The identity confirmation of 99gTc-DMSA complexes by using NMR and HPLC–MS/MS methods

The analyses of ^99gTc-DMSA complexes prepared under alkali and acidic reactions were reported. Modern analytical, separation and spectral methods such as NMR (¹H-NMR, ¹³C-NMR, APT, COSY and HSQC) and Q-TOF HPLC–MS/MS system with ESI were employed to determine the identity and characterization of the products. The structure of ^99gTc(V)DMSA was clearly confirmed and its fragmentation path in negative and positive ionisation mode was suggested. The effect of ascorbic acid and new alternative labelling with the use of NH ₄ ^99g TcOCl₄ was examined. Surprisingly, ^99gTc(III)DMSA complex was not formed under acidic reaction conditions. ^99gTc(V)DMSA complex was the main reaction product under both experimental conditions. This result suggests the key role of ^99g/99mTc concentration during the process of radiopharmaceuticals preparation.     

Chromatographic and biological comparison of99mTc-DMSA prepared by direct labelling and ligand exchange reaction

^99mTc-dimercaptosuccinic acid /DMSA/ is one of the most favourable agents used for renal scintigraphy. This radiopharmaceutical was prepared in two different ways: by direct labelling of tin/II/-dimercaptosuccinic acid and by ligand exchange reaction from^99mTc-gluconate at tracer concentration of technetium under acidic condition. The complexes formed were compared using paper chromatography, thin layer chromatography, gel filtration and electrophoresis. Their biodistribution was studied on rats.     

A Freeze Dried Kit for 99mTc(V) Dimercaptosuccinic Acid

^99mTc pentavalent dimercaptosuccinic acid [^99mTc(V) DMSA], a useful agent for imaging thyroid medullary carcinoma and other tumors can be reliably prepared by addition of Na^99mTcO₄ to a freeze-dried mixture of DMSA and Sn (2:1 molar ratio). The radiochemical purity, stability and animal bio-distribution behaviour is similar to that of the agent made by addition of NaHCO₃ to DMSA (III) renal imaging freeze-dried kit.     

<Superscript>99m</Superscript>Tc-labelled biguanide derivatives: chemical speciation modelling thereof and evaluation in vervets

^99mTc-DMSA (dimercaptosuccinic acid) is known to be a safe and effective agent for static renal imaging. However, it has a long uptake time which is a limiting factor in diagnostic procedures and also leads to a relatively high radiation dose being administered to patients. There is a constant search for possible new renal imaging agents with a good resolution, kidney/liver contrast and low radiation dose to all organs. A series of biguanide derivatives (potential as non-insulin-dependent diabetes mellitus agents) labelled with ^99mTc were investigated as potential alternative kidney-imaging agents on theoretical grounds (in silico) and their biodistribution (in vivo) verified in a limited number of animal experiments. Such a dual approach has the benefit that it reduces the number of animal experiments needed to evaluate a potential radiopharmaceutical. The blood plasma model shows little or no complexation of the biguanide type ligands by the metal ions in blood plasma. It was therefore expected that these ligands will clear rapidly through the kidneys and liver (increased lipophilicity). These predictions were verified by studies on single vervets comparing them with ^99mTc-DMSA as gold standard. All the biguanide derivatives labelled with ^99mTc show liver, kidney and gallbladder uptake in vervets. It was shown that the agent ^99mTc-CBIG (carboxylbiguanide) has a very fast kidney clearance, which will reduce the dose to organs (as experienced for ^99mTc-DMSA), although it’s potential as a kidney agent is limited by its gallbladder uptake.     

The interaction of <Superscript>99m</Superscript>Tc with pyrimidine compounds

The reaction of ^99mTc of different oxidation states (+7, +4) with 2-thiouracil and 5-nitrobarbituric acid have been studied at different temperatures, pH and concentrations. The reaction mixtures have been analyzed at different times using thin layer chromatography (TLC) and a radio detector to show the peaks at the plates. ^99mTc is obtained from the Mo generators with oxidation state (+7). The use of SnCl₂ as a reducing agent gave ^99mTc with oxidation state (+4). It is very difficult to separate the complexes formed from the reactions in very small concentration. The percentage of ^99mTc and its oxidation state involved in the complexes can be determined. The labeling efficiencies (percentage of complex) in the reaction of ^99mTc⁺⁷ with 5-nitro-barbituric-acid increases mostly at pH  10. Both oxidation states of ^99mTc(+7, +4) can be detected at pH’s 4 and 10, but at pH  4, the reduced form ^99mTCO₂, is more pronounced. At pH  7 no complexes were detected and most of ^99mTc remains as ^99mTCO₄ ⁻ . By increasing the ligand concentration, the labeling efficiencies of the complex increases. For the reaction of ^99mTc of oxidation states (+4, +7) with 2-thiouracil at different temperatures and analytical times it is concluded that several complexes with different R_f values were observed in equilibrium and most of these complexes were unstable.     

