The effect of DMSO treatment on the composition of 99mTc(Sn)EHDP

To find an explanation for the reported positive effect of a dimethyl sulfoxide (DMSO) treatment on the performance of the bone scan agent technetium-(tin)-ethane-1-hydroxy-1, 1-diphosphonate, we compared the composition of the agent, prepared with and without treatment with DMSO by using high performance ion-pair chromatography (IPC).The preparation obtained with the DMSO treatment appeared to contain a larger fraction of large and highly charged polynuclear complexes than the preparation without the DMSO treatment. According to experiments by other investigators the smaller ^99mTc(Sn)EHDP complexes (early eluting components in IPC) give lower bone/blood ratios than the larger ones. The results presented in this paper show that the explanation for the effect of the DMSO treatment may be that the Sn-EHDP complexes which are removed by extraction with DMSO, give relatively small ^99mTc(Sn)EHDP complexes. Without these small complexes a superior bone scan agent is obtained.From experiments in which an excess of ⁹⁹TcO₄ over Sn(II) was used, it was concluded that at least one of the technetium complexes contains Sn(IV). On the other hand, the absence of Sn from a late eluting technetium complex was proven.     

Gel chromatographic analysis of the bone seeking radiopharmaceutical 99mTc(Sn)-EHDP: The influence of pH and EHDP-concentration on the size of its constituents

^99mTc(Sn)-EHDP complexes have been produced by reduction of TcO₄⁻ with Sn(II) in the presence of EHDP at varying pH and EHDP concentration. The mixture was separated by means of gel-chromatography with an eluent of the same composition and pH as the reaction mixture.It appears that at neutral pH larger complexes are formed than under acidic or basic reaction conditions. Larger complexes are also formed at higher EHDP concentrations.     

The separation of 99mTc(Sn) EHDP complexes by HPLC and GPC

For the characterization of the multi-component bone-scan agent ^99mTc(Sn)EHDP we have analysed the complex mixture with reversed phase ion pair chromatography (IPC) and soft gel permeation chromatography (GPC).With IPC five major complexes were found within a separation-time of 40 min. To avoid decomposition of the complexes during separation, the concentrations of EHDP and the reductant Sn(II) in the eluent had to be identical to the EHDP and Sn(II) concentrations used for the preparation of the complexes.To investigate the stability of the complexes we applied separation by IPC, followed by re-analysis of the fractions within 1 h and after 21–25 h.It appears that there is a state of equilibrium between the five complexes. Within 1 h after isolation the separated complexes were still more than 90% in their original form while after 20 h considerable amounts of the other complexes are found. The two complexes with the largest retention time with IPC are the most stable ones. When the total mixture was re-analysed after 26 h all five components appeared to be still present, but the relative amount of the most stable component had increased.Using GPC for the separation of the complex mixture, we found four major peaks within a separation time of 14 h.The elution orders of the complexes with the two separation methods are opposite.     

Different routes in aqueous media to some 99Tc complexes investigated through ultraviolet absorption spectroscopy

Different synthetic routes were followed for preparing ⁹⁹Tc complexes with iminodiacetate (IDA), N-(2,6-dimethylphenylcarbamoylmethyl)iminodiacetate (HIDA) and salicylate (SAL). Each ligand was added to ⁹⁹TcO₄⁻ and the reduction performed with Sn(II) or hydrazine; alternatively in the ligand exchange reaction it was mixed with chloro-complexes of ⁹⁹Tc in the oxidation states +3, +4 and +5. Comparative analyses, performed through u.v. absorption spectra, showed that the ligand exchange products of the Tc(III)-IDA, Tc(III)-HIDA and Tc(V)-SAL systems were coincident with those obtained from the reduction procedure, though yield and purity were different.     

Anion exchange chromatography of 99mTc(Sn)-MDP complexes: Influence of eluent composition,determination of void volume and charge of the main component

^99mTc(Sn)-MDP complexes have been prepared by reduction of ^99mTcO₄⁻ by Sn(II) in the presence of MDP. These complexes were separated on an anion exchange column. The necessity of addition of the ligand and the reducing agent to the eluent to avoid decomposition during chromatography is demonstrated.For the main component that is found at pH 5 the ionic charge was calculated according to the method of Wilson and Pinkerton [Anal. Chem. 57, 246 (1985)] and the method of Russell and Bishoff [Int. J. Appl. Radiat. Isot. 35, 859 (1985)]. With the first method a charge of −4.2 ± 0.3 was obtained, with the second a charge of −5.1 ± 0.6.An accurate method to determine the void volume of the ion exchange column is described.     

