Method of producing technetium-99m labelled nanocolloid

FIELD: chemistry.

SUBSTANCE: invention relates to a method of producing radioactive technetium-99m labelled nanocolloid. The method is characterised by adding a technetium-99m eluate to an aqueous suspension prepared from nanosized gamma-aluminium oxide powder with particle size of 7-10 nm and concentration of 0.5-0.7 mg/ml after bringing the pH of the medium to 4-5, followed by successive addition of ascorbic acid, tin (II) chloride dihydrate and gelatine in defined amounts. The obtained mixture is heated in a water bath at temperature 70-80°C for 30 minutes, cooled to room temperature in an ultrasonic bath and then subjected to sterilisation filtration.

EFFECT: invention enables to obtain 99mTc-Al2O3 nanocolloid, used based on its size to conduct diagnostic research for which not less than 80% of the particles have size in the range of 20-100 nm, and the radiochemical purity of the preparation is higher than 90%.

1 dwg, 2 tbl, 3 ex

 

The invention relates to medicine, in particular to a method for producing radioactive drugs for medical purposes, which can be used for radionuclide diagnostics, including for carrying out lymphoscintigraphy in Oncology.

In practice nanocolloids drugs labeled with short-lived radionuclide technetium-99m (99mTc), are widely used for diagnostics in Oncology, cardiology, for the detection of inflammatory diseases of musculoskeletal disorders anatomical and morphological patterns in tumors, cirrhosis, hepatitis and other diseases. Among such drugs commercially available in Russia is99mTc-Technetic (OOO "diamed"), which is a colloidal solution on the basis of phytin (salts insidestory acid). In addition, a method of obtaining nanocolloid for preparation of radiopharmaceuticals based rhenium sulfide [Patent RU No. 2315624 C2, 27.01.2008]. Data nanocolloids medicines99mTc, manufactured in Europe, presented in table 1 [V.I. Chernov, Afanasiev YEAR, Sinilkin A.A. and other Radionuclide research methods in identifying sentinel lymph nodes // Siberian Oncology journal. - 2008. V.28. No. 4. - C.5-10].

Australia is mechen the th technetium-99m nanocolloid drug-based fluoride, tin, used for more than 20 years in clinical practice for labeling white blood cells in the diagnosis of infection and inflammation [.Tsopelas. The radiopharmaceutical chemistry of99mTc-tin fluoride colloid-labeled-leukocytes // The quarterly journal of nuclear medicine and molecular imaging. - 2005. Vol.49, P.319-324].

All these nanocolloids drugs are based on compounds that form stable hydrosols. The deciding factor of success is not their chemical composition and size of nanoparticles. It is known, for example, that the optimal particle size for holding lymphoscintigraphy is 20-100 nm. Such particles are derived from tissues with speed, not allowing them to enter the bloodstream. Particles with sizes less than 20 nm can pass into the bloodstream, which prevents the visualization of lymph nodes [Sampson SV Textbook of Radiopharmacy Theory and Practice. Vol.3, 2nded. London, United Kingdom: Gordon and Breach; 1994: 196]. Nanoparticles with sizes larger than 200 nm (Hepatocis) can be used for labelling auto leukocytes to identify foci of inflammation in cardiology.

The majority of known nanocolloids radiopharmaceuticals is a simple inorganic complexes99mTc with rhenium sulfides and antimony obtained sufficiently complex technologies. For example, known is a method of obtaining nanocolloid sulphide of antimony [Lin Y., Zhang X., Li J. et al. Appl. Radiat. Isot, 58 (2003), 347-352], according to which the and the first stage gaseous hydrogen sulfide is bubbled through 100 ml of distilled water, cooled to 0°C, followed by introduction to this solution 5 ml of a 3.5% solution of polyvinylpyrrolidone. Then add 10 ml of 1% solution of antimony-potassium tartrate. To remove excess of hydrogen sulfide through the educated Hydrosol miss nitrogen. In the second stage, 10 ml of the obtained nanocolloid sulphide of antimony mixed with the previously prepared mixture consisting of 60 ml of 0.1 M solution of thioacetamide with 10 ml of 1%aqueous solution of antimony-potassium tartrate. The mixture is neutralized to pH 5-6 with a solution of 0.1 M HCl, heated on a water bath for 6 h and cooled to room temperature. And, finally, in the third stage, carry out the tagging of the obtained colloid technetium-99m. With this purpose in a vial with 1.5 ml of nanocolloid antimony administered 0.5 ml of the eluate99mTc, add 0.1 ml of 0.1 M HCl solution and heated in a water bath for 30 min and then adding to the cooled mixture of 0.5 ml of acetate buffer until neutral.

