CA1133812A - Composition for labeling of red blood cells with radioactive technetium - Google Patents
Composition for labeling of red blood cells with radioactive technetiumInfo
- Publication number
- CA1133812A CA1133812A CA340,133A CA340133A CA1133812A CA 1133812 A CA1133812 A CA 1133812A CA 340133 A CA340133 A CA 340133A CA 1133812 A CA1133812 A CA 1133812A
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- Prior art keywords
- labeling
- composition
- blood cells
- red blood
- radioactive
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
- A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
- A61K51/025—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus inorganic Tc complexes or compounds
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
- A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
- A61K51/04—Organic compounds
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2123/00—Preparations for testing in vivo
Abstract
COMPOSITION FOR LABELING OF RED BLOOD
CELLS WITH RADIOACTIVE TECHNETIUM
Abstract of the Disclosure The invention relates to a non-radioactive composition for intracorporeal labeling of red blood cells with radio-active technetium, The composition comprises (a) pyridoxal or a salt thereof, (b) at least one stannous salt, and (c) at least one .alpha.-amino acid as the essential components.
The compound can be administered in aqueous solution into a vein in a living body and assures an efficient intra-corporeal labeling of red blood cells in the living body with radioactive technetium subsequently administered to the vein. This enables technetium to be efficiently used as a labeling medium without the disadvantages of extra-corporeal labeling previously employed.
CELLS WITH RADIOACTIVE TECHNETIUM
Abstract of the Disclosure The invention relates to a non-radioactive composition for intracorporeal labeling of red blood cells with radio-active technetium, The composition comprises (a) pyridoxal or a salt thereof, (b) at least one stannous salt, and (c) at least one .alpha.-amino acid as the essential components.
The compound can be administered in aqueous solution into a vein in a living body and assures an efficient intra-corporeal labeling of red blood cells in the living body with radioactive technetium subsequently administered to the vein. This enables technetium to be efficiently used as a labeling medium without the disadvantages of extra-corporeal labeling previously employed.
Description
:~133812 The present invention relates to a non-radioactive composition useful for labeling red blood cells with radio-active technetium, and its preparation and use.
More particularly, the invention relates to a non-radioactive composition which is useful for the efficient attainment of intracorporeal labeling of red blood cells radioac~, v~
in living bodies with ratioJctivc technetium, and which has a high stability and no material toxicity, and its preparation and use.
Radioactive substances which are not taken into any organs when administered into b100d vessels and remain in the blood circulation system are useful for obtaining medically valuable informations, for instance, for extra-corporeal observation of blood pools in the heart and of cerebral and peripheral blood vessels and for determin-ation of the volume of the whole blood circulation system.
for development of such radioactive substances, a variety of investigations have been made in the field of nuclear medicine. l3lI-Labeled human serum albumin (13lI-HSA) is now widel~ used for such purposes, but this labeled agent is not satisfying because 13lI has a long half life ~i.e. about ~3 days) and emits B-rays which expose the patients to high levels of radiation. In addition, its y-ray energy spectrum is not suitable for widely employed y-cameras for low energy.
Since technetium-99m (99mTc3 emits only y-rays of about 140 KeY (not accompanied with B-rays) and has a short half life (i.e. about 6 hours3, it is quite suitable as a radioactive nuclide for diagnostic agents to be admini-30 stered to human bodies, and its utilization in the fieldof nuclear medicine has been rapidly increasing in recent years.
~13381Z
Attempts have been made to label the blood circulation system with 99mTc in place of l3lI-HSA, and the pro-duction of 99mTc-HSA and 99mTc-red blood cells by the extracorporea1 labeling method have been proposed. How-ever, 99mTc-HSA is inferior in labeling efficiency and stability in living bodies. 99mTc-red blood cells produced by the extracorporeal labeling method are also not satisfying because a complicated series of operations is required for their production: i.e collection of blood, separation of blood cells, add;tion of stannous compounds, addition of pertechnetate (99mTc), reac-t;on, isolation and purification, and administration.
In addition, there is a high possibility of bacterial contamination during these operations. Thus, the use of these labeling agents is not yet widely accepted.
On the other hand, intracorporeal labeling of red blood cells with 99m~c has been attempted by adm;n-istering into a blood vessel an injectable liquid composition containing a stannous compound in the form of a phosphate chelate and then administering a pertechnetate (99mTco4 ) so as to label red blood cells with 99mTc specifically. ~his procedure of labeling has been attracting much interest because of its simplicity and high labeling efficiency, the high stability after labeling, etc. Stannous pyrophosphate (Sn-PPi), stannous ethane-l-hydroxy-1,1-diphosphonate (Sn-EHD~), stannous methylene-diphosphonate (Sn-MDP), etc. have been suggested dS the stannous phosphate chelate to be used in this procedure. Of these, Sn-PPi is employed most frequently.
The main object of the present invention is to provide a novel composition for attaining efficient intracorporeal 99mTc-labeling of red blood cells with a high stability and no material toxicity. Another object of this inven-tion is to provide a process for industrial preparation of such a composition. A further object of the invention is to provide a method for intracorporeal 99mTc-labeling of red blood cell~s in living bodies which such a composition.
According to one aspect of the invention there is provided a non-radioactive composition for intracorporeal labeling of red blood cells with radioactive technetium, which comprises a lyophilized product containing la) pyridoxal and/or at least one salt thereof, (b1 at least one stannous s~lt, and (c) at least one a-amino acid as the essential components.
According to another aspect of the invention there is provided a kit for the preparation of a non-radioactive composition which comprises (a) pyridoxal and/or at least one salt thereof, Ib) at least one stannous salt, and (c) at least one ~-amino acid as the essential components;
which ~it comprises (A) an aqueous solution comprising at least one of the essential components (a), (~) and (c) or a lyophilized product therefrom, and ~B) an aqueous 501u-tion comprising the remaining of those essential components or a lyophilized product therefrom.
The pyridoxal salts may be salts formed with inorganic acids such as mono~asic inorganic acids ~e g. hydrocnloric acid, nitric acid) and poly~asic inorganic acids (e.g.
sulfuric acid, phosphoric acid) and witn organic acids such as acetic acid and oxa~ic acid. ~he term stannous salts means the salts of the divalent tin ion (Sn+ ), of which stannous halides (e.g. stannous chloride, stannous fluoride), stannous sulfate, stannous nitrate, stannous acetate, stannous citrate, stannous tartrate, etc. are examples. The ~-amino acid may be a natural or synthe-tic D-, L- or DL-amino acid having an amino group at the a-position to a carboxyl group. A natural L-amino acid is desirable for administration to human bodies.