Improvement of the reproducibility of ion-pair HPLC of 99mTc(Sn)EHDP complexes and the influence of the Sn(II) concentration on the composition of the reaction mixture

The use of a non-volatile modifier and the simultaneous exclusion of oxygen by flushing with helium in ion-pair HPLC of ^99mTc(Sn)EHDP-complexes prevents spurious oxidation during the separation process. It thus enables the omission of the reducing agent from the eluent solution and the determination of the labelling percentage from the TcO₄¹-peak. In addition, retention times become more reproducible.     

Oxidation states of 99mTc in technetium chelates studied by the ICES method

The chemical shifts of ^99mTc core-electron binding energies were measured in solid samples by means of the internal conversion electron spectroscopy (ICES) method. Technetium chelates with citric acid, DTPA, EDTA and ethylenediamine-N,N′-tetraacetohydroxamic acid (EDTAHA) as ligands were prepared in solution at (2–4) × 10⁻⁷ M Tc concentration (no-carrier-added). The samples for ICES measurements were made by evaporation of the solution in vacuum to dryness. The following chemical shifts ΔE_B [K^99mTcO₄-^99mTc(chelate)] were found in the systems investigated: 1.9 and 4.1 eV for ^99mTc(Sn)citr; 2.1, 3.2 and 4.3 eV for ^99mTc(Sn)DTPA; 1.9 and 3.6 eV for ^99mTc(Sn)EDTA; 1.8 and 3.3 eV for ^99mTc(Sn)EDTAHA. Standard deviations of the shifts are 0.2 eV. These shifts were compared with those of inorganic technetium oxocompounds and correlated with oxidation states via a potential model. It was concluded, that these shifts refer to technetium oxidation states in chelates as follows: 1.8–2.1 eV to ^99mTc(V), 3.2–3.6 eV to ^99mTc(IV) and 4.1–4.3 eV to ^99mTc(III).     

The binding of 99mTc(Sn)-MDP complexes to human serum albumin and other blood proteins determined with gel chromatography and ultrafiltration

The binding of ^99mTc(Sn)-MDP to human serum albumin and other blood proteins was investigated by gel chromatography and ultrafiltration.During gel chromatography dissociation of the ^99mTc(Sn)-MDP-protein complex occurs: thus, it is not a suitable technique for the determination of protein binding.The values found with ultrafiltration have to be corrected for non-ultrafiltrable TcO₂·nH₂O. From the corrected values it can be concluded that binding of ^99mTc(Sn)-MDP to blood proteins does not play a role in the biodistribution.     

The influence of Sn(IV) on the mean size of 99mTc(Sn)MDP constituents at neutral pH

The effect of addition of Sn(IV), Mg(II) and Ca(II) on the mean size of the ^99mTc-containing constituents of a ^99mTc(Sn)MDP reaction mixture was determined by a gel chromatographic batch procedure that does not disturb the equilibria in the mixture.     

Preparation of [99mTc(DMPE)2Cl2]+ for myocardial imaging by using Fe-(DMPE) complexes

The preparation of [^99mTc(1,2-bis(dimethylphosphino)ethane)₂Cl₂]⁺ ([^99mTc(DMPE)₂Cl₂]⁺) for imaging the myocardium is investigated. Starting from Fe(III)- or Fe(II)-DMPE and ^99mTcO₄⁻ different preparation variants are compared. In these reactions either ascorbic acid or DTPA serves as an agent for complexing iron.A simple procedure using lyophilized initial components is represented. Its application yields [^99mTc(DMPE)₂Cl₂]⁺ with a radiochemical purity of more than 95%. Organ distribution studies performed in rats emphasize the high myocardial accumulation of this preparation.     

