Passage of inhaled particles into the blood circulation in humans.

BACKGROUND: Pollution by particulates has been consistently associated with increased cardiovascular morbidity and mortality. However, the mechanisms responsible for these effects are not well-elucidated. METHODS AND RESULTS: To assess to what extent and how rapidly inhaled pollutant particles pass into the systemic circulation, we measured, in 5 healthy volunteers, the distribution of radioactivity after the inhalation of "Technegas," an aerosol consisting mainly of ultrafine (99m)Technetium-labeled carbon particles (<100 nm). Radioactivity was detected in blood already at 1 minute, reached a maximum between 10 and 20 minutes, and remained at this level up to 60 minutes. Thin layer chromatography of blood showed that in addition to a species corresponding to oxidized (99m)Tc, ie, pertechnetate, there was also a species corresponding to particle-bound (99m)Tc. Gamma camera images showed substantial radioactivity over the liver and other areas of the body. CONCLUSIONS: We conclude that inhaled (99m)Tc-labeled ultrafine carbon particles pass rapidly into the systemic circulation, and this process could account for the well-established, but poorly understood, extrapulmonary effects of air pollution.

Self-injection ictal SPECT during partial seizures.

The authors compared ictal SPECT injection performed by medical personnel with self-injection ictal SPECT in six patients with refractory temporal lobe epilepsy. Self-injection was safe and started faster. Self-injection subtraction ictal SPECT coregistered to MRI (SISCOM) was localizing in three patients who had a complex partial seizure, but only one of three patients who had a simple partial seizure, which may limit its usefulness in clinical practice. The localizing information of self-injection was better in three patients, and obviated the need for depth-EEG studies in one patient.

Scintigraphic evaluation in rabbits of nasal drug delivery systems based on carbopol 971p((R)) and carboxymethylcellulose.

The residence time of apomorphine mucoadhesive preparations incorporating 99mTc labeled colloidal albumin in rabbit nasal cavity was evaluated by gamma scintigraphy. This technique was used to compare the nasal clearance of preparations based either on Carbopol 971P((R)) or lactose (control), each with and without apomorphine, or carboxymethylcellulose with apomorphine. The planar 1-min images showed an excipient-dependent progressive migration of radioactivity with time from the nasal cavity to the stomach and intestine. Thirty minutes post insufflation, the percentages of the formulations cleared from the nasal cavity were 47% for lactose, 26% for lactose/apomorphine, 10% for Carbopol 971P((R)), and 3% for both Carbopol 971P((R))/apomorphine and carboxymethylcellulose/apomorphine. Three hours post insufflation, the percentages of the formulations cleared from the nasal cavity were 70% for lactose, 58% for lactose/apomorphine, 24% for Carbopol 971P((R)), 12% for Carbopol 971P((R))/apomorphine, and 27% for carboxymethylcellulose/apomorphine. Apomorphine inhibited nasal mucociliary clearance since migration of the radioactivity administered with apomorphine containing preparations was in all cases slower than that of the corresponding powder without apomorphine. The peak plasma concentration of apomorphine was attained while all the formulations were still within the nasal cavity. The use of mucoadhesive polymers such as Carbopol 971P((R)) or carboxymethylcellulose in nasal dosage forms increases their residence time within the nasal cavity and provides the opportunity for sustained nasal drug delivery.

Preparation and preliminary evaluation of 99mTc-EC-For-MLFK.