In addition to the said essential components, additional materials may be incorporated into the composition, e.g.
a stabilizing agent or a compound having an oxidation-preventing activity, for example ascorbic acid. An isotonization agent, for example sodium chloride, and a preservative, for example, benzyl alcohol, may also be added without any disadvantage. The order of incor-poration of the essential and optional components has no influence upon the production of the composition of the invention, and the components may be mixed together in any optional order. The pH value of the composition is not particularly irnportant, and any optional pH value is suit-able for the purpose of use of the composition, but it is preferred to adjust the pH value within a range of 8 to 9, for instance, by the addition of sodium hydroxide or hydro-chloric acid.
The molar ratio of pyridoxal, or a salt thereof, and the ~-amino acid may be varied within the range of 0.3 to 3.0, A particularly desirable molar ratio is lØ The stannous salt may be used in an amount sufficient to reduce the whole pertechnetate administered into a blood vesse1. When the stability of the composition of the invention and the toxicity of the stannous salt are taken into consideration, the stannous salt may be used in an amount of 0.001 to 0.01 mol per mol of pyridoxa1 or its salt. The concentration of the pyridoxal or its salt may be not less than 0.5 mmol/liter and should be sufficiently dissolved to give a clear solution [e.g. not more than S00 mmol/liter). From the point of view of the amount of the stannous salt to be added and the stability of the composition of the invention, a concentration of 80 to 10 mmol/liter is preferable. Although the composition of the invention is usually administered into blood vessels of living bodies and therefore should be ulti~ately prepared in the form of an aqueous solution, it may be formulated in powder form as an intermediate stage for convenience during storage. ~or instance, the composition of the invention once prepared in an aqueous solution may be subjected to lyophilization, and the lyophilized product may be diluted with water to make an aqueous solution suitable for use. Alternatively, for instance, an aqueous solution comprising pyridoxal or its salt and a stannous salt or a lyophilized product therefrom and an aqueous solution comprising an -amino acid or a lyophilized product therefrom may be separately produced and stored as a kit. When required for use, these are combined together, if necessary with an additional amount of an aqueous medium, to make the composition of the invention in the form of an aqueous solution.
For intracorporeal labeling of red blood cells in living bodies with radioactive technetium, the composi-tion of the invention in aqueous solution form is usually first administered into a blood vessel, e.g. a vein, and then an aqueous solution comprising 99mTc in the form of pertechnetate is administered into the blood vessel.
The amount of the composition of the invention to be administered may be such that the stannous salt therein can sufficiently reduce 99mTc in the form of the per-technetate to be subsequently administered. When, forinstance, the administration is carried out intravenously, the composition of the invention may be employed in such an amount as to contain 10 to 20 ~9 of stannous ion per kg of bodyweight, which corresponds to a 2.0 to 4.0 ml injec-tion of 2.gS mM/liter stannous ion solution to a 70 kg man. In such a case, the concentration of 99mTc in the aqueous sslJtion is normally from about 0.1 to 50 mCi in about 0.5 to 5.0 ml at the time of administration. Need-less to say, the doses of the composition of the invention and of the 99mTc aqueous solution may be varied with the concentrations of the active ingredients, the ~odyweignts of the patients, etc. Although there is no special limitation regarding the time difference between the adm;nistrations of the composition of the invention and of the 99mTc aqueous solution, there is normally a period of about 10 to 60 minutes, preferably about 15 to 30 minutes, hetween them.
The composition of the invention is quite suitable for intracorporeal 99mTc-labeling of red blood cells because of the following advantageous characteristics:
1) The composition is stable for a long period of time after production;
More particularly, the invention relates to a non-radioactive composition which is useful for the efficient attainment of intracorporeal labeling of red blood cells radioac~, v~
in living bodies with ratioJctivc technetium, and which has a high stability and no material toxicity, and its preparation and use.
Radioactive substances which are not taken into any organs when administered into b100d vessels and remain in the blood circulation system are useful for obtaining medically valuable informations, for instance, for extra-corporeal observation of blood pools in the heart and of cerebral and peripheral blood vessels and for determin-ation of the volume of the whole blood circulation system.
for development of such radioactive substances, a variety of investigations have been made in the field of nuclear medicine. l3lI-Labeled human serum albumin (13lI-HSA) is now widel~ used for such purposes, but this labeled agent is not satisfying because 13lI has a long half life ~i.e. about ~3 days) and emits B-rays which expose the patients to high levels of radiation. In addition, its y-ray energy spectrum is not suitable for widely employed y-cameras for low energy.
Since technetium-99m (99mTc3 emits only y-rays of about 140 KeY (not accompanied with B-rays) and has a short half life (i.e. about 6 hours3, it is quite suitable as a radioactive nuclide for diagnostic agents to be admini-30 stered to human bodies, and its utilization in the fieldof nuclear medicine has been rapidly increasing in recent years.
~13381Z
Attempts have been made to label the blood circulation system with 99mTc in place of l3lI-HSA, and the pro-duction of 99mTc-HSA and 99mTc-red blood cells by the extracorporea1 labeling method have been proposed. How-ever, 99mTc-HSA is inferior in labeling efficiency and stability in living bodies. 99mTc-red blood cells produced by the extracorporeal labeling method are also not satisfying because a complicated series of operations is required for their production: i.e collection of blood, separation of blood cells, add;tion of stannous compounds, addition of pertechnetate (99mTc), reac-t;on, isolation and purification, and administration.
In addition, there is a high possibility of bacterial contamination during these operations. Thus, the use of these labeling agents is not yet widely accepted.
On the other hand, intracorporeal labeling of red blood cells with 99m~c has been attempted by adm;n-istering into a blood vessel an injectable liquid composition containing a stannous compound in the form of a phosphate chelate and then administering a pertechnetate (99mTco4 ) so as to label red blood cells with 99mTc specifically. ~his procedure of labeling has been attracting much interest because of its simplicity and high labeling efficiency, the high stability after labeling, etc. Stannous pyrophosphate (Sn-PPi), stannous ethane-l-hydroxy-1,1-diphosphonate (Sn-EHD~), stannous methylene-diphosphonate (Sn-MDP), etc. have been suggested dS the stannous phosphate chelate to be used in this procedure. Of these, Sn-PPi is employed most frequently.