Aspects of 99mTechnetium binding from an ethane-1-hydroxy-1,1-diphosphonate-99mTc complex to bone

In vivo and in vitro experiments were under-taken to study factors influencing ^99mTc binding from an EHDP complex to bone. In vivo ^99mTc (IV) complexes of EHDP, ATP, citrate and ascorbate were injected into rats. The effects of pretreatment of rats with EHDP on EHDP- and ATP-Tc (IV) injection were examined. Citrate and ascorbate gave no bone scintigram, ATP gave a bone scintigram inferior to EHDP-Tc (IV). EHDP pretreatment deteriorated EHDP- and ATP-Tc (IV) scintigrams. Micro-autoradiography showed that Tc-activity is only found in the bone matrix of 15-day-old rats and that EHDP-pretreatment diminishes this labelling. In vitro experiments showed that EHDP also diminishes labelling of hydroxyapatite by EHDP-Tc (IV), but that an excess of Ca can augment this labelling. It is suggested that Tc (IV) binds to bone because EHDP binds to bone, permitting released Tc (IV) to bind to bone separately.     

The effect of formulation conditions on the distribution of 99mTc(NaBH4)-HEDP components separated by anion exchange HPLC

Reduction of ^99mTcO₄⁻ by NaBH₄ in the presence of HEDP leads to a mixture of ^99mTc-containing complexes which can be separated by anion exchange high performance liquid chromatography (HPLC). The distribution of complexes within this mixture can be varied by controlling pH, the concentration of HEDP, the concentration of technetium, and the presence of air. Suitable control of these formulation conditions can yield mixtures which consist of essentially (85%) one component. Arguments are presented to support the view that the components of ^99mTc(NaBH₄)-HEDP and ^99mTc(NaBH₄)-MDP mixtures are in fact oligomeric or polymeric complexes that can contain technetium centers in at least two different oxidation states.     

Radiochemical studies of 99mTc complexes of modified cysteine ligands and bifunctional chelating agents

The synthesis of four novel ligands using the amino-acid cysteine and its ethyl carboxylate derivative is described. The synthetic method involves a two-step procedure, wherein the intermediate Schiff base formed by the condensation of the amino group of the cysteine substrate and salicylaldehyde is reduced to give the target ligands. The intermediates and the final products were characterized by high resolution nuclear magnetic resonance spectroscopy. Complexation studies of the ligands with ^99mTc were optimized using stannous tartrate as the reducing agent under varying reaction conditions. The complexes were characterized using standard quality control techniques such as thin layer chromatography, paper electrophoresis, and paper chromatography. Lipophilicities of the complexes were estimated by solvent extraction into chloroform. Substantial changes in net charge and lipophilicity of the ^99mTc complexes were observed on substituting the carboxylic acid functionality in ligands I and II with the ethyl carboxylate groups (ligands III and IV). All the ligands formed ^99mTc complexes in high yield. Whereas the complexes with ligands I and II were observed to be hydrophilic in nature and not extractable into CHCl₃, ligands III and IV resulted in neutral and lipophilic ^99mTc complexes. The ^99mTc complex with ligand III was not stable and on storage formed a hydrophilic and nonextractable species. The biodistribution of the complexes of ligands I and II showed that they cleared predominantly through the kidneys, whereas the complexes with ligands III and IV were excreted primarily through the hepatobiliary system. No significant brain uptake was observed with the ^99mTc complexes with ligands III and IV despite their favorable properties of neutrality, lipophilicity, and conversion into a hydrophilic species. These ligands offer potential for use as bifunctional chelating agents.     

Determination of 99mTc valent form in solids by measurement of internal conversion electrons

The core-level internal conversion electron spectroscopy (ICES) was used to study no-carrier-added compounds of ^99mTc in the form of solid deposits. The following chemical shifts of the electron binding energies were determined: ΔE_B(NH₄TcO₄NaTcO₄) = 0.0±0.3 eV, ΔE_B(NH₄TcO₄TcO₂·2H₂ O) = 3.±0.3 eV. Furthermore, two valent forms were distinguished in the decay-induced ^99mTc species created from the Na₂⁹⁹MoO₄ in the NaOH matrix. The smallest detectaable amount of ^99mTc in the minor valent form was ∼30μCi (6 × 10⁻¹²g). The observed shifts ΔE_B(NH₄TcO₄NaTcO_x) = ±0.0±3 eV and ΔE_B(Nh₄TcO₄-NaTcO_y) = 2.1±0.3 eV indicate the chemical form NaTc(VII)O₄ for the NaTcO_x and either Na₂Tc(Vi) or NaTc(V0O₃ for the NaTcO_y. Shift calculations based on the ground -state potential model prefer NaTcO₃.     