Characterization of 99mTc(CO)3-iminodiacetic acid (IDA) and its comparison with 99mTc-(IDA)2 using compounds for hepatobiliary scintigraphy

The organometallic precursor of fac-[^99mTc(CO)₃(H₂O)₃]⁺ has attracted much attention because of the robustness and small size of Tc(I)-tricarbonyl complexes compared to Tc(V) complexes and the good labeling affinity with a variety of donor atoms. Among various ligand systems, an iminodiacetic acid (IDA) was proven as a good chelating group to form a Tc(III)-compelx as well as has been shown its potential as a chelating system for fac-[^99mTc(CO)₃] precursor. In an attempt to confirm the similarity and the difference between ^99mTc(CO)₃-IDA and ^99mTc-(IDA)₂-complex, M(CO)₃-IDA (M = ^99mTc, Re) complexes of disofenin, mebrofenin and N-(3-iodo-2,4,6-trimethyl phenylcarbamoylmethyl) iminodiacetic acid were prepared, and the biological evaluation of ^99mTc(CO)₃-disofenin was performed. The ^99mTc(CO)₃-IDA complexes were prepared with a high radiolabeling yield (>98%) in a quantitative manner and showed a negative charge. The in vivo pharmacokinetic behavior of ^99mTc(CO)₃-disofenin showed a similar biological activity to ^99mTc-(disofenin)₂ in that those complexes were quickly cleared from the blood by the hepatocytes and excreted into the gallbladder and intestine. Accordingly, the ^99mTc(CO)₃-IDA derivatives of disofenin and mebrofenin might be used as hepatobiliary imaging agents. Since an IDA is a promising chelator for ^99mTc-based radiopharmaceutical and the biological properties of ^99mTc(CO)₃-IDA derivative shows similar to that of ^99mTc-complex, a biomolecule containing IDA can be freely radiolabeled with fac-[^99mTc(CO)₃]-precursor or ^99mTc. However, the radiolabeling efficiency and the biological behavior demonstrates the favorable properties of ^99mTc(CO)₃-IDA compound for the development of a new imaging agent.     

Synthesis, quality control and biodistribution of 99mTc-Kanamycin

Summary Kanamycin is an antibiotic used for treatment of infections when penicillin or other less toxic drugs cannot be used. Kanamycin was labeled with technetium-99m pertechnetate using SnCl₂^. 2H₂O as reducing agent. The labeling efficiency depends on the ligand/reductant ratio, pH, and volume of reaction mixture. Radiochemical purity and stability of ^99mTc-Kanamycin was determined by thin layer chromatography. Biodistribution studies of ^99mTc-Kanamycin were performed in rats and rabbits. A significantly higher accumulation of ^99mTc-Kanamycin was seen at sites of S. aureusinfected animals (rat/rabbit).     

Radiochemical quality control of99mTc-labelled radiopharmaceuticals

Two methods for the determination of radiochemical purity of preparations labeled with^99mTc are described. Paper chromatography was used for separation of reduced^99mTc and free pertechnetate from the labeled radiopharmaceutical. Whatman 3MM paper was used first with a acetone and then with 1N NaCl, as the mobile phase. Instant thin layer chromatography was performed for comparison. The electrophoretic method was also applied, using 0.05 M Na-acetate and 0.017 M NaCl. Biodistribution of^99mTc-DMSA in the organs of experimental animals was followed in order to verify chemical control methods.     

Synthesis and biodistribution of 99mTc(CO)3-DMSA-MIBI in mice

Mixed ligand fac-tricarbonyl complex of [^99mTc(CO)₃-DMSA-MIBI] has been prepared starting from the precursor [^99mTc(OH₂)₃(CO)₃]⁺. The complex can be obtained in good yield and purity in a two-step procedure by first attaching meso-2,3-dimercaptosuccinic acid (DMSA, HOOCCH(SH)CH(SH)COOH) with [^99mTc(OH₂)₃(CO)₃]⁺, followed by addition of MIBI [tetrakis-2-methoxyisobutylisonitrile (CH₃OC(CH₃)₂CH₂-N≡C) copper(I) tetrafluoroborate] solution. The complex was characterized by TLC and HPLC and was studied by means of octanol-water partition coefficient, electrophoresis, stability in vitro, and normal mice experiment. Biodistribution in mice demonstrated that the complex showed higher myocardial uptake after 0.5-hour p.i. The ratios of heart/liver (%ID/g) in the case of ^99mTc(CO)₃-DMSA-MIBI was higher (1.88) than that observed in case of ^99mTc-MIBI¹ (0.93) after 0.5-hour p.i. (P<0.05). Results showed that the complex may be developed to a novel myocardial perfusion-imaging agent.     