The main object of the present invention is to provide a novel composition for attaining efficient intracorporeal 99mTc-labeling of red blood cells with a high stability and no material toxicity. Another object of this inven-tion is to provide a process for industrial preparation of such a composition. A further object of the invention is to provide a method for intracorporeal 99mTc-labeling of red blood cell~s in living bodies which such a composition.
According to one aspect of the invention there is provided a non-radioactive composition for intracorporeal labeling of red blood cells with radioactive technetium, which comprises a lyophilized product containing la) pyridoxal and/or at least one salt thereof, (b1 at least one stannous s~lt, and (c) at least one a-amino acid as the essential components.
According to another aspect of the invention there is provided a kit for the preparation of a non-radioactive composition which comprises (a) pyridoxal and/or at least one salt thereof, Ib) at least one stannous salt, and (c) at least one ~-amino acid as the essential components;
which ~it comprises (A) an aqueous solution comprising at least one of the essential components (a), (~) and (c) or a lyophilized product therefrom, and ~B) an aqueous 501u-tion comprising the remaining of those essential components or a lyophilized product therefrom.
The pyridoxal salts may be salts formed with inorganic acids such as mono~asic inorganic acids ~e g. hydrocnloric acid, nitric acid) and poly~asic inorganic acids (e.g.
sulfuric acid, phosphoric acid) and witn organic acids such as acetic acid and oxa~ic acid. ~he term stannous salts means the salts of the divalent tin ion (Sn+ ), of which stannous halides (e.g. stannous chloride, stannous fluoride), stannous sulfate, stannous nitrate, stannous acetate, stannous citrate, stannous tartrate, etc. are examples. The ~-amino acid may be a natural or synthe-tic D-, L- or DL-amino acid having an amino group at the a-position to a carboxyl group. A natural L-amino acid is desirable for administration to human bodies.
In addition to the said essential components, additional materials may be incorporated into the composition, e.g.
a stabilizing agent or a compound having an oxidation-preventing activity, for example ascorbic acid. An isotonization agent, for example sodium chloride, and a preservative, for example, benzyl alcohol, may also be added without any disadvantage. The order of incor-poration of the essential and optional components has no influence upon the production of the composition of the invention, and the components may be mixed together in any optional order. The pH value of the composition is not particularly irnportant, and any optional pH value is suit-able for the purpose of use of the composition, but it is preferred to adjust the pH value within a range of 8 to 9, for instance, by the addition of sodium hydroxide or hydro-chloric acid.
The molar ratio of pyridoxal, or a salt thereof, and the ~-amino acid may be varied within the range of 0.3 to 3.0, A particularly desirable molar ratio is lØ The stannous salt may be used in an amount sufficient to reduce the whole pertechnetate administered into a blood vesse1. When the stability of the composition of the invention and the toxicity of the stannous salt are taken into consideration, the stannous salt may be used in an amount of 0.001 to 0.01 mol per mol of pyridoxa1 or its salt. The concentration of the pyridoxal or its salt may be not less than 0.5 mmol/liter and should be sufficiently dissolved to give a clear solution [e.g. not more than S00 mmol/liter). From the point of view of the amount of the stannous salt to be added and the stability of the composition of the invention, a concentration of 80 to 10 mmol/liter is preferable. Although the composition of the invention is usually administered into blood vessels of living bodies and therefore should be ulti~ately prepared in the form of an aqueous solution, it may be formulated in powder form as an intermediate stage for convenience during storage. ~or instance, the composition of the invention once prepared in an aqueous solution may be subjected to lyophilization, and the lyophilized product may be diluted with water to make an aqueous solution suitable for use. Alternatively, for instance, an aqueous solution comprising pyridoxal or its salt and a stannous salt or a lyophilized product therefrom and an aqueous solution comprising an -amino acid or a lyophilized product therefrom may be separately produced and stored as a kit. When required for use, these are combined together, if necessary with an additional amount of an aqueous medium, to make the composition of the invention in the form of an aqueous solution.
For intracorporeal labeling of red blood cells in living bodies with radioactive technetium, the composi-tion of the invention in aqueous solution form is usually first administered into a blood vessel, e.g. a vein, and then an aqueous solution comprising 99mTc in the form of pertechnetate is administered into the blood vessel.
The amount of the composition of the invention to be administered may be such that the stannous salt therein can sufficiently reduce 99mTc in the form of the per-technetate to be subsequently administered. When, forinstance, the administration is carried out intravenously, the composition of the invention may be employed in such an amount as to contain 10 to 20 ~9 of stannous ion per kg of bodyweight, which corresponds to a 2.0 to 4.0 ml injec-tion of 2.gS mM/liter stannous ion solution to a 70 kg man. In such a case, the concentration of 99mTc in the aqueous sslJtion is normally from about 0.1 to 50 mCi in about 0.5 to 5.0 ml at the time of administration. Need-less to say, the doses of the composition of the invention and of the 99mTc aqueous solution may be varied with the concentrations of the active ingredients, the ~odyweignts of the patients, etc. Although there is no special limitation regarding the time difference between the adm;nistrations of the composition of the invention and of the 99mTc aqueous solution, there is normally a period of about 10 to 60 minutes, preferably about 15 to 30 minutes, hetween them.
The composition of the invention is quite suitable for intracorporeal 99mTc-labeling of red blood cells because of the following advantageous characteristics:
1) The composition is stable for a long period of time after production;
2) Labeling of red blood cells with 99mTc can be attained in a high yield by a simple operation comprising administration of the composition into a blood vessel and subsequent administration of an aqueous solution of 99mTc in the form of pertechnetate into the blood vessel;
3) The 99mTc-labe1ed red blood cells thus obtained are sufficiently stable in the living body; and
4) The toxicity is extremely low.
The present invention will be explained further in detail by way of the following Examples.
