundly alter the retention of label on a protein in vivo.

After suboccipital injection of Iotrol and Iopamidol labelled with iodine 125 in rabbits, we measured residual radioactivity in the whole brain and optical density on autoradiographs of brain sections obtained 2, 8 and 24 h after injection. Residual radioactivity is higher with Iotrol than with Iopamidol after 8 h and 24 h. At densitometry, while the penetration of the cortex is the same with both media at 2 h (although subcortical passage of Iotrol is greater), by 8 h the concentration of Iopamidol is twice that of Iotrol, and at 24 h it is three times as high. A similar pattern was seen in the subcortical region. These densitometric findings are in agreement with previous electrophysiological studies, in which changes were less severe and more transient with Iotrol than with Iohexol. There is nevertheless an apparent lack of agreement between the studies of radioactivity and the electrical findings. The lower neurotoxicity of Iotrol may be explained by: a longer half-life in the subarachnoid space; its larger molecules, which inhibit diffusion in the extracellular fluid, and its more hydrophilic nature, which reduces intracellular penetration.

125I-labelled o-iodobenzoate (OIB) was prepared by means of an isotopic exchange reaction; its distribution and excretion were determined and its pharmacokinetic parameters in rats were calculated. The calculated value of the half-life of OIB elimination was 38.7 +/- 0.7 min, the distribution volume was 278.2 +/- 53.5 ml . kg-1. The rate of elimination activity in urine was in agreement with the above values. On the basis of the developed technique of separation of OIB metabolites by thin-layer chromatography, their relative proportion in rat urine was determined; within 24 h 50% of the eliminated activity was in the original form (as OIB) and the metabolites of o-iodohippurate and o-iodobenzoylglucuronide formed approximately 25% of the activity eliminated in urine each.

1. Pig heart cytosolic malate dehydrogenase was radiolabelled with O-(4-diazo-3,5-di-[125I]iodobenzoyl)sucrose and intravenously injected into rats. Enzyme activity and radioactivity were cleared from plasma identically, with first-order kinetics, with a half-life of about 30 min. 2. The tissue distribution of radioactivity was determined at 2 h after injection. All injected radioactivity was recovered from the tissues. A high percentage of the injected dose was found in liver (37%), spleen (6%) and bone including marrow (19%). 3. Radioactivity in liver and spleen increased up to 2 h after injection and subsequently declined, with a half-life of about 20 h. 4. After differential fractionation of liver, radioactivity was largely found in the mitochondrial and lysosomal fraction. 5. Liver cells were isolated 1 h after injection of labelled enzyme. We found that Kupffer cells, endothelial cells and parenchymal cells had endocytosed the enzyme at rates corresponding to 2725, 94 and 63 ml of plasma/day per g of cell protein respectively. 6. Radioautography indicated that in spleen and bone marrow the enzyme is mainly taken up by macrophages. 7. Internalization of the enzyme by liver, spleen and bone marrow was saturable. This indicates that the enzyme is taken up in these tissues by adsorptive endocytosis. 8. The present results closely resemble those obtained previously for the mitochondrial isoenzyme of malate dehydrogenase and for lactate dehydrogenase M4. Since those enzymes are positively charged at physiological pH, whereas cytosolic malate dehydrogenase is negative, net charge cannot be the major factor determining the rate of uptake of circulating enzymes by reticuloendothelial macrophages, as has been suggested in the literature [Wachsmuth & Klingmüller (1978) J. Reticuloendothel. Soc. 24, 227-241].