Evaluation of labelling conditions,quality control and biodistribution study of 99mTc-5-aminolevulinic acid (5-ALA): a potential liver imaging agent

Labelling of 5-aminolevulinic acid (5-ALA) with ^99mTc was achieved by using SnCl₂·2H₂O as reducing agent. Radiochemical purity and labelling efficiency was determined by instant thin layer chromatography/paper chromatography. Efficiency of labelling was dependent on many parameters such as amount of ligand, reducing agent, pH, and time of incubation. ^99mTc labelled 5-ALA remained stable for 24 h in human serum. Tissue biodistribution of ^99mTc-5-ALA was evaluated in Sprague–Dawley rats. Biodistribution study (% ID/g) in rats revealed that ^99mTc-5-ALA was accumulated significantly in liver, spleen, stomach and intestine after half hour, 4 and 24 h. Significant activity was noted in bladder and urine at 4 h. High liver uptake of ^99mTc-5-ALA makes it a promising liver imaging agent.     

99mTc-labelled compounds prepared with tungsten(III) as the reducing agent

The application of K₃W₂Cl₉ as reducing agent in preparation of^99mTc-labelled compounds is described. Pertechnetate reduction was carried out in solutions of pH 2 and also of pH 5.5. DTPA, citrate, gluconate, HEDSPA and MDP were successfully labelled. Two types of labelled gluconate complexes were obtained. Complex I exhibited in rats an increased^99mTc affinity to kidneys (21%). Complex II exhibited an increased^99mTc affinity to bone (8.7%). Two types of labelled HEDSPA preparations were obtained: one yielded 29%^99mTc activity in bone, the other type exhibited only 13.4%. The results indicated the absence of mixed complexes.     

In vitro incorporation studies of 99mTc–alendronate sodium at different bone cell lines

Bisphosphonates can be labeled with Technetium-99m (^99mTc) and are used for bone imaging because of their good localization in the skeleton and rapid clearance from soft tissues. Over the last decades bone scintigraphy has been used extensively in the evaluation of oncological patients to provide information about the sites of bone lesions, their prognosis and the effectiveness of therapy by showing the sequential changes in tracer uptake. Since the lesion visualization and lesion/bone ratio are important utilities for a bone scanning radiopharmaceutic; in this study incorporation of ^99mTc labeled alendronate sodium (^99mTc–ALD) was evaluated in U₂OS (human bone osteosarcoma) and NCI-H209 (human bone carcinoma) cell lines. ALD was directly labeled by ^99mTc, radiochemical purity and stability of the complex were analyzed by radioactive thin layer chromatography and radioactive high performance liquid chromatography studies. For cell incorporation study, NCI-H209 and U₂OS cell lines were used with standard cell culture methods. The six well plates were used for all experiments and the integrity of each cell monolayer was checked by measuring its transepithelial electrical resistance (TEER) with an epithelial voltammeter. Results confirmed that ALD was successfully radiolabeled with ^99mTc. ^99mTc–ALD incorporated with NCI-H209 and U₂OS cells. The uptake percentages of ^99mTc–ALD in NCI-H209 and U₂OS cell lines were found significantly different. Since ^99mTc–ALD highly uptake in cancer cell line, the results demonstrated that radiolabeled ALD may be a promising agent for bone cancer diagnosis.     

Technetium-99m labeling of hippuric and p-amino-hippuric acid in the presence of DTPA: Biological and pharmacokinetic evaluation and comparison with99mTc-DTPA