For-Met-Leu-Phe-Lys (For-MLFK), a chemotactic peptide that binds with high affinity to granulocytes and monocytes, was labeled with 99mTc using ethylene dicysteine (EC) as the metal chelating system. EC was selected because of the rapid renal excretion of its 99mTc-complex and therefore, was expected to enhance the degree of urinary elimination of the peptide-conjugate. 99mTc-EC-For-MLFK was prepared using a preformed chelate approach. After incubation of 99mTc-EC-For-MLFK with total blood, 68.1% of the labeled peptide was associated with WBC and 86% of this cell-associated activity was bound to granulocytes. Biodistribution studies in normal mice revealed a very fast blood clearance (4.1% and 0.6% of I.D. in blood at respectively 5 and 60 min p.i.). However, elimination of the labeled peptide proceeds mainly via the hepatobiliary system (24.5% of I.D. in liver and 48.8% of I.D. in intestines at 60 min p.i.) and to a much lower degree via the kidneys (17.9% in renal system at 60 min p.i.). From these results, it is concluded that 99mTc-EC-For-MLFK is not suited to image infections, despite its high binding to granulocytes, since it leads to high, non-specific, abdominal activity.

Synthesis and biological evaluation of the four isomers of technetium-99m labeled ethylenecysteamine cysteine ((99m)Tc-ECC), the mono-acid derivative of (99m)tc-L,L-ethylenedicysteine.

A few years ago (99m)Tc-ethylenedicysteine ((99m)Tc-L,L-EC) had been proposed as an interesting substitute for technetium-99m labeled mercaptoacetyltriglycine (MAG3) as renal function tracer agent. It possesses in its structure two carboxylate functions and is in this respect different from other renal tracers such as (99m)Tc-N, N'-bis-(mercaptoacetyl)-2,3-diaminopropionate ((99m)Tc-CO(2)DADS), (99m)Tc-MAG3, and Hippuran, which have only one carboxylic group. To study whether both carboxylic acid groups of (99m)Tc-L,L-EC contribute to the efficient renal handling of this compound we synthesized and biologically evaluated the technetium-99m labeled isomers of L- and D-ethylenecysteamine cysteine (ECC), the mono-acid derivative of (99m)Tc-L,L-EC. Labeling of L-ECC or D-ECC with (99m)Tc using a direct or exchange labeling method yields for each of them two diastereomeric (99m)Tc complexes (A and B, in the order of elution during reversed phase high performance liquid chromatography) in relative amounts depending on the pH during labeling. In mice, all four isomers of (99m)Tc-ECC (LA, LB, DA, and DB) are cleared rapidly from the blood, mainly by the renal system. The isomers LB and DB show the most efficient renal handling, but none of the mono-acid derivatives has a urinary excretion rate as high as that of (99m)Tc-L,L-EC. The renal handling of the isomers of (99m)Tc-ECC is partly due to tubular secretion because the urinary excretion of these compounds is significantly lower in mice pretreated with probenecid. In the baboon, isomers DA and DB show a plasma clearance comparable to that of (99m)Tc-L,L-EC. The plasma clearance of isomers LA and LB is lower but still comparable to or higher than that of (99m)Tc-MAG3. In a human volunteer, isomer DB shows a plasma clearance rate only slightly lower than that of (99m)Tc-L,L-EC. Thus, it appears that the presence of one carboxylate in (99m)Tc-EC-like compounds can be sufficient for efficient renal handling. However, it is also evident that the configuration at the chiral carbon atom and the orientation of the oxotechnetium core determine in a significant way the biological characteristics.

Importance of the two ester functions for the brain retention of 99mTc-labelled ethylene dicysteine diethyl ester (99mTc-ECD).