Approximately the same number of stages includes a method of obtaining99mTc-rhenium sulfide nanocolloid [Tsopelas C.J. Nucl. Med., 42 (2001), 3, 460-466]. However, we conducted preliminary studies have shown that stable colloidal compounds can be obtained in a simple way - by conducting adsorption restored99mTc on gamma-alumina [Skuridin B.C., Stasiuk Y.S., Sadkin V.L., Lapwings E.V., Horns A.S., Ikawa .. The study of static and dynamic adsorption of technetium-99m aluminum oxide // Izvestiya vuzov, W. Physics - 2010. - t. No. 10/2, - s-300]. When the amount of adsorption of the radionuclide on the surface of the oxide exceeds 93%. To restore99mTc (VII)present in the original eluate99mTc, we used tin (II) chloride dihydrate (SnCl2·2H2O, hereinafter referred to as Sn (II)). The maximum adsorption was observed at pH 4-6.

The main prerequisites for the use of gamma-oxides of Al2O3as the "media" tag99mTc is its lower toxicity than the sulphides, in combination with good adsorption properties, availability, and low cost. Suffice it to say that at this sorbent "holds" all the world production99Mo/99mTc-generators for medicine.

Ways to get labeled99mTc nanocolloid gamma oxide Al2O3our literature is not detected, which determined the task of developing a method of obtaining nanocolloid99mTc-Al2O3acceptable sizes for holding lymphoscintigraphy and other diagnostic tests, and meet the following requirements:

- at least 80% of the particles should have a size in the range of 20-100 nm;

- radiochemical purity of the drug should be not less than 90%.

In the accordance with this, the task of obtaining nanocolloid 99mTc-Al2O3with the given dimensions is solved as follows. In the way with technetium-99m of nanocolloid in aqueous suspension prepared from nanosized powder of gamma-aluminum oxide with a particle diameter of 7-10 nm and a concentration of 0.5-0.7 mg/ml, after adjusting pH to 4-5, enter the eluate of technetium-99m, then in quantities per 1 ml of the mixture sequentially injected ascorbic acid 0,20-0,25 mg tin (II) chloride dihydrate 0,00875-0,0175 mg and gelatin 2,5-4,0 mg, the resulting mixture is heated on a water bath at a temperature of 70-80°C for 30 min, cooled to room temperature in an ultrasonic bath and spend a sterilizing filter.

Among the factors complicating obtain the desired nanocolloid99mTc-Al2O3with the given dimensions, is that when you restore99mTc (VII) tin (II) optionally formed large colloid (>220 nm) type (-O-TcO-O-SnCl2-O-TcO-)nwhere n=2, 3... - the number depending on the pH of the solution [Zaitsev L.L., Velichko A.V., Vinogradov I. Compounds of technetium and their applications // Results of science and technology. - M.: VINITI, 1984. - V.9. - S]. In addition, the formation of tin colloid due to hydrolysis of the excess SnCl2proceeding according to the equation: SnCl2+H2O→Sn(OH)Cl+HCl.

Therefore, we have conducted preliminary research the Oia to establish necessary and sufficient amount of Sn (II), providing a full recovery99mTc (VII) in the eluate from99Mo/99mTc generator. To this end, in the eluate99mTc was injected different amounts of Sn (II) with subsequent determination of the content of unrecovered99mTc (VII) by thin layer chromatography on silica gel plates in the environment of acetone [GF XII, part 1, s]. For stabilization of the formed complex99mTc (IV) in the eluate before the introduction of the Sn (II) was added ascorbic acid based 0,20-0,25 mg per 1 ml of the eluate. The results are shown in table 2.

It follows that the optimal amount of Sn (II) in the reaction mixture, ensuring that99mTc (VII) less than 10%corresponds to a value ranging from 0,00875 to 0,0175 mg/ml

Introduction to the composition of the reaction mixture of gelatin due to the following. According to the chemical nature of the gelatin is a biopolymer - polydispersed mixture of low molecular weight peptides. The length of the gelatin molecules have an average 2850 Å, with a diameter of 14 Å. The average size of interstices is about 80-90 Å. Therefore, the matrix system based on gelatin characterized by nanostructural organization that provides a fairly uniform distribution of the immobilized substance and good steric accessibility of molecules for different chemical processes, interfering in the data case, the formation of large colloid tin with 99mTc [Mikhailov O.V. How to combine chemical jug" of fragments. // Nature. No. 12, in 2003. - P.17-24]. The content of gelatin in the composition known nanocolloids drugs, for example, "Nanosys" with a particle size of 100 nm is in the range of 2.5-4 mg/ml, which, as was confirmed by our research, is enough to reduce the number of large colloid tin with99mTc to an acceptable value of 20%.

The technical result from the proposed invention consists in receiving technetium-99m of nanocolloid that meets the following requirements:

- at least 80% of the particles have a size in the range of 20 to 100 nm;

the relative content of particles with sizes less than 20 nm does not exceed 6%;

- radiochemical purity of the radiopharmaceutical is over 90% and is maintained for at least 4 hours.

The invention is illustrated by figure 1, which presents lymphoscintigram rats 1 hour after injection under the skin of the animal nanocolloidal isotope99mTc-Al2O3marked: 1 - lymph node, 2 - site of administration of the drug. Accumulation in the lymph node of 1.5%.