^99mTc-Hipp as a modified^99mTc-DTPA, and^99mTc-PAH as a new renal agent are developed. Each veal of lyophilised kit contain hippuric (Hipp) or p-aminohippuric acid (PAH), diethyltriaminepentaacetic acid as calciumtrisodium salt (CaNa₃DTPA) and stannouschloride (SnCl₂·xH₂O), in molar ratio Hipp/PAH:DTPA=4∶1. They are high radiochemical purity radiopharmaceuticals, with hydrophilic character and low percentage of protein binding. The ITL chromatography and HPLC analyses of these labeled compounds have shown almost identical results as^99mTc-DTPA, but their biological behavior in rats confirm certain differences.^99mTc-Hipp is a renal agent clearing by the glomerular system, with better pharmacokinetical parameters than^99mTc-DTPA:t _1/2(α)=4.1 min,t _1/2(β)=198.6 min,K _cl=1.2·10⁻²min⁻¹ and a twofold value for blood clearance (Cl=2.07 ml/min).^99mTc-PAH is a quite different renal agent, rapidly secreted by kidney as a tubular secretion agent. Its pharmacokinetical parameters:t _1/2(α)=2.5 min,t _1/2(β)=41.7 min andK _cl=5.1·10⁻⁴ min⁻¹ are almost equal to those of^99mTc-MAG₃, but the blood clearance of Cl=5.22 ml/min is even higher than that of IOH clearance.     

Practicality of Tetragonal Nano-Zirconia as a Prospective Sorbent in the Preparation of 99Mo/99mTc Generator for Biomedical Applications

The feasibility of using tetragonal nano-zirconia (t-ZrO₂) as an effective sorbent for developing a ⁹⁹Mo/^99mTc chromatographic generator was demonstrated. The structural characteristics of the sorbent matrix were investigated by different analytical techniques such as XRD, BET surface area analysis, FT-IR, TEM etc. The material synthesized was nanocrystalline, in tetragonal phase with an average particle size of ~7 nm and a large surface area of 340 m² g^?1. The equilibrium sorption capacity of t-ZrO₂ is >250 mg Mo g^?1. The present study indicates that ⁹⁹Mo is both strongly and selectively retained by t-ZrO₂ at acidic pH and ^99mTc could be readily eluted from it, using 0.9% NaCl solution. A 9.25 GBq (250 mCi) t-ZrO₂ based chromatographic ⁹⁹Mo/^99mTc generator was developed and its performance was repeatedly evaluated for 10 days. ^99mTc could be eluted with >85% yield having acceptable radionuclidic, radiochemical and chemical purity for clinical applications. The compatibility of the product in the preparation of ^99mTc labeled formulations such as ^99mTc-EC and ^99mTc-DMSA was evaluated and found to be satisfactory.     

Studies on the porphine labeled with <Superscript>99m</Superscript>Tc–pertechnetate

The aim of this research is to use acetylacetonate as a ^99mTc chelating agent label with porphyrin and evaluate its radiochemical and biological characteristics. Stannous chloride was used as a reductant to determine the chemical and biological characterization of ^99mTc-complexes from labeling porphine{4′,4′′,4′′′-(2lH,23H-Porphine-5,10,15,20-terayl)tetrakis-(benzoic acid), TPPB} with ^99mTc–pertechnetate. Instant thin layer chromatography (ITLC), size exclusion chromatography (SEC), paper electrophoresis, and UV/Vis spectrophotometry were used to evaluate chemical characterization. Finally, biodistribution and liver function tests were applied to evaluate biological characteristics. The results of this study show that the labeling efficiency of ^99mTc(acac)–TPPB was nearly 100% when using acetylacetone (acac) as a conjugator. Three major ^99mTc(acac)–TPPB complexes were separated by SEC, and all of them were hydrophilic. The UV-Vis spectra of ^99mTc(acac)–TPPB complexes closely resembled those of the TPPB, but the wave lengths of their peaks changed 430, 521, 556, 591 and 647 nm after complexation. The biodistribution study selected the liver as the target organ. The ^99mTc(acac)–TPPB complex may cause short-term liver injury. However, this injury can be repaired, and the reagent is quickly metabolized. Hence, the toxicity of the ^99mTc(acac)–TPPB complex is within an acceptable range, and making it a promising liver imaging agent.     

Evaluation and comparison of 99mTc-labeled d-glucosamine derivatives with different 99mTc cores as potential tumor imaging agents