Evaluation of 99mTc(CO)5I as a potential lung perfusion agent

IntroductionThe use of ^99mTc-macroggregated albumin for lung perfusion imaging is well established in nuclear medicine. However, there have been safety concerns over the use of blood-derived products because of potential contamination by infective agents, for example, Variant Creutzfeldt Jakob Disease. Preliminary work has indicated that Tc(CO)₅I is primarily taken up in the lungs following intravenous administration. The aim of this study was to evaluate the biodistribution and pharmacokinetics of ^99mTc(CO)₅I and its potential as a lung perfusion agent.Methods^99mTc(CO)₅I was synthesized by carbonylation of ^99mTcO_4? at 160 atm of CO at 170°C in the presence of HI for 40 min. Radiochemical purity was determined by HPLC using ⁹⁹Tc(CO)₅I as a reference. ^99mTc(CO)₅I was administered by ear-vein injection to three chinchilla rabbits, and dynamic images were acquired using a gamma camera (Siemens E-cam) over 20 min. Imaging studies were also performed with ^99mTc-labeled macroaggregated albumin (^99mTc-MAA) and ^99mTcO_4? for comparison. ^99mTc(CO)₅I was administered intravenously to Sprague–Dawley rats, and tissue distribution studies were obtained at 15 min and 1 h postinjection. Comparative studies were performed using ^99mTc-MAA.ResultsRadiochemical purity, assessed by HPLC, was 98%. The retention time was similar to that of ⁹⁹Tc(CO)₅I. The dynamic images showed that 70% of ^99mTc(CO)₅I appeared promptly in the lungs and remained constant for at least 20 min. In contrast, ^99mTcO_4? rapidly washed out of the lungs after administration. As expected ^99mTc-MAA showed 90% lung accumulation. The percentage of injected dose per gram of organ ±S.D. at 1 h for ^99mTc(CO)₅I was as follows: blood, 0.22±0.02; lung, 12.8±2.87; liver, 0.8±0.15; heart, 0.15±0.01; kidney, 0.47±0.08. The percentage of injected dose per organ ±S.D. at 1 h was as follows: lung, 22.47±2.31; liver, 10.53±1.8; heart, 0.18±0.01; kidney, 1.2±0.17. Tissue distribution studies with ^99mTc-MAA showed 100% lung uptake.Conclusion^99mTc(CO)₅I was synthesized with a high radiochemical purity and showed a high accumulation in the lungs. Further work on the mechanism and optimization of lung uptake of ^99mTc-pentacarbonyl complexes is warranted.     

99mTc(L-L)3+ complexes containing ether analogs of DMPE

Novel ^99mTc(L-L)₃⁺ complexes have been investigated for potential use in myocardial perfusion imaging. Bidentate chelators have been prepared that are based on substituent analogs of 1,2-bis(dimethylphosphino)ethane, onto which alkyl ether groups have been incorporated. The new ligands are: (1) MMPE, 1,2-bis(methyl methoxyethyl phosphino)ethane, (2) MIBPE, 1,2-bis(methyl methoxyisobutyl phosphino)ethane, (3) FURPE, 1,2-bis(methyl tetrahydrofuran phosphino)ethane, and (4) PYRPE, 1,2-bis(methyl tetrahydropyran phosphino)ethane. These ligands have been reacted with ^99mTc and the resulting complexes evaluated for myocardial imaging properties. ^99mTc(MMPE)₃⁺ exhibited the most favorable myocardial imaging characteristics in animal models. Results indicate that pendent ether moieties can improve the myocardial imaging properties of cationic technetium complexes.     