99mTc-ethylene dicysteine diethyl ester (99mTc-L,L-ECD) is a neutral lipophilic tracer agent that crosses the blood-brain barrier and is retained in the brain of primates following enzymatic hydrolysis of one of the ester functions to the ionized mono-ester, mono-acid metabolite. Up to now, it is not clear whether the second ethylcarboxylate group is essential for brain uptake and retention. Therefore, we have synthesized and studied two derivatives of 99mTc-L,L-ECD that contain only one ethylcarboxylate function, namely 99mTc-labelled L- and D-ethylene cysteamine cysteine ethyl ester (99mTc-ECCE). Direct labelling of L- or D-ECCE at neutral pH and room temperature resulted for each of them in the formation of two probably diastereomeric 99mTc-complexes in a 1:1 ratio. This means that four different isomers could be isolated. The 99mTc-labelled complexes formed after labelling ECCE (A and B, in order of elution during HPLC) are slightly less lipophilic than 99mTc-L,L-ECD. In mice, all four isomers show a low brain activity at 10 min post injection (p.i.), approximately 0.1% to 0.3% of the injected dose versus 0.9% for 99mTc-L,L-ECD. The clearance from the blood is comparable with (isomers LA and LB) or slower (isomers DA and DB) than that of 99mTc-L,L-ECD. Isomers LA and DA show high liver uptake and rapid excretion to the intestines. Both 99mTc-ECCE-LB and 99mTc-ECCE-DB, the most lipophilic isomers, are cleared from the blood mainly by the kidneys and excreted more efficiently to the urine. 99mTc-ECCE-LB is characterized by a surprisingly high heart uptake (about 1.5% of i.d.) at 10 min p.i. versus 1.0% for 99mTc-methoxyisobutylisonitrile (99mTc-MIBI) and 0.2-0.3% for the other isomers, but also lung uptake is relatively high. In the baboon, brain and heart uptake of isomers LA and LB of 99mTc-ECCE were negligible. Activity concentrated mostly in the hepatobiliary system for isomer LA and in the renal system for isomer LB. The results indicate a clear difference in biological behaviour between 99mTc-L,L-ECD and the four isomers of 99mTc-ECCE. This shows that the presence of both ester functions in 99mTc-L,L-ECD is an essential structural requirement for its brain uptake and retention in primates.

Complexes of technetium-99m with tetrapeptides, a new class of 99mTc-labelled agents.

Tetrapeptides are a class of N4-tetraligands that can efficiently bind 99mTc. In fact, tetrapeptides can be considered as derivatives of mercaptoacetyltriglycine (MAG3) in which the mercaptoacetyl moiety is replaced by a more stable and easier to handle aminoacyl group. Direct labelling of tetrapeptides with 99mTc in alkaline medium (pH > or = 11) in the presence of stannous ions gave a high yield (> 95%) of one or two (probably isomeric) radiochemical species. Exchange labelling at different pH values in the presence of stannous tartrate resulted in lower yields of the same 99mTc-labelled products as those formed during direct labelling. In addition, other radiochemical species were formed of which one was characterized as an oxotechnetium-complex with the cyclisized tetrapeptide. Tetrapeptides with a chiral centre in the first amino acid yield upon labelling with 99mTc two radiochemical species, probably the two diastereomers with an oxotechnetium core respectively syn and anti with respect to the substituent on the amino acid. Only one diastereomer was observed when the chiral carbon atom is located in the second or third amino acid. Electrophoresis indicated that these new 99mTc-labelled complexes are neutral in acidic medium and negatively charged in neutral and alkaline conditions. This correlates with a complex in which an oxotechnetium(V) group is bound to the ligand through three deprotonated nitrogen atoms of the amide functions and the free electron pair of the amine nitrogen atom. Biodistribution in mice showed for all studied 99mTc-labelled tetrapeptides a rapid clearance from the blood mainly by the renal system. The presence of a methyl substituent in the tetrapeptide increased the urinary excretion. 99mTc-labelled L-glycylalanylglycylglycine showed in mice a urinary excretion comparable to that of 99mTc-MAG3. Further rise of lipophilicity by introduction of a dimethyl, isopropyl or isobutyryl group leads to increased hepatobiliary handling. It is concluded that tetrapeptides are an interesting group of technetium complexing agents which can easily be labelled with 99mTc at room temperature in alkaline medium. This class offers the possibility of a wide variety of derivatives, just by substituting one or more amino acids. This group of ligands thus opens a new research field of 99mTc-complexes with potential usefulness in several areas.

Investigation of the labelling characteristics of 99mTc-mercaptoacetyltriglycine.