E_ample 1 Preparation of d composition for labeling containing L-isoleucine as the -amino acid and ascorbic acid as the stabilizing agent (this composition being hereinafter referred to as "Sn-P.Ile.A."):-Nitrogen gas, sterilized by passing it through a filter having a pore size of 0.22 ~m, was introduced into pyrogen-free sterile water to eliminate dissolved oxygen therefrom. Pyridoxal hydrochloride (3665 mg, 18 mmol~, anhydrous stannous chloride (37.9 mg, 0.2 mmol), and L~ ascorbic acid as the stabilizer (70 mg, 0.4 mmol) were dissolved in the thus treated water (100 ml) under sterile conditions in a nitrogen atmosphere to form a solution (hereinafter referred to dS "solution A").
L-lsoleucine (2361 mg, 18 mmol) and sodium hydroxide (1440 mg, 36 mmol) were separately dissolved into the said sterile, pyrogen-free and oxygen-free water (100 ml) under sterile conditions in a nitrogen atmosphere to form a solution (hereinafter referred to as "solution B"). The solutions A and ~ were mixed together in a nitrogen atmosphere to form the desired composition for labeling, Sn-P.lle.A., which was used to fill vials (each 2.0 ml) having rubber caps, through a filter of 0.22 ~m in pore size in a nitrogen atmosphere. The thus obtained Sn-P.lle.A. was a clear yellow solution having a pH value of 8 to 9.
Example 2 _ _ _ _ _ Preparation of compositions for labeling containing -amino acids other than L-isoleucine:-In the same manner as in Example 1 but using L-leucine, L-valine, L-phenylalanine, L-alanine, L-glycine, and mono-sodium L-glutamate in place of L-isoleucine in the same molar amount (18 mmol), compositions for labeling (i.e., Sn-P.Leu.A., Sn-P.Yal.A., Sn-P.Phe.A., Sn-P.Ala.A., Sn-P.Gly.A. and Sn-P.~lu.A.) were prepared. These compositions ~ere each a clear yellow solution having a pH value of 8 to 9.
Example 3 Lyophilization of compositions for la~eling obtained in Examples 1 and 2:-The Sn-P.Ile.A. solution obtained in Example 1 was 11338~2 charged in portions (each 2.0 ml) into vials for lyo-philization and subjected to lyophilization by means of a lyophilizer. After the lyophilization, the pressure was elevated to atmospheric pressure by the introduction of nitrogen, and the vials were sealed with a rubber cap in a nitrogen atmosphere.
The compositions for labeling obtained in Example 2 were also treated in the same manner as above for lyophilization ~xample 4 _, , Production of a kit for the preparation of a composition for labeling (Sn-P.lle,A.):-The solution A obtained in Example 1 was charged inportions (each 1.~ ml) into vials for lyophilization through a filter of 0.22 ~m pore size in a nitrogen atmosphere and subjected to lyophilization by means of a lyophilizer. After the lyophilization, the pressure was elevated to atmospheric pressure by the introduction of nitogen, and the vials were sealed with a rubber cap in a nitrogen atmosphere to obtain a preparation (hereinafter referred to as "Sn-P.A."). Separately, the solution B
obtained in Example 1 WdS diluted two fold with sterile, pyrogen-free and oxygen-free water ~10~ ml) with suf-ficient stirring. The diluent was filled in portions (each 2.5 ml) into ampoules through a filter of 0.22 ~m pore size in a nitrogen atmosphere and melt-sealed in a nitrogen atmosphere to obtain another preparation (here-inafter referred to as "Ile."). During use, ~le. ~2.~ ml) was added to Sn-P.A. to obtain Sn-P.Ile.A.
Ex_mple 5 Production of kits for the preparation cf compositions 1~33812 for labeling:-ln the production of the solution B describedin Example lt L-leucine, L-valine, L-phenylalanine, L-alanine, L-glycine and L-monosodium glutamate were used in place of L-isoleucine in the same molar amount to obtain preparations Leu., Val., Phe., Ala., Gly. and Glu.
During use, each of these preparations (2.0 ml) was added to the Sn-P.A. obtained in Example 4 to form Sn-P.Leu.A., Sn-P.Val.A., Sn-P.Phe.A., Sn-P.Ala,A., Sn-P.Gly.A. and Sn-P.Glu.A.
Example 6 _ _ _ Intracorporeal 99m~c-labeling of red blood cells with compositions for labeling:-Various labeling compositions of the invention wereinjected into the tail veins of female rats of the Sprague-Dawley strain (body weight, 145 - 175 9). The dose of the composition was adjusted so that the amount of Sn(ll) per kg of body weight was 20 ug (as mentioned below). After a suitable time (30 minutes), normal saline solutions (0.2 ml) containing 99mTc in the form of sodium pertechnetate (about 0.5 mCi) were injected into the opposite tail veins. ~ne hour after the administra-tion of the 99mTc, the rats were subjected to abdominal incision, and blood (5 - 7 ml) was collected from the aorta with a heparinized syringe. A part of the collected blood (1.~ ml) was put into an ampoule with a pipet for radioactivity determination and counted on a gamma coun-ter. 8y comparing the determined ~alue thus obtained with that of a separately prepared standard sample, the percen-tage of 99mTc present in 1 ml of blood (after correctionfor decay) to the whole injected dose was calculated.
This value was indicated as %ID/ml 81cod (ID = injected ~e). For comparison of the data obtained in animals having different body weights, this value was then nor-malized to an average body weight of 160 g according to the following equation:
~ID/ml Blood (norm) = [% ID/ml Blood] x 160 W: body weight of rat (g) The remaining blood (3 to 5 ml) was centrifuged (gO0 g, 20 min) so as to be separated into plasma and blood cells. After centrifuging, the hematocrit value (volume percent of blood cells in blood, % Hct) was obtained. A part of the separated plasma (1.0 ml) was placed into an ampoule with a pipet for radioactivity determination. The plasma (1.0 ml) and the above mentioned blood (1.0 ml) were counted in the same geometrical position (the same count efficiency position) in the gamma counter, and ~rom the determined values, the background values were subtracted to obtain net count val~Jes per min., C(plasma) and C(blood). From these values, the percentages of radioactivity distributed in the red blood cells and in the plasma (~/R~C and ~/Plasma), respectively were calculated according to the following equations.