The influence of the method of preparation of the bone scanning agent 99mTc(Sn)EHDP on its tissue distribution in the rat

The tissue distribution in the rat of the bone scanning agent 99mTc(Sn)EHDP is investigated by three methods, viz. by dissecting the animals 3 and 24 h post injection, by scanning the animals with a rectilinear scanner and by whole body autoradiography, particular of micro-sections of the skeleton. The effect of varying the experimental conditions in the preparation of the bone scanning agent (pH and concentrations of Sn(II) and EHDP) is investigated. In some of the experiments additional amounts of Sn(II) and EHDP are added to the reaction mixture after the preparation of the bone scanning agent. In this way the influence of the Sn(II) and EHDP concentrations on the composition of the bone scanning agent, and their influence as such on tissue uptake can be separately investigated. The 99mTc-uptake in bone is compared with the distribution of the 99mTc over the two main products in the reaction mixture, viz. 99mTc(IV)-Sn-EHDP2 and 99mTc(IV)-Sn-EHDP. In addition, the stability in vitro of some of the preparations in urine and plasma and the in vivo binding of the 99mTc to plasma proteins are determined. A procedure for the preparation of the bone scanning agent is recommended.     

99mTc bone scanning agents--III. Preparation and gel chromatography of 99mTc(Sn)MDP complexes

The preparation of 99mTc(Sn)MDP was investigated as a function of pH, MDP concentration and Sn(II) concentration. The labeling efficiency was over 90% in the majority of the experiments and remained constant over the range pH 2-9. The MDP concentration had little effect, while the Sn(II) concentration had a significant positive influence. The complex formation appeared to be partly reversible. The formation of different complexes was investigated by means of gel chromatography under various experimental conditions. Altogether six complexes were found. At acid conditions two major complexes were found and at neutral pH one major complex. The presence or absence of a particular complex was mainly determined by the pH and by the MDP concentration. The Sn(II) concentration had very little effect. The results are compared with previous results of similar experiments with 99mTc(Sn)pyrophosphate.     

Biokinetics of bone tracers by means of deconvolution analysis--comparison of 99mTc MDP, 99mTc DPD and 99mTc EHDP

Transfer functions of 99mTc methylene diphosphonate (MDP), 99mTc 2,3-dicarboxypropane-1,1-diphosphonate (DPD) and 99mTc ethane-1-hydroxy-1,1-diphosphonate (EHDP) into bone and extravascular fluid of soft tissues were determined in 5 dogs by deconvolution analysis of the time-course of plasma, soft tissue and bone radioactivity. The transfer rates 5 min after injection--indicating the rapid exchange of the tracer between plasma and the extravascular fluid--decrease in the order MDP greater than EHDP greater than DPD (P less than 0.05). The transfer rates into bone--determined from transfer rates between 30 and 60 min--decreased in a different order, i.e. MDP greater than DPD greater than EHDP (P less than 0.05). The fractional bone uptake of diphosphonates estimated from the ratio of early to late transfer rates was slightly greater for DPD than for MDP and EHDP respectively. The difference between DPD and MDP was not significant (P greater than 0.05). The average bone and soft tissue concentrations of DPD 60 min after injection were greater than that of MDP and EHDP due to different plasma concentrations (DPD greater than EHDP greater than MDP), whereas the bone-to-soft tissue ratios decreased in the sequence MDP greater than DPD greater than EHDP (P less than 0.05).--Our results reveal different biokinetics of MDP, DPD and EHDP explaining variations in osseous and soft tissue uptake suggesting that deconvolution analysis could play an important role in bone scan interpretation.     

A micro-autoradiographical study of the localization of 99mTc(Sn)-MDP and 99mTc-MDP in undecalcified bone sections

The localization of ^99mTc(Sn)-MDP in bone tissue was compared with ^99mTc-MDP by means of microautoradiography of undecalcified bone sections. Sections of good histological quality were obtained by a rapid embedding method in methylmethacrylate. No differences were found in the localization of these radiopharmaceuticals in fetal rat calvariae after incubation in vitro or in rat femora after administration in vivo. In the incubation experiment, hydrolyzed ^99mTc was formed. The uptake was high in areas of new bone formation. No uptake was seen in cells or in resorbing areas. In compact bone ^99mTc(Sn)-MDP was predominantly taken up in the vicinity of blood vessels.