Pharmacokinetical properties of eight triiodobenzene derivatives, X-ray contrast agents, were studied in the hemoglobin-free perfused rat liver with emphasis on the structural relation to biliary transport. With chemical modification of the basic structure, these agents showed different characteristics in the processes of diffusion into hepatocytes, accumulation in the cells and active transport into the bile, and were separated into four groups; [I]: Iotroxic acid (1), Iodipamic acid (2), Iodoxamic acid (3), and Ioglycamic acid (4) which showed faster rates of diffusion into hepatocytes [(1) greater than or equal to (2) greater than (3) greater (4)] and also of biliary excretion [(1) greater than (2) greater than (4) greater than (3)], [II]: Diatrizoic acid and Metrizamide showed poor diffusion and biliary excretion, [III]: Iopodic acid showed the highest permeability into and accumulation in hepatocytes with little biliary excretion, [IV]: ZK73 215 was slowly transported into the bile, yet, showed little permeation through the cell membrane. Characteristics of (1), (2) and (3) observed in the perfused liver were, in principle, confirmed in the pharmacokinetical profile observed in vivo. However, the fast diffusion of (2) into the hepatocytes appears to be hampered by high binding ability with serum proteins, whereas the relatively poor profile of the biliary excretion of (3) was improved by its low protein-binding in blood in vivo. Superiority of (1) as a cholangiographic agent was demonstrated by the fast biliary excretion in both the case of experimental systems and moderate protein-binding.

The bile acid taurocholate increases the biliary excretion of organic anions, such as sulfobromophthalein (BSP), bilirubin and iopanoic acid. In the present study has been investigated the effect of taurocholate on 1. Canine biliary excretion and concentration of the i.v. contrast medium ioglycamide and 2. Canine bile flow. The experimental model consisted of cholecystectomized, anaesthetized dogs with a fistula, through which the common bile duct could be catheterized and drained. One hour after cannulation, i.v. infusion of ioglycamide at a rate of 4 mumol/min./kg. was started. Two hours after the infusion start a control group received i.v. infusion of saline, while in another a 1.5% sodium taurocholate infusion was started with stepwise increases with 30 min. intervals from 0.4 to 0.8, 1.6 and 3.2 mumol/min./kg. Compared with control, all rates of taurocholate infusion increased bile flow and decreased biliary ioglycamide concentration. Although the bile flow with increasing taurocholate infusion rates was enhanced, the biliary ioglycamide excretion did not increase. The results indicate that ioglycamide and taurocholate are excreted into bile by separate excretion mechanisms. As taurocholate increases the biliary excretion of some other organic anions, it supports the hypothesis that organic anions are excreted into bile by more than two excretion mechanisms, taurocholate affecting only some of them.

The concentration of iodine in the bile of patients with indwelling T-tubes has been measured following administration of the new cholangiographic agent Iotroxamide. Studies have been performed after administration of the contrast agent by both 10 min bolus injection and 1 h drip infusion techniques. Comparison has been made with the iodine concentrations obtained after administration of equimolar amounts of Ioglycamide. There was no significant difference in the biliary iodine levels obtained with the two methods of administration when Iotroxamide was the contrast agent employed. However, with Ioglycamide the slow infusion technique produced higher iodine levels than the bolus injection method (P less than 0.05). Comparison between the two agents reveals that, whichever administration technique is used, Iotroxamide provides higher iodine levels than Ioglycamide. After bolus injection the superiority for Iotroxamide is in the region of 20% and after slow infusion is of the order of 10--15%. It is concluded that Iotroxamide is likely to prove superior to Ioglycamide as a cholangiographic agent. In terms of opacification of the biliary tree there is little to choose between bolus injection and slow infusion techniques when using Iotroxamide and the relative toxicity of the two techniques should be the major factor in determining which method is employed in clinical practice.

The absorption of the nonionic contrast medium iohexol, the clearance tracer 51Cr-ethylenediaminetetraacetic acid, and the blood-pool marker 125I-human serum albumin was studied after subarachnoid and subdural injection in rabbits. Subdural deposition of the contrast medium and 51Cr-ethylenediaminetetraacetic acid resulted in a faster absorption rate and higher achieved blood levels than a subarachnoid injection of the two substances, where a slow absorption to lower blood concentrations was observed. No significant differences in absorption rate could be shown after subdural and subarachnoid administration of iodine-labelled albumin. The excretion of iohexol was observed for 1 week after the intrathecal injection. For both the subdural and subarachnoid depositions, about 83% of the injected iohexol was found in urine within 24 hr after injection. The total recovery of iohexol after 1 week was 96% (range, 87%-101%).