C(Plasma) x ~1 _ lo~t) %/Plasma = C(Blood) x 1~0 ~/~BC = 100 - (%/Plasma) RBC: red blood cells Then, the percentage of radioactivity present in 1 ml of red blood cells to the whole dose of radioactivity i133812 [%/ml RBC (norm)] was calculated according to the following equation:
%ID/ml RBC ~norm) = [%/ml Blood (norm)] x ~i7'-ct) Finally, the percentage of radioactivity present in the whole red blood cells to the whole dose of radio-activity (%ID in whole RBC) was calculated according to the following equation, on the assumption that the whole amount of blood (ml) was 6.5 % of the body weight (g):
~I~ in whole RBC
1~ = 1%ID/ml Blood) x W x lo5 x (1/0 C) = ~%ID/ml Blood (norm)] x 160 x 1OO x (l/oB ) Using the experimental procedure described above, the following evaluations were performed on the compositions of the invention:
(1) The results of intracorporeal labeling of red blood cells with seven of the compositions for label ing of the invention are shown in Table 1.
Table 1 Intracorporeal 99mTc-labeling of red blood cells with compositions for labeling:-romposition ¦ ~ID/ml ~/RBC j~/Plasma ¦ ~ID/ml ¦ ~ID in for labell.ng, Blood RBC ¦ whole ¦ (norm) (norm) I RBC
. .~ . __ .. _ Sn-P.Ile.A. 9.58 98.95 1.05 20.77 98.52 - ~ __ _ Sn-P.Val.A. 9.04 99.04 0.96 19.62 93.05 __ ...... . _ .. _ _ . _ Sn-P.Leu.A. ¦ 8.79 98.98 ¦ 1.02 19.09 90.51 __ I __, Sn-P.Phe.A. ¦ 8.75 98.90 ¦ 1.10 18.98 90.01 Sn-P.Ala,AO ¦ 9.51 ~99 ! 1 l0 L20 64 97~94 Sn-P.Gly.A. ¦ 9.51 ¦98.71 ¦ 1.29 ¦ 20.60 97.71 _, I
Sn-P.Glu.A.9.34 ¦ 98.75 1.25¦ 20.23 96.00 ¦Sn-PPi ) 8.38 1 98.70 1.30¦ 18.17 86.18 Note: Each numeral indicates an average value for five rats.
For comparison, the results obtained in the already known Sn-PPi ~*) used under the same conditions are also shown in Table 1. In this experiment, each of the compositions for labeling was administered in an amount of 20 ~g as Sn(ll) ion per kg of body weight of rat, then 99mTc04 was administered after 30 minutes and the blood was collected 1 hour thereafter, With the amount of Sn(II) of 20 ug/kg, as mentioned below, the efficiency of labeling of red blood cells reached the maximum with each of the compositions for labeling, As can be understood from Table 1, all of the compositions for labeling of the invention showed a labeling efficiency of 90 ~ or more in the ~ ID in whole RBC.
From these results, it will be apparent to those skilled in the art th~t the series of the compositions of the invention exhibits an extremely good effect in intracorporeal labeling of red blood cells with 99mTc.
~2) The relationship between the dose of Sn-Plle.A. as a typical representative of the composition for labeTing of 11338~Z
the invention and the efficiency of intracorporeal labeling of red blood cells is shown in Table 2.
Table 2 Intracorporeal 99mTc-labeling red blood cells with Sn-P.Ile.A.:-Amount of ¦ %ID/ml %/RBC %/Plasma ~ID/ml %ID in Sn~II) Blood RBCwhole RBC
(~g/kg body (norm) (norm) weight) _ 1 1.12 40.1659.04 0.984.64 6.26 98.381.62 13.5064.03 9.52 99.100.90 20.6898.08 9.58 98.951.05 20.7798.52 8.56 98.611.39 18.4987.71 Note: Each numeral indicates an average valuefor five rats.
As is apparent from this Tablel administration of 10 to 20 ~glkg of Sn(ll) in the form of Sn-P.lle.A. attains a high yield of red blood cell labeling. Similar results were also obtained on the compositions for labeling other than Sn-P.lle.A. shown in Example ~.
It was shown by these experiments that intracorporeal labeling of red blood cells with 99mTc could be realized with extreme simplicity by the use of the composition for labeling of the invention.
Example 7 __ _ Stability of compositions for labeling:-(1) Stability in a solution form:
The Sn-P.Ile.A. produced as in Example 1 was stored at 4 to 8 C for 50 or }00 days. 8y the use of the resulting preparation, the intracorporeal labeling of red blood cells with 99mTc was effected as in Example 6. The results are shown in Table 3. As can be seen from this ~1338~2 Table, Sn-P-lle.A. in solution form is extremely stable, not showing any signs o~ deterioration during a period immediately after its production to the 100th day.
It was also confirmed that ~any kinds of compositions for labeling of the invention including the other six compositions shown in Example 2 showed a similar stability in the sollJtion form.
Table 3 Stability of ~n-P.Ile.A. in a solution form (stored at 4 to 8C):-After production 50 days ¦ 100 days %ID/ml Blood (norm) 9.57 9.52 ~/~BC 98.26 98.32 ~/Plasma 1.74 1.68 ... .. __ I "
~ID/ml RBC (norm) 20.21 20.11 %ID in whole RBC 98.22 98.16 Note: Average values for five rats Experimental procedure: administration of Sn-P.Ile.A. in an amount of 20 ~g as Sn(II) per kg of thebody weight of a rat, further administration of 9gmTc04 after 30 minutes and collection of blood 1 hour thereafter.
(2) Stability in a lyophilized state:
The lyophilized product of Sn-P.Ile.A. produced as in Example 3 was stored at 4 to 8C for 100 or 200 days.
Using the resultant product, the intracorporeal labeling of red blood cells with 99mTc was effected as in Example 6.
The resul-ts are shown in Table 4.
Table 4 3~1 Stability of Sn-PIle.A. in a lyophilized state ~stored at 4 to 8C):-¦ After production ¦100 days 200 days ,._ . . . . _ _ ~ID/ml Blood (norm~ 9.63 9.52 ._ . . ._ %/RBC _ _ !98-42_ 98.31 ~/Plasma 1.58 1.69 ._.
%ID/ml RBC (norm) 20.24 20.21 ___ ~ID in whole RBC ! 98.31 98.35 Note: Average values for five rats.