The invention is illustrated by the following examples.

Example 1. In a bottle with a capacity of 20 ml contribute a portion of the gamma oxide Al2O3with a particle diameter of 7-10 nm mass of ~5 mg and throw it in 10 ml of water for injection. Then bring the pH of 0.05 M rest the rum HCl to a value of 4-5. 2 ml of the resulting suspension is transferred to a separate vial and consistently add 2 ml of the eluate99mTc activity 1.5 to 2 GBq, 100 μl of an aqueous solution of ascorbic acid at a concentration of 10 mg/ml and 10 ál of freshly prepared solution of Sn (II) concentration 7 mg/ml of SnCl2·2H2O). After intensive mixing, the mixture is heated on a water bath (70-80°C) for 30 min and then cooled to room temperature in an ultrasonic bath and filtered through a sterilizing filter (0, 22 μm) under aseptic conditions in a sterile vial.

Radiochemical yield with a particle size less than 100 nm relative to the total activity of the prepared drug was determined by filtering the mixture through a filter with pore diameter of 100 nm. The content of particles smaller than 20 nm was controlled according to the accumulation of99mTc in the blood after 1 hour after drug administration to experimental animals. In the above example, the entered quantity of ascorbic acid and Sn (II) in terms of 1 ml of prepared mixtures are respectively equal to 0.25 and 0,0175 mg of the product with dimensions less than 100 nm was 23%, and particles less than 20 nm - 6%. The radiochemical purity of the filtrate is equal to 76% and remains at this level for 4 hours.

Example 2. The reaction mixture is prepared as in example 1 with the difference that PEFC is the introduction of a freshly prepared solution of Sn (II) the mixture was added 100 μl of an aqueous solution of gelatin with a concentration of 10 mg/ml The resulting mixture is heated on a water bath (70-80°C) for 30 min and then cooled to room temperature and filtered through a sterilizing filter (0.22 μm) under aseptic conditions in a sterile vial.

In the above example, the entered quantity of ascorbic acid, Sn (II) and gelatin in terms of 1 ml of the prepared mixture, respectively, equal to 0.25; 0,0175 and 2.5 mg of the product with dimensions less than 100 nm was 83%, and particles less than 20 nm - 6%. The radiochemical purity of the drug is equal to 91% and remains at this level for 4 hours.

Example 3. The reaction mixture is prepared as in example 2, with the difference that after the introduction of a freshly prepared solution of Sn (II) the mixture was added 160 μl of an aqueous solution of gelatin with a concentration of 10 mg/ml of the resulting mixture is heated on a water bath (70-80°C) for 30 min and then cooled to room temperature and filtered through a sterilizing filter (0.22 μm) under aseptic conditions in a sterile vial.

In the above example, the entered quantity of ascorbic acid, Sn (II) and gelatin in terms of 1 ml of the prepared mixture, respectively, equal to 0.25; 0,0175 and 4.0 mg of the product with dimensions less than 100 nm was 84%, and particles less than 20 nm - 6%. The radiochemical purity of the drug is equal to 92% and remains at this level for 4 hours.

Presented p is Kerov should introduction in the composition of the reaction mixture of gelatin in the amount of 2.5 to 4.0 mg per 1 ml of the mixture enhances the yield of the target nanocolloid99mTc-Al2O3with particle sizes less than 100 nm up to 83-84% with radiochemical purity of the product is 92%.

In General, the proposed method allows to obtain technetium-99 m nanocolloidal the drug on the basis of gamma-alumina, suitable for carrying out gamma scintigraphic studies, as evidenced by lymphoscintigram in figure 1, obtained within 1 hour after administration of the radiopharmaceutical experimental animal (rat). Accumulation in the lymph node of 1.5%.

Table 1
RadiopharmaceuticalsManufacturerThe particle sizes
NanocollGE Amersham<80 nm
99mTc-nanocolloidGE, Uppsala, Sweden<80 nm
NanocisCIS bio International~100 nm
HepatocisCIS bio International~500 nm
MicroliteDu Font~10 nm
Sulfur colloidCIS bio International40-10000 nm

Table 2
The concentration of Sn (II), mg/ml0,008750,01750,0350,070,1050,14
The contents of Tc(VII), %10730,400

The method with technetium-99m of nanocolloid, characterized in that the aqueous suspension prepared from nanosized powder of gamma-aluminum oxide with a particle diameter of 7-10 nm and a concentration of 0.5-0.7 mg/ml, after adjusting pH to 4-5, enter the eluate of technetium-99m, then in quantities per 1 ml of the mixture sequentially injected ascorbic acid 0,20-0,25 mg tin (II) chloride dihydrate 0,00875-0,0175 mg and gelatin 2,5-4,0 mg, the resulting mixture is heated on a water bath at a temperature of 70-80°C for 30 min, cooled to room temperature and the temperature of the ultrasonic bath and spend sterilizing filtration.



 

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