The biotransformation of the 131I-labeled cholegraphic media ioglycamic acid, iodoxamic acid and iotroxic acid in man is investigated. Plasma, urine and fistular bile were analyzed for unchanged and metabolized constituents of the administered substances using thin layer chromatography. No metabolites were found in plasma, but up to two were found in urine in addition to unchanged contrast media (a total of 50% of the total elimination in 24 hr. urine). A metabolite was only found in the fistular bile after the injection of iotroxic acid.

The diffusion of water-soluble contrast media (CM) into the extracellular space of the central nervous system following injection into the subarachnoid space has previously been shown. As a result of this, water-soluble CM will come in direct contact with the neurons and may interfere with their normal function. The toxic effects would thus be a result both of the molecular properties of the CM as well as the local tissue concentration. The neuronal tissue uptake and clearance of metrizamide in rabbits following lumbar myelography was described in a previous study by our group. This study indicated some retention of metrizamide in the spinal cord probably as a result of binding of the CM to the cell membrane. The mechanism for this has not yet been shown although it may relate to the binding of metrizamide via its 2-deoxy-D-glucose (2-DG) portion and the specific glucose membrane carrier. The present investigation was performed to evaluate the diffusion kinetics of a new non-ionic CM. With iohexol, which lacks a 2-DG component in its molecule a direct relationship between the neural tissue and CSF concentration was found which seems to follow a simple diffusion model. Since iohexol shows no sign of entrapment in the tissue, the contact time for neurons will be shorter than that seen with metrizamide assuming that their rate of drainage from the CSF is identical.

Theophylline (TH), which has been shown in experimental dogs to increase bile-salt-independent bile flow, was studied in its effect on the biliary excretion and concentration of the intravenous contrast medium ioglycamide in cholecystectomized anesthetized dogs equipped with a Thomas cannula through which the common bile duct could be cannulated. One hour after cannulation, i.v. infusion of ioglycamide at the rate of 4 mol/min/Kg was started. Two hours later, 10 mg/kg of TH was injected intravenously and the experiment continued for a further 75 minutes. Bile was collected at 15 min. intervals throughout the whole experiment and simultaneous intravenous blood samples were taken. In this study, TH increased bile flow and decreased biliary ioglycamide concentration. Although TH increased bile flow, it had no effect on the biliary excretion of ioglycamide. It may be postulated that the organic anion ioglycamide, and possibly other organic anions, are secreted into the bile by mechanisms, unaffected by drugs which increase bile-salt-independent bile flow in a similar manner to TH.

The excretion of ioglycamate in the bile of the rhesus monkey was measured at 5% and at 100% bile diversion following an intravenous bolus injection of ioglycamate. At 100% diversion the bile volume was reduced and the concentration of ioglycamate was increased, but the quantity excreted was unchanged. A similar study using iodipamide reported previously gave the same result. When the ioglycamate was given by intravenous infusion, the effect of 100% bile diversion was quite different. The concentration of ioglycamate in the bile was unchanged by the bile diversion but the excretion was reduced. These results indicate that the transport maximum for the excretion of ioglycamate in bile is not a constant and is reduced by interruption of the enterohepatic circulation of bile salts. The maximum concentration of ioglycamate in bile was constant and was independent of the reduction in bile salt output produced by 100% bile diversion. Following a single bolus injection however, the reduction in bile flow produced by 100% bile diversion increased the biliary concentration of ioglycamate. These results suggest that the excretion of ioglycamate is limited by a maximum concentration rather than a transport maximum. The maximum rate of transport (Tm) is dependent on two factors--the maximum concentration of ioglycamate in the bile and the rate of bile flow. The maximum concentration is achieved by an infusion technique and not by a single bolus injection and this supports the view that an infusion technique should be used for intravenous cholangiography.