S-Benzyl-, S-benzamidomethyl- and S-benzoylmercaptoacetyltriglycine were synthesized and compared in exchange labelling experiments for the preparation of 99mTc-MAG3. The rate of exchange from 99mTc-tartrate to 99mTc-MAG3 starting from the respective precursors was determined in different conditions. Labelling proceeded most rapidly starting from the S-benzoyl protected precursor but efficient labelling was also accomplished using the more stable S-benzamidomethyl- and S-benzylmercaptoacetyltriglycine. 99mTc-MAG3 was also prepared by direct labelling of unprotected mercaptoacetyltriglycine at alkaline pH. Radiochemical purity in these conditions is mainly dependent on the pH during labelling.

Synthesis and evaluation of the (99m)tc-complexes of L-cysteine acetyldiglycine (a hybrid of MAG3 and L,L-EC) and of L-beta-homocysteine acetyldiglycine.

L-Cysteine acetyldiglycine (L-CAG2), a hybrid compound of L,L-EC and MAG3, and its L-beta-homocysteine analogue L-HAG2 were synthesized. After labeling with (99m)Tc, (99m)Tc-L-CAG2 and (99m)Tc-L-HAG2 gave two peaks on high performance liquid chromatography. Urinary excretion of both isomers of (99m)Tc-L-CAG2 and (99m)Tc-L-HAG2 was slower than for the "parent" complexes (99m)Tc-MAG3 or (99m)Tc-L,L-EC. Isomer B of (99m)Tc-L-CAG2 showed pronounced kidney retention in mice (57% of ID in kidneys at 30 min postinjection), but further evaluation in baboon did not reproduce this phenomenon.

Influence of a 99mTcN core on the biological and physicochemical behavior of 99mTc complexes of L,L-EC and L,L-ECD.

99mTc-nitrido complexes of L,L-ethylene dicysteine (99mTcN-L,L-EC) and 99mTcN-L,L-ethylene dicysteine diethylester (99mTcN-L,L-ECD) were prepared and their characteristics compared to those of the respective 99mTc-oxo complexes. 99mTcN-L,L-EC and 99mTcO-L,L-EC migrate to similar extents during electrophoresis at pH 12, but, at pH6, 99mTcN-L,L-EC migrates further than 99mTcO-L,L-EC. Renal excretion of 99mTcN-L,L-EC is inferior to that of 99mTcO-L,L-EC, indicating that the TcN-glycine sequence has lower affinity for the renal tubular system. Both 99mTcO-L,L-ECD and 99mTcN-L,L-ECD are neutral, but 99mTcN-L,L-ECD is hydrophilic and shows minimal brain uptake in both mice and the baboon.

Coronary radiation therapy with liquid rhenium-186: a first clinical experience.

BACKGROUND: Coronary radiation therapy (CRT) is a new, attractive approach for the treatment and prevention of restenosis after percutaneous coronary interventions (PCI). The RadioCath device consists of a standard balloon dilatation catheter that can be charged with a solution of sodium 186Re perrhenate, a predominant beta emitter. The safety and performance of this new device was evaluated in a pilot trial. METHODS AND RESULTS: Thirty-three patients with a de novo lesion in a native coronary artery were treated with the RadioCath device after successful angioplasty. The average dwell time to deliver a dose of 20 Gy at 0.5 mm into the vessel wall was 418+/-64 seconds. The treatment was well tolerated by most of the patients. In 79%, only one inflation cycle was required to deliver the prescribed dose. There were two procedural device-related complications (5.9%) and three minor procedural related in-hospital complications (9%). CONCLUSIONS: CRT using a balloon catheter device, charged with a sodium 186Re perrhenate solution, seems feasible and safe. Clinical and angiographic 6-month follow-up data are pending.

RP-HPLC separation of the diastereomers of technetium-99m labelled tropanes and identity confirmation using radio-LC-MS.