Isocyanide is a strong coordination ligand that can coordinate with [^99mTc(I)(CO)₃]⁺ core and [^99mTc(I)]⁺ core to produce stable ^99mTc complexes, therefore developing a ^99mTc-labeled isocyanide complex for single-photon emission computed tomography (SPECT) imaging is considered to be of great interest. In order to develop potential tumor imaging agents with satisfied tumor uptake and suitable pharmacokinetic properties in vivo, a novel d -glucosamine isocyanide derivative, 4-isocyano-N-(2,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-3-yl)butanamide (CN3DG), was synthesized and radiolabeled with [^99mTc(I)]⁺ and [^99mTc(CO)₃]⁺ cores to obtain [^99mTc(CN3DG)₆]⁺ and [^99mTc(CO)₃(CN3DG)₃]⁺ in high radiolabeling yields (>95%). Both of the complexes showed good hydrophilicity and great stability in vitro. Cell uptake studies performed in S180 cells demonstrated they were transported into cells by glucose transporters. Biodistribution studies of the two complexes in mice bearing S180 tumor showed they had high tumor uptakes and rapid clearance from muscle and blood so that the tumor/blood and tumor/muscle ratios were high. By comparison, [^99mTc(CN3DG)₆]⁺ was superior to [^99mTc(CO)₃(CN3DG)₃]⁺ in regard to tumor uptake, tumor/blood and tumor/liver ratios. S180 tumors could be seen clearly from the SPECT/CT images with [^99mTc(CN3DG)₆]⁺. Considering its favorable properties, [^99mTc(CN3DG)₆]⁺ would be a promising tumor imaging agent and needs to be further studied.     

Biodistribution and toxicity assessment of radiolabeled and DMSA coated ferrite nanoparticles in mice

DMSA-coated Fe₃O₄ nanoparticles were synthesized by wet-chemical method. The chemical interaction between Fe₃O₄ and DMSA were investigated by FTIR. They were directly radiolabeled with ^99mTc radioisotope (Fe₃O₄@DMSA–^99mTc) at room temperature in the presence of stannous solution as a reducing agent. Magnetic and structure properties of Fe₃O₄@DMSA–^99mTc nanoparticles were investigated by AGFM, TEM, and XRD. Biodistribution and toxicity assessment of Fe₃O₄@DMSA–^99mTc were studied in mice by intravenous and intraperitoneally injections, respectively. Blood, kidney, and liver factors were measured 4 days post injection and at the mean-while tissue sections were prepared from their kidney and liver. The results indicate that, the Fe₃O₄@DMSA–^99mTc nanoparticles were passed through the membrane of different cells but do not create any disorder in the kidney and liver function even in high doses such as 300 mg/kg.     

Synthesis of 99mTc-cationic steroid antimicrobial-107 and in vitro evaluation

Ceragenins/cationic steroid antimicrobials (CSAs) are a group of cholic acid derivatives with many properties that make them favourable for application as anti-infective agents. CSA-107 is also a member of this group that was labelled with ^99mTc by using SnCl₂·2H₂O as reducing agent and Na–K tartrate as transchelating agent. Labelling efficiency was optimized by varying the amount of reducing agent, pH, and time of incubation. Labelling efficiency and the stability of ^99mTc-CSA-107 in human serum was determined by paper and thin layer chromatography, which were >95 and >90 % respectively. In vitro binding of ^99mTc-CSA-107 was >95 % determined by using Staphylococcus aureus bacteria.     

Synthesis and biodistribution of the 99m TcN-DMSA complex as a potential bone imaging agent

In this study, the preparation of the 99m TcN complex of DMSA (dimercaptosuccinicacid) was carried out as a freeze-dried formulation, through a simple procedureinvolving the initial of 99m TcO 4 – with succinic dihydrazide in thepresence of stannous chloride as reducing agent, followed by the additionof the ligand DMSA to afford the final product. The radiochemical purity ofthe 99m TcN-DMSA complex was over 90% determined by thin layer chromatography.It was stable over 8 hours at room temperature. Its partition coefficientindicated that it was a good hydrophilic complex. Biodistribution in miceshowed that the 99m TcN-DMSA complex was accumulated in bone with high uptakeand good retention, suggesting it would be potentially useful as a bone imagingagent containing the [ 99m TcN] 2+ core. The biodistribution comparison inmice of the 99m TcN-DMSA complex and the 99m Tc-DMSA complex indicate thatthe presence of the 99m Tc nitrido group significantly alters the biologicalproperties of the 99m Tc complex.     

Chemical characterization of complexation behavior of pertechnetate with cysteine

The labeling behavior of cysteine with⁹⁹TcO ₄ ^– ion and/or^99mTcO ₄ ^– ion at different cysteine concentrations reductant and pH values has been studied by chromatography, and the labeling yield was calculated. Three major Tc-complexes, yellow, reddish brown and green can be separated by gel filtration chromatography (GFC). Thin layer chromatography (TLC), high performance liquid chromatography (HPLC) and ion-exchange chromatography (IC) were used to separate the complexes collected from GFC. The TLC, HPLC data show the pertechnetate accompanied with a yellow complex; the green and purple complex contain more than two complexes. Electrophoresis and IC data show that the complexes carry a negative charge. The conductivity, UV-VIS, flow beta-detector with HPLC and autoradiography are also applied to analyze complex formation.