Lesions to the blood-brain barrier have been investigated in rabbits following constant or intermittent injections of sodium metrizoate into the internal carotid artery at different injection rates. Under certain experimental conditions the lesion to the blood-brain barrier was predominantly located to the brain area supplied by the contralateral carotid artery or the vertebral arteries. It is shown by serial angiography that the application time was shorter within the vascular territory of the paradoxical lesion. The results may imply that a mixture of blood and contrast medium is more vulnerable to the blood-brain barrier than contrast medium alone.

Plasma ioglycamide concentration was linearly related to the rate of intravenous infusion over a dose range of from 1 to 4 mg per kg per min. The relation between plasma and biliary concentration of ioglycamide was studied in 15 anicteric patients with a T-tube in situ. Peak biliary concentrations and excretory rates of ioglycamide were seen when the plasma concentration was greater than 1500 micrograms per ml. The mean biliary transport maximum (Tm) for ioglycamide in man was 31.6 mg/min (range 22.0-40.4). The results suggest that near optimal concentrations of iodine in the bile duct can be obtained during intravenous cholangiography if ioglycamide is infused for one hour at a rate of about 4 mg per kg per min.

Iodipamide and iodoxamate were compared in equimolar clinical dosages in five cholecystectomized chronic bile fistula dogs in which hepatic dysfunction was produced by oral administration of a total dose of 480 and 960 microliters dimethylnitrosamine (DMNA), respectively. After both DMNA dosages, the peak biliary excretion rate for iodoxamate was significantly higher than for iodipamide (p less than 0.01). The peak bile iodine concentration was not significantly different for the two agents (480 microliter DMNA: p less than 0.1; 960 microliter DMNA: p = 0.07). On the basis of this investigation, it is suggested that iodoxamate should not significantly improve the opacification of the biliary system in patients with hepatic dysfunction.

16 experiments were performed in order to investigate the mechanisms of biliary excretion of Ioglycamide in the mini-pig. The results confirm the mechanisms known from other species for this experimental animal. Additionally, however, they demonstrate a significant linear correlation between the spontaneous bile flow and the biliary transport-maximum of Ioglycamide. It is concluded that this correlation is due to the fact that the bile flow as well as the transport-maximum represent independent parameters of the excretory capacity of the liver cells. This conclusion is supported by further experiments that were unable to show an improvement of the biliary transport-maximum of Ioglycamide by stimulation of the bile flow during infusions of bile acids and secretin. These results prove the impossibility to enhance the biliary excretion of Ioglycamide by stimulation of bile flow. This, however, indicates that it is possible to dilute or concentrate the biliary Ioglycamide concentration by induced variations of bile flow, a fact that might become important to improve the efficiency of i.v. cholegraphy.

Of 50 successive in-patients with normal blood urea levels who received ioxaglate (Hexabrix 320; May & Baker Ltd), 60% showed gall-bladder opacification 12-25 h later. None of these patients had been asked to fast after receiving the ioxaglate. This has not been reported previously with any of the presently available urographic media, either conventional or the new low-osmolality agents. This should be regarded as a normal occurrence and does not imply impairment of renal function.

Pharmacokinetic data after injection of the new cholegraphic contrast medium iotroxic acid (Biliscopin) in man are reported and compared with the results of injections of ioglycamate and iodoxic acid. Iotroxic acid is less completely bound to plasma proteins than ioglycamate, but significancy more so than iodoxamate. Plasma protein binding depends on contrast concentration in the plasma, as does excretion in the urine. Biliary transport rate and maximal iodine concentration in the gall bladder are higher after injections of iotroxinate and iodoxamate than after ioglycamate.

The dynamic method originally developed for studying the capacity-limited kinetics of the cholecystographic agents iodoxamic acid and iopanoic acid was applied to study the in vivo interactions of these two compounds following coadminstration in the monkey. Results indicate that these interactions are complex. The compounds appear to compete for plasma protein binding sites as well as for binding sites on intrahepatic proteins. The biliary excretion data apparently fit the "ligand exclusion" model in which iopanoic acid acts as an inhibitor and competes with iodoxamic acid for binding to either of two identical sites within the liver. This competition probably is the rate-limiting step in the liver's overall elimination of these radiographic contrast agents.