99mTc-TRODAT-1 (technetium(V)-oxo-2-[[2-[[[3-(4-chlorophenyl)-8-methyl-8-azabicyclo[3.2.1]oct-2-yl]methyl](2-mercaptoethyl)amino]ethyl]]amino]-ethanethiolato(3-)) and 99mTc-TRODAT-M, the 4-methylphenyl derivative of 99mTc-TRODAT-1, are at this moment being evaluated in clinical trials as imaging agents for the central nervous dopamine transporter system. Both compounds are formed as a mixture of two major diastereomers. As the tracer concentration in preparations for clinical investigations is very low (30-150 pmol/ml), identification of these 99mTc-complexes was, up to now, carried out indirectly using X-ray diffraction analysis of the corresponding rhenium complexes which can be synthesized in gram amounts. In this study, we developed a convenient and practical reversed phase HPLC-method for purification and isolation of the respective diastereomers of 99mTc-TRODAT-1 and three of its derivatives using mixtures of solvents which are compatible with biological studies, i.e. aqueous buffers and ethanol. Furthermore, direct identity confirmation of the 99mTc-complexes using radio-LC-MS was successfully elaborated.

An efficient HPLC method for the analysis of isomeric purity of technetium-99m-exametazime and identity confirmation using LC-MS.

99mTc-exametazime (99mTc-d,l-HMPAO, 99mTc-d,l-hexamethylpropyleneamine oxime) is a neutral rather unstable complex of short-lived 99mTc (t(1/2)=6 h) with the d,l-isomer (mixture of D,D- and L,L-isomers) of a bis-amine bis-oxime tetraligand. It is widely used for measurement of regional cerebral perfusion in nuclear medicine. The meso-isomer (D,L-form) should not be present in a preparation as it is not retained in brain and thus does not provide clinically useful information. Meso-HMPAO is removed from the ligand during the synthesis procedure by repeated recrystallization, but can still be present as impurity in d,l-isomer. Due to the lack of a suitable chromatographic method for analysis of the isomeric purity of 99mTc-exametazime preparations, United States Pharmacopoeia 25 (USP 25) prescribes a biological test in rats for quality control purpose. In this study, we developed a suitable high-performance liquid chromatography (HPLC) method which allows to demonstrate the relative amounts of d,l- and meso-isomer in 99mTc-exametazime and so obviates the need for a biodistribution test in animals as part of the quality control. Due to the low concentrations in which 99mTc-d,l-HMPAO is obtained (typically 2-6 ng/ml), confirmation of the identity of 99mTc-d,l-HMPAO in the monograph of the European Pharmacopoeia is now performed only indirectly by TLC and assessment of its retention time on RP-HPLC. To investigate the potential of radio-LC-MS for assessment of the identity of 99mTc-exametazime, 99mTc-d,l-HMPAO and 99mTc-meso-HMPAO prepared using a Tc-rich eluate were analyzed using a radio-LC-MS system equipped with a time-of-flight mass spectrometer with electrospray ionization. The main peak in the radiometric channel coincided with the molecular ion mass of 99mTc-d,l-HMPAO in the mass spectrometer channel and the measured accurate mass differed only by 0.26 ppm from the theoretical mass. The identity of 99mTc-meso-HMPAO was also confirmed. Thus, radio-LC-MS allowed to obtain strong evidence for the structure of 99mTc-d,l-HMPAO and 99mTc-meso-HMPAO at nanomolar concentration. It is concluded that radio-LC-MS can become a sensitive aid in quality control of "no carrier added" radiopharmaceutical preparations.

99Tcm-dimercaptopropionyl-HSA prepared from a labelling kit: preliminary investigations as a blood pool agent.