Experimental procedure: administration of Sn-10 P.Ile.A. in an amount of 20 ~g as Sn~II) per kg of the body weight of a rat, further administration of 99mTcO4 after 30 minutes and collection of blood 1 hour thereafter.
The lyophilized product was subjected to the ex-periment in the form of a solution which was prepared by adding distilled water for injection (2.0 ml) immediately before use. As can be seen from Table 4, the Sn-P.Ile.A.
in the lyophilized state is extremely stable, not showing any sign of deterioration during the period immediately after its preparation to the 200th day.
It was also confirmed that many kinds of compositions for labeling of the invention including the other six compositions shown in Example 3 showed a similar stability in the lyophilized state.
(3) Stability of a kit for preparation of a composition for labeling:
A kit for the preparation of d composition for labeling comprising Sn-P.A. and Ile. which was produced as in Example 4 was stored at 4 to 8 C for 100 or 200 days. Immediately before use, Ile (2.0 ml) ~Jas added to 30 Sn-P.A. to obtain Sn-P.Ile.A., by the use of which the 1133~31Z
intracorporeal labeling of red blood cells with 99mTc was effected as in Example 6. The results are shown in Tdble 5. As is apparent from this Table, the kit for the preparation of a composition for labeling comprising Sn-P.A. and 1le. was extremely stable, not showing any signs of deterioration during the period from its pro-duction time to the 200th day.
Table 5 Stability of a kit for preparation of a composition for labeling comprising Sn-P.A. and Ile. (stored at 4 to g C ) :--. . ._ .
After production 100 days ¦ 20Q days . .
%ID/ml Blood (norm) 9.55 9.49 ~/R~C g8.43 19g.66 ~/Plasma 1.57 1.34 _ __ %ID/ml RBC ~norm) 20.41 20.52 ._.
%ID in whole RBC 99.22 99.24 Note: Average values for five rats.
Experimental procedure: administration of Sn-P.Ile.A. in an amount of 20 ~g as Sn(II) per kg of the body ~eight of a rat, further administration of 99mTcO4 after 30 minutes and collection of blood 1 hour thereafter.
It was also confirmed that many kinds of ~its - shown in Example 5 sho~ed the same stabili~y.
Example 8 Toxicity of compositions for labeling of the invention:-In the same manner as in Example 1 or 2 but using the materials (other than water) in 5 times the amount,seven kinds of compositions for labeling were produced having 5 fold concentrations in comparison with the con~entrations of the compositions for labeling obtained in Examp1e 1 ~r 2. Each of the resulting so1utions was intravenously administered to groups consisting of lO
~prague Dawley strain ma1e rats, of lO S-D strain female rats, o~ lO ICR strain male mice and of lO ICR strain female mice at a dose of 0.5 ml per lO0 g of body weight (corresponding to an amount 1000 fold the intended dose to human beings) as well as to groups consisting of lO0 Hartley strain male guinea pigs, of lO Hartley strain female guinea pigs, of 10 male rabbits and of lO female rabbits at a dose of 0.25 ml per lO0 g Ot body w~ight (corresponding to an amount of abo~t 500 fold the inten-ded dose to human beings (per ~nit body weight)). As a control, the same vo1ume of a norm31 sa1ine so1ution was ;ntravenously administered t~ the separate groups of the same kinds of animals as above. ~hese animals were fed and observed for lO days, and the changes of the body weight wer~ recorded every d~y.
As the result of this toxicity test, no signifi-cant difference in the appearance of the animals and the change of the body weiyht was recognized between the yroups treated with the composition for labeling and the control groups. After 10 days of feeding and observation, all the animals were subjected to dissection, and observation of various organs was effected to examine histological abnormal-ities, whereby no abnormality was seen in any of the animals.
0 As shown from these results of the toxicity test, 1~3381Z
the composition for labeling of the invention produces no abnormality, even when administered to four kinds of animals (both male and female) in an amount of 500 to 1000 fold the designed dose to human beings per unit body ~eight. Thus, its toxicity is proven to be extremely low.
It should be understood by those skilled in the art that the above mentioned Examples are intended only to illustrate and explain the invention in detail and not to limit the scope of the invention.
The present invention will be explained further in detail by way of the following Examples.
E_ample 1 Preparation of d composition for labeling containing L-isoleucine as the -amino acid and ascorbic acid as the stabilizing agent (this composition being hereinafter referred to as "Sn-P.Ile.A."):-Nitrogen gas, sterilized by passing it through a filter having a pore size of 0.22 ~m, was introduced into pyrogen-free sterile water to eliminate dissolved oxygen therefrom. Pyridoxal hydrochloride (3665 mg, 18 mmol~, anhydrous stannous chloride (37.9 mg, 0.2 mmol), and L~ ascorbic acid as the stabilizer (70 mg, 0.4 mmol) were dissolved in the thus treated water (100 ml) under sterile conditions in a nitrogen atmosphere to form a solution (hereinafter referred to dS "solution A").
L-lsoleucine (2361 mg, 18 mmol) and sodium hydroxide (1440 mg, 36 mmol) were separately dissolved into the said sterile, pyrogen-free and oxygen-free water (100 ml) under sterile conditions in a nitrogen atmosphere to form a solution (hereinafter referred to as "solution B"). The solutions A and ~ were mixed together in a nitrogen atmosphere to form the desired composition for labeling, Sn-P.lle.A., which was used to fill vials (each 2.0 ml) having rubber caps, through a filter of 0.22 ~m in pore size in a nitrogen atmosphere. The thus obtained Sn-P.lle.A. was a clear yellow solution having a pH value of 8 to 9.
Example 2 _ _ _ _ _ Preparation of compositions for labeling containing -amino acids other than L-isoleucine:-In the same manner as in Example 1 but using L-leucine, L-valine, L-phenylalanine, L-alanine, L-glycine, and mono-sodium L-glutamate in place of L-isoleucine in the same molar amount (18 mmol), compositions for labeling (i.e., Sn-P.Leu.A., Sn-P.Yal.A., Sn-P.Phe.A., Sn-P.Ala.A., Sn-P.Gly.A. and Sn-P.~lu.A.) were prepared. These compositions ~ere each a clear yellow solution having a pH value of 8 to 9.