Metrizamide, a new non-ionic, triodinated contrast agent has been considerable for intravenous urography. Theory predicts that lower osmotic diuresis with this agent should lead to higher urinary iodine concentrations than with ionic agents, at equivalent iodine doses. However, a distinct advantage with metrizamide has not been observed with doses of 175 mgI/kg or less. In this study metrizamide and sodium locarmate were given intravenously to dehydrated dogs at a dose level of 600 mgI/kg. During the studies the concentrations and outputs of major urinary solutes were compared. Comparisons of urinary iodine concentrations and outputs showed that at this dose the predicted advantage with metrizamide can be observed.

The effect of furosemide on biliary excretion of ioglycamide and bile flow during experimental cholangiography was studied in cholecystectomized and anaesthetized dogs equipped with a Thomas cannula through which the common bile duct could be cannulated. Ioglycamide infusion was started one hour after cannulation of the common bile duct, the infusion rate being 4 micromol/min/kg. Two hours later, 2 mg/kg furosemide was injected intravenously. After furosemide administration biliary ioglycamide concentration and bile flow remained the same as before the furosemide administration. Our evidence suggests that furosemide has no effect on the biliary excretion of ioglycamide or bile flow during ioglycamide cholangiography on dogs.

Fifty patients underwent myelography with the newly developed CSF-isotonic, dimeric, non-ionic contrast medium Iotrolan. Repeat spinal and cranial computerised tomographic studies with measurement of the attenuation values were conducted to demonstrate the administered contrast medium in the spinal canal and intracranial subarachnoid space. The patients were examined neurologically, observed clinically and asked about concomitant symptoms. The period of retention and the spread of the contrast medium in the subarachnoid space was not found to have any influence on the side effects rate. In particular, no association was confirmed between intracranially demonstrable contrast medium and the occurrence of side effects.

Active tubular secretion of iodamide, a water-soluble contrast medium, was demonstrated by comparing the clearances of iodamide, iothalamate and inulin. Passive tubular back diffusion was not found. The fraction excreted by tubular secretion was significantly reduced at "clinical" plasma concentrations. The glomerular filtration rate was slightly depressed at these concentrations of iodamide and diatrizoate.

When five patients with varying degrees of hepatic impairment and a T-tube in situ were given intravenous ioglycamide at a rate of 2 mg/kg/min for two hours the mean biliary excretion in the first two hours was only 3.2% of the administered dose. In contrast, in five T-tube patients with relatively normal liver function the mean biliary excretion over the same time interval was 20.6%. The mean plasma concentration of ioglycamide achieved at the end of a two-hour intravenous infusion at 2 mg/kg/min was 1427 +/- 187 microgram/ml in six anicteric patients and 1262 +/- 82 in six jaundiced patients. Despite these very similar plasma levels the 24-hour urinary excretion of ioglycamide was 42.3 +/- 3.8% of the administered dose in the patients with jaundice compared with only 18.1 +/- 2.4% in the anicteric group. These differences probably reflect the fact that the percentage of unbound contrast agent in the plasma of the jaundiced group (11.9 +/- 1.9%) was significantly higher than that of the anicteric group (6.4 +/- 0.9%). It is suggested that bilirubin and possibly other substances in the plasma are competing with ioglycamide for binding sites on albumin. These factors need to be borne in mind when performing intravenous cholangiograms on jaundiced patients.

Contrast media injected into the subarachnoid space may be eliminated in two different ways, by diffusion through the meningeal membranes or by transport through these CSF pathways. The latter mode of transport may be examined using computer assisted tomography of the head. Diffusion predominants with contrast media of low molecular weight, such as methiodal sodium. The transport through the CSF cihculation increases when contrast media of high molecular weight such as meglumine iocarmate and metrizamide are used.