The blood retention of 99Tcm-dimercaptopropionyl human serum albumin (99Tcm-DMP-HSA), prepared from a kit, was compared with that of five other 99Tcm-labelled blood pool tracers in two healthy volunteers. 99Tcm-DMP-HSA showed an almost identical behaviour to in vitro labelled red blood cells (RBCs), which are generally considered the reference standard for blood pool agents. The mean apparent blood mass of 99Tcm-DMP-HSA was 2.1% higher 10 min post-injection (p.i.) than that of in vitro 99Tcm-RBCs, 2.0% higher 30 min p.i., 4.7% higher 60 min p.i. and 2.3% higher 120 min p.i. In vivo labelling of RBCs yielded a labelling efficiency of 75-98%, depending on the stannous agent used. About 20 min after pertechnetate administration, the intravascular activity as a percentage of injected dose stabilized at levels close to that of in vitro labelled RBCs. One commercially available 99Tcm-HSA kit was found to be unsuitable as a blood pool tracer. As 99Tcm-DMP-HSA offers the same practical advantages as 99Tcm-HSA, but better biological characteristics, it shows promise as a new tracer for radionuclide ventriculography and further large-scale investigations are warranted.

Effect of clopidogrel on naproxen-induced gastrointestinal blood loss in healthy volunteers.

The effect of clopidogrel, a potent inhibitor of platelet aggregation, on naproxen-induced faecal blood loss was investigated in 30 healthy volunteers in a randomized, double-blind, placebo-controlled, two parallel treatment groups study. All subjects first received naproxen 250 mg b.i.d. during 7 days, after which they were randomly allocated to additionally receive either clopidogrel 75 mg once daily (n = 15) or matching placebo (n = 15) for 11 days. Faecal blood loss was measured by the 51Cr-labelled erythrocyte method during the last four days of each of the four study periods, i.e. baseline, treatment with naproxen alone, combined treatment and wash-out. Mean daily faecal blood loss was below 0.5 ml/day during the baseline period in both treatment groups and increased during treatment with naproxen alone to (mean +/- SD) 1.14 +/- 0.58 ml/day in the naproxen + placebo group and to 1.93 +/- 1.51 ml/day in the naproxen + clopidogrel group. Addition of clopidogrel to naproxen treatment was associated with an increase of the mean daily blood loss to 6.83 +/- 9.32 ml/day, which was statistically significantly higher than 1.75 +/- 1.40 ml/day observed during treatment with naproxen + placebo. During the wash-out period, mean daily blood loss decreased to 0.98 +/- 0.51 ml/day in the naproxen + placebo group and to 1.07 +/- 0.46 ml/day in the naproxen + clopidogrel group. Based on these results, it can be concluded that clopidogrel increases naproxen-induced gastrointestinal blood loss in healthy volunteers. Caution should therefore be called for when these drugs are coadministered.

The effect of long-term treatment with spironolactone on variceal pressure in patients with portal hypertension without ascites.

The effect of spironolactone on esophageal variceal pressure (VP) in patients without ascites was investigated. VP was assessed using a noninvasive endoscopic gauge. Spironolactone was administered during a 6-week period at a dosage of 100 mg/d. This treatment decreased VP from 16.8 +/- 1.9 (SD) to 14.1 +/- 2.7 mm Hg (P < .001) in a group of 12 patients and from 18.6 +/- 2.1 to 13.7 +/- 4.1 mm Hg (P < .01) in another group of 8 patients who still had high VP despite chronic intake of propranolol. In both groups, placebo administration to 12 and 8 comparable patients did not significantly alter VP. Spironolactone induced a significant reduction of plasma volume (42.1 +/- 5.5 to 36.1 +/- 6.6 mL/kg body weight, P < .01) and of the concentration of alpha-atrial natriuretic peptide (alpha-ANP) (39.8 +/- 22 to 27.7 +/- 20 pg/mL, P < .01); in addition, a pronounced increase in plasma renin activity (PRA) (1.1 +/- 0.9 to 7.5 +/- 3.4 ng/mL/h, P < .001) was induced by the treatment. No significant changes in systemic hemodynamics were observed during the studies. Severe side effects were not observed except for a high incidence (55%) of painful gynecomasty in the male patients. In conclusion, chronic spironolactone administration effectively lowers VP, even in patients under chronic propranolol therapy. The combination of propranolol and spironolactone deserves further study as a prophylactic therapy of variceal hemorrhage, but development of gynecomasty might be a problem. Finally, we confirmed the reproducibility of VP measurements with the noninvasive gauge in chronic conditions.