Example 3 Lyophilization of compositions for la~eling obtained in Examples 1 and 2:-The Sn-P.Ile.A. solution obtained in Example 1 was 11338~2 charged in portions (each 2.0 ml) into vials for lyo-philization and subjected to lyophilization by means of a lyophilizer. After the lyophilization, the pressure was elevated to atmospheric pressure by the introduction of nitrogen, and the vials were sealed with a rubber cap in a nitrogen atmosphere.
The compositions for labeling obtained in Example 2 were also treated in the same manner as above for lyophilization ~xample 4 _, , Production of a kit for the preparation of a composition for labeling (Sn-P.lle,A.):-The solution A obtained in Example 1 was charged inportions (each 1.~ ml) into vials for lyophilization through a filter of 0.22 ~m pore size in a nitrogen atmosphere and subjected to lyophilization by means of a lyophilizer. After the lyophilization, the pressure was elevated to atmospheric pressure by the introduction of nitogen, and the vials were sealed with a rubber cap in a nitrogen atmosphere to obtain a preparation (hereinafter referred to as "Sn-P.A."). Separately, the solution B
obtained in Example 1 WdS diluted two fold with sterile, pyrogen-free and oxygen-free water ~10~ ml) with suf-ficient stirring. The diluent was filled in portions (each 2.5 ml) into ampoules through a filter of 0.22 ~m pore size in a nitrogen atmosphere and melt-sealed in a nitrogen atmosphere to obtain another preparation (here-inafter referred to as "Ile."). During use, ~le. ~2.~ ml) was added to Sn-P.A. to obtain Sn-P.Ile.A.
Ex_mple 5 Production of kits for the preparation cf compositions 1~33812 for labeling:-ln the production of the solution B describedin Example lt L-leucine, L-valine, L-phenylalanine, L-alanine, L-glycine and L-monosodium glutamate were used in place of L-isoleucine in the same molar amount to obtain preparations Leu., Val., Phe., Ala., Gly. and Glu.
During use, each of these preparations (2.0 ml) was added to the Sn-P.A. obtained in Example 4 to form Sn-P.Leu.A., Sn-P.Val.A., Sn-P.Phe.A., Sn-P.Ala,A., Sn-P.Gly.A. and Sn-P.Glu.A.
Example 6 _ _ _ Intracorporeal 99m~c-labeling of red blood cells with compositions for labeling:-Various labeling compositions of the invention wereinjected into the tail veins of female rats of the Sprague-Dawley strain (body weight, 145 - 175 9). The dose of the composition was adjusted so that the amount of Sn(ll) per kg of body weight was 20 ug (as mentioned below). After a suitable time (30 minutes), normal saline solutions (0.2 ml) containing 99mTc in the form of sodium pertechnetate (about 0.5 mCi) were injected into the opposite tail veins. ~ne hour after the administra-tion of the 99mTc, the rats were subjected to abdominal incision, and blood (5 - 7 ml) was collected from the aorta with a heparinized syringe. A part of the collected blood (1.~ ml) was put into an ampoule with a pipet for radioactivity determination and counted on a gamma coun-ter. 8y comparing the determined ~alue thus obtained with that of a separately prepared standard sample, the percen-tage of 99mTc present in 1 ml of blood (after correctionfor decay) to the whole injected dose was calculated.
This value was indicated as %ID/ml 81cod (ID = injected ~e). For comparison of the data obtained in animals having different body weights, this value was then nor-malized to an average body weight of 160 g according to the following equation:
~ID/ml Blood (norm) = [% ID/ml Blood] x 160 W: body weight of rat (g) The remaining blood (3 to 5 ml) was centrifuged (gO0 g, 20 min) so as to be separated into plasma and blood cells. After centrifuging, the hematocrit value (volume percent of blood cells in blood, % Hct) was obtained. A part of the separated plasma (1.0 ml) was placed into an ampoule with a pipet for radioactivity determination. The plasma (1.0 ml) and the above mentioned blood (1.0 ml) were counted in the same geometrical position (the same count efficiency position) in the gamma counter, and ~rom the determined values, the background values were subtracted to obtain net count val~Jes per min., C(plasma) and C(blood). From these values, the percentages of radioactivity distributed in the red blood cells and in the plasma (~/R~C and ~/Plasma), respectively were calculated according to the following equations.
C(Plasma) x ~1 _ lo~t) %/Plasma = C(Blood) x 1~0 ~/~BC = 100 - (%/Plasma) RBC: red blood cells Then, the percentage of radioactivity present in 1 ml of red blood cells to the whole dose of radioactivity i133812 [%/ml RBC (norm)] was calculated according to the following equation:
%ID/ml RBC ~norm) = [%/ml Blood (norm)] x ~i7'-ct) Finally, the percentage of radioactivity present in the whole red blood cells to the whole dose of radio-activity (%ID in whole RBC) was calculated according to the following equation, on the assumption that the whole amount of blood (ml) was 6.5 % of the body weight (g):
~I~ in whole RBC
1~ = 1%ID/ml Blood) x W x lo5 x (1/0 C) = ~%ID/ml Blood (norm)] x 160 x 1OO x (l/oB ) Using the experimental procedure described above, the following evaluations were performed on the compositions of the invention:
(1) The results of intracorporeal labeling of red blood cells with seven of the compositions for label ing of the invention are shown in Table 1.
Table 1 Intracorporeal 99mTc-labeling of red blood cells with compositions for labeling:-romposition ¦ ~ID/ml ~/RBC j~/Plasma ¦ ~ID/ml ¦ ~ID in for labell.ng, Blood RBC ¦ whole ¦ (norm) (norm) I RBC
. .~ . __ .. _ Sn-P.Ile.A. 9.58 98.95 1.05 20.77 98.52 - ~ __ _ Sn-P.Val.A. 9.04 99.04 0.96 19.62 93.05 __ ...... . _ .. _ _ . _ Sn-P.Leu.A. ¦ 8.79 98.98 ¦ 1.02 19.09 90.51 __ I __, Sn-P.Phe.A. ¦ 8.75 98.90 ¦ 1.10 18.98 90.01 Sn-P.Ala,AO ¦ 9.51 ~99 ! 1 l0 L20 64 97~94 Sn-P.Gly.A. ¦ 9.51 ¦98.71 ¦ 1.29 ¦ 20.60 97.71 _, I
Sn-P.Glu.A.9.34 ¦ 98.75 1.25¦ 20.23 96.00 ¦Sn-PPi ) 8.38 1 98.70 1.30¦ 18.17 86.18 Note: Each numeral indicates an average value for five rats.