Sixteen healthy men received iohexol intravenously at a concentration of 346 mg of iodine/mL. Doses of 500, 750, 1000, and 1500 mg of iodine/kg of body weight were administered to four volunteers each. Neither clearance nor percent of dose excreted in the urine showed any significant correlation with size of the dose. The overall mean (+/- SD) renal and total body clearances were 120 +/- 18.6 and 131 +/- 18.6 mL/min, respectively. The overall mean apparent volume of distribution was 165 (+/- 30.7) mL/kg. Urine contained 92.3 +/- 4.4% of the dose. Most of the drug (89.9%) was excreted within the first 12 h. An open three-compartment body model gave the best fit to the experimental data. The mean apparent first-order terminal elimination (gamma-phase) half-life was 12.6 h.

The maximun rate of excretion of ioglycamate in the bile of the rhesus monkey was achieved when the rate of adminstration was at least twice the rate of excretion. A maximum concentration of ioglycamate in the bile was also established, and this is more important to the visualization of the bileducts than the quanity of ioglycamate excreted. The maximum concentration was obtained at the same rate of infusion as that which produced the maximum rate of excretion. The peak concentration was sustained longer with an infusion lasting two hours than with one lasting 36 minutes, althought the same quanity of ioglycamate had been administered. It is concluded that an infusion at a rate equal to twice the maximum excretory rate and continued for two hours or longer is a rational approach to intravenuous cholangiography particularly in those patients where there has been some diffculty in bile-duct visulization.

Alcaligenes denitrificans NTB-1, previously isolated on 4-chlorobenzoate, also utilized 4-bromo-, 4-iodo-, and 2,4-dichlorobenzoate but not 4-fluorobenzoate as a sole carbon and energy source. During growth, stoichiometric amounts of halide were released. Experiments with whole cells and cell extracts revealed that 4-bromo- and 4-iodobenzoate were metabolized like 4-chlorobenzoate, involving an initial hydrolytic dehalogenation yielding 4-hydroxybenzoate, which in turn was hydroxylated to 3,4-dihydroxybenzoate. The initial step in the metabolism of 2,4-dichlorobenzoate was catalyzed by a novel type of reaction for aerobic organisms, involving inducible reductive dechlorination to 4-chlorobenzoate. Under conditions of low and controlled oxygen concentrations, A. denitrificans NTB-1 converted all 4-halobenzoates and 2,4-dichlorobenzoate almost quantitatively to 4-hydroxybenzoate.

The penetration into rabbit spinal cord of two nonionic contrast media, iohexol and metrizamide, and a reference tracer, technetium DTPA, were compared. The spinal subarachnoid space was perfused for 4 hours with a CSF solution to which technetium DTPA and either iohexol or metrizamide had been added. The contrast media and technetium DTPA concentrations reached a plateau level in CSF outflow within 80 minutes. The contrast media concentrations in CSF were higher than the technetium DTPA (P less than .001). In the cord tissue, technetium DTPA reached higher concentrations than the contrast media (P less than .001), and iohexol reached higher concentrations relative to technetium DTPA than metrizamide (P less than .001). The mean contrast media distribution volumes in the thoracic cord were 13% (iohexol) and 12% (metrizamide). The smaller distribution volume observed for metrizamide could be related to the larger effective size of "associated" metrizamide molecules or an interference with diffusion perhaps related to binding to glucose carriers.

Several classes of mutants of Pseudomonas putida (JT810) defective in the utilization of p-cymene as sole carbon source have been isolated. Selective enrichment of the mutants and for strains putatively cured of a degradative plasmid was achieved by incubation of cells in minimal growth media containing p-cymene (or p-cumate) and various halogenated analogs of the growth substrates or pathway intermediates. Analogs which led to successful enrichments included: p-chlorotoluene, p-bromotoluene, alpha-chloro-p-xylene, and p-iodobenzoate. A mutant strain, PpJT811, constitutive for the p-cymene pathway gave significantly greater enrichments of defective mutants than the wild-type parent PpJT810 after incubation with the halogenated analogs. It is suggested that the defective mutants are enriched because of the genetic alterations they possess, which confer immunity to a lethal synthesis performed by transformation of the analogs in clones possessing an intact p-cymene pathway. A nomenclature for the genetic organization of p-cymene pathway is described.