Technetium-99m mercaptoalbumin as a potential substitute or technetium-99m labelled red blood cells.

Technetium-99m labelled red blood cells (99mTc-RBCs) are far superior to 99mTc-labelled human serum albumin (99mTc-HSA) for radionuclide ventriculography, but their labelling is more complex, time consuming and risk bearing (in vitro labelling) or suffers from interference by some medications (in vivo labelling). We have now modified HSA by the introduction of mercapto groups with the purpose of preparing stable and practical 99mTc-mercaptoalbumin with long retention in the vascular system, that could replace 99mTc-RBCs. HSA was incubated with N-succinimidyl S-acetylthioacetate (SATA) or N-succinimidyl 2,3-di(S-acetylthio) propionate (SATP) to introduce a chain containing one or two protected sulfhydryl groups on some of the lysine amino groups. After purification by size-exclusion chromatography (SEC), the mercapto groups were deprotected by incubation at alkaline pH or by treatment with hydroxylamine. The reaction products were used with or without SEC purification for direct or exchange labelling experiments with 99mTc at neutral pH. SEC-HPLC was used to determine labelling yields and to isolate pure 99mTc-mercaptoalbumin. Stable 99mTc-mercaptoalbumin complexes could be formed in 90%-95% yield after coupling albumin with SATA or SATP in all molar ratios used followed by deacetylation in one of the mentioned conditions. The most favourable results were obtained after reaction of SATA or SATP with HSA in a 25:1 ratio and deprotection with NH2OH. The stability of the resulting 99mTc-mercaptoacetyl-albumin (99mTc-MA-HSA) and 99mTc-dimercaptopropionyl-albumin (99mTc-DMP-HSA) and their retention in vivo in plasma of mice and rabbits are clearly higher than that of conventional 99mTc-HSA preparations. 99mTc-DMP-HSA approaches the behaviour of 125I-HSA quite well in both animal species. A preliminary study with 99mTc-DMP-HSA in a volunteer showed a retention in the vascular compartment almost identical to that of 99mTc-RBCs and clearly higher than that of a common 99mTc-HSA preparation. The results indicate that these 99mTc-mercaptoalbumins and especially 99mTc-DMP-HSA are very promising as a practical alternative to 99mTc-RBCs.

Synthesis and biological evaluation of the four isomers of 99mTc-cysteinyltriglycine, an amino substituted derivative of 99mTc-MAG3.

Cysteinyltriglycine (CYSG3) is a derivative of MAG3 in which the mercaptoacetyl group is replaced by a cysteinyl moiety. This implies the presence of a primary amino group on the ligand, as in case of p-amino-hippuric acid, the compound with the highest renal tubular secretion known. The present study was undertaken to investigate the influence of this amino group on the biological behaviour of complexes of 99mTc with MAG3-like molecules. The L- and D-isomers of cysteinyltriglycine were synthesized as S-benzyl N1-CBO protected precursors. After removal of the protective groups with Na/NH3, the isomers were labelled with 99mTc. This resulted in the formation of two diastereomeric complexes (A and B in the order of HPLC-elution) for each of them. The biodistribution of the four HPLC-purified isomers was tested in mice. Isomers DA and LB showed slightly superior or similar renal excretion characteristics compared to 99mTc-MAG3, whereas the two other isomers were cleared at a lower rate by the kidneys and more through the liver and the intestines. The results indicate that substitution of 99mTc-MAG3 with an amino function may somewhat improve the rate of renal excretion, but the configuration of the 99mTc-labelled complexes appears to be more important to its biological behaviour.

Technetium-99m labelled human serum albumin for ventriculography: a comparative evaluation of six labelling kits.

In this study we have compared the characteristics of six labelling kits for the preparation of technetium-99m labelled human serum albumin (99mTc-HSA) and evaluated the usefulness of the various 99mTc-HSA preparations as blood pool tracer agents. The amount of the principal ingredients, i.e. HSA and stannous ions, varies largely between the studied kits and this is probably a reason for the observed differences in the labelling rate. Analysis of the reaction mixtures after labelling of the respective kits with 99mTc showed in each preparation the presence of four to five radioactive components in variable relative amounts. The retention time of the main component on size-exclusion high-performance liquid chromatography (SEC-HPLC) was identical for all preparations. Biodistribution of the HPLC-isolated fractions was studied in mice. The components with the shortest and longest retention times on HPLC show poor retention in the plasma. The three intermediate fractions, including the principal peak, are initially retained relatively well in the blood (60%-70% of the injected dose after 10 min), but clearly to a lower degree than iodine-125 labelled HSA. Moreover, they diffuse out of the vascular compartment at a much higher rate than 125I-HSA. The biological behaviour of the main component of the various preparations was clearly different, despite the identical retention time on SEC-HPLC. Study of the total preparations in mice and a rabbit showed that two of them are cleared rapidly from the blood and cannot be considered valuable blood pool tracers. Diffusion of the other preparations out of the blood is slower but also considerable and compromises their use for ventriculography.

Stabilisation of high-activity 99mTc-d,l-HMPAO preparations with cobalt chloride and their biological behaviour.

It has been reported that the stability of a 1.11-GBq (30 mCi) technetium-99m d,l-hexamethyl-propylene amine oxime (HMPAO) preparation can be improved to up to 5 h by the addition of 200 micrograms CoCl2.6H2O within 2 min after reconstitution. However, it is not clear whether this method is also efficient for high-activity preparations (5.55 GBq) and whether this modified 99mTc-d,l-HMPAO has the same biological properties and can safely be used. We have now studied CoCl2-stabilised 99mTc-d,l-HMPAO preparations containing different amounts of "in-house" HMPAO ligand and SnCl2 and reconstituted with activities from 1.11 GBq to 5.55 GBq 99mTc. The characteristics of the generator eluates were also divergent, ranging from fresh eluates from a generator eluted less than 2 h previously to 4-h-old eluates from a generator not eluted during the preceding 72 h. Preparations containing up to 5.55 GBq 99mTc and as low as 2 micrograms SnCl2.2H2O can be efficiently stabilised for at least 6 h by the addition of CoCl2 shortly after reconstitution. Interestingly, it was found that the stabilisation method is not efficient if the cobalt ions are added prior to reconstitution of the preparation. This implies that the cobalt chloride cannot be incorporated in the labelling kit. Also, preparations with amounts of the ligand lower or higher than 0.5 mg formed the 99mTc-d,l-HMPAO complex with low radiochemical yield or showed rapid degradation. Therefore, combination of a subdivision and storage of Ceretec kits in fractions (as reported in the literature) is contra-indicated with this CoCl2 stabilisation method. CoCl2-stabilised Ceretec kits reconstituted with 5550 MBq 99mTcO4- and used 4-5 h after preparation retain the diagnostic usefulness of the fresh 1110-MBq preparation with regard to leucocyte labelling and brain imaging. Although baboon brain uptake of the stabilised preparation was 6%-9% lower, this small difference could not be distinguished in the tomographic images. The data obtained with both inhouse prepared d,l-HMPAO and Ceretec kits suggest that the eluate restrictions recommended by the Ceretec manufacturer can be neglected if the preparation is stabilised with Co2+ ions. Studies with 57Co-spiked CoCl2 added to d,l-HMPAO preparations demonstrated that the Co2+ ions clearly interact with the d,l-HMPAO ligand, probably to form one or more complexes. From biodistribution studies in mice it became evident that the toxicological profile of the Co2+ ions in the presence of d,l-HMPAO should be more favourable than that of cobaltous ions. For these reasons, it seems justifiable that CoCl2-stabilised 99mTc-d,l-HMPAO preparations should undergo rigorous studies to elucidate their clinical usefulness and pharmacological safety.