For comparison, the results obtained in the already known Sn-PPi ~*) used under the same conditions are also shown in Table 1. In this experiment, each of the compositions for labeling was administered in an amount of 20 ~g as Sn(ll) ion per kg of body weight of rat, then 99mTc04 was administered after 30 minutes and the blood was collected 1 hour thereafter, With the amount of Sn(II) of 20 ug/kg, as mentioned below, the efficiency of labeling of red blood cells reached the maximum with each of the compositions for labeling, As can be understood from Table 1, all of the compositions for labeling of the invention showed a labeling efficiency of 90 ~ or more in the ~ ID in whole RBC.
From these results, it will be apparent to those skilled in the art th~t the series of the compositions of the invention exhibits an extremely good effect in intracorporeal labeling of red blood cells with 99mTc.
~2) The relationship between the dose of Sn-Plle.A. as a typical representative of the composition for labeTing of 11338~Z
the invention and the efficiency of intracorporeal labeling of red blood cells is shown in Table 2.
Table 2 Intracorporeal 99mTc-labeling red blood cells with Sn-P.Ile.A.:-Amount of ¦ %ID/ml %/RBC %/Plasma ~ID/ml %ID in Sn~II) Blood RBCwhole RBC
(~g/kg body (norm) (norm) weight) _ 1 1.12 40.1659.04 0.984.64 6.26 98.381.62 13.5064.03 9.52 99.100.90 20.6898.08 9.58 98.951.05 20.7798.52 8.56 98.611.39 18.4987.71 Note: Each numeral indicates an average valuefor five rats.
As is apparent from this Tablel administration of 10 to 20 ~glkg of Sn(ll) in the form of Sn-P.lle.A. attains a high yield of red blood cell labeling. Similar results were also obtained on the compositions for labeling other than Sn-P.lle.A. shown in Example ~.
It was shown by these experiments that intracorporeal labeling of red blood cells with 99mTc could be realized with extreme simplicity by the use of the composition for labeling of the invention.
Example 7 __ _ Stability of compositions for labeling:-(1) Stability in a solution form:
The Sn-P.Ile.A. produced as in Example 1 was stored at 4 to 8 C for 50 or }00 days. 8y the use of the resulting preparation, the intracorporeal labeling of red blood cells with 99mTc was effected as in Example 6. The results are shown in Table 3. As can be seen from this ~1338~2 Table, Sn-P-lle.A. in solution form is extremely stable, not showing any signs o~ deterioration during a period immediately after its production to the 100th day.
It was also confirmed that ~any kinds of compositions for labeling of the invention including the other six compositions shown in Example 2 showed a similar stability in the sollJtion form.
Table 3 Stability of ~n-P.Ile.A. in a solution form (stored at 4 to 8C):-After production 50 days ¦ 100 days %ID/ml Blood (norm) 9.57 9.52 ~/~BC 98.26 98.32 ~/Plasma 1.74 1.68 ... .. __ I "
~ID/ml RBC (norm) 20.21 20.11 %ID in whole RBC 98.22 98.16 Note: Average values for five rats Experimental procedure: administration of Sn-P.Ile.A. in an amount of 20 ~g as Sn(II) per kg of thebody weight of a rat, further administration of 9gmTc04 after 30 minutes and collection of blood 1 hour thereafter.
(2) Stability in a lyophilized state:
The lyophilized product of Sn-P.Ile.A. produced as in Example 3 was stored at 4 to 8C for 100 or 200 days.
Using the resultant product, the intracorporeal labeling of red blood cells with 99mTc was effected as in Example 6.
The resul-ts are shown in Table 4.
Table 4 3~1 Stability of Sn-PIle.A. in a lyophilized state ~stored at 4 to 8C):-¦ After production ¦100 days 200 days ,._ . . . . _ _ ~ID/ml Blood (norm~ 9.63 9.52 ._ . . ._ %/RBC _ _ !98-42_ 98.31 ~/Plasma 1.58 1.69 ._.
%ID/ml RBC (norm) 20.24 20.21 ___ ~ID in whole RBC ! 98.31 98.35 Note: Average values for five rats.
Experimental procedure: administration of Sn-10 P.Ile.A. in an amount of 20 ~g as Sn~II) per kg of the body weight of a rat, further administration of 99mTcO4 after 30 minutes and collection of blood 1 hour thereafter.
The lyophilized product was subjected to the ex-periment in the form of a solution which was prepared by adding distilled water for injection (2.0 ml) immediately before use. As can be seen from Table 4, the Sn-P.Ile.A.
in the lyophilized state is extremely stable, not showing any sign of deterioration during the period immediately after its preparation to the 200th day.
It was also confirmed that many kinds of compositions for labeling of the invention including the other six compositions shown in Example 3 showed a similar stability in the lyophilized state.
(3) Stability of a kit for preparation of a composition for labeling:
A kit for the preparation of d composition for labeling comprising Sn-P.A. and Ile. which was produced as in Example 4 was stored at 4 to 8 C for 100 or 200 days. Immediately before use, Ile (2.0 ml) ~Jas added to 30 Sn-P.A. to obtain Sn-P.Ile.A., by the use of which the 1133~31Z
intracorporeal labeling of red blood cells with 99mTc was effected as in Example 6. The results are shown in Tdble 5. As is apparent from this Table, the kit for the preparation of a composition for labeling comprising Sn-P.A. and 1le. was extremely stable, not showing any signs of deterioration during the period from its pro-duction time to the 200th day.
Table 5 Stability of a kit for preparation of a composition for labeling comprising Sn-P.A. and Ile. (stored at 4 to g C ) :--. . ._ .
After production 100 days ¦ 20Q days . .
%ID/ml Blood (norm) 9.55 9.49 ~/R~C g8.43 19g.66 ~/Plasma 1.57 1.34 _ __ %ID/ml RBC ~norm) 20.41 20.52 ._.
%ID in whole RBC 99.22 99.24 Note: Average values for five rats.
Experimental procedure: administration of Sn-P.Ile.A. in an amount of 20 ~g as Sn(II) per kg of the body ~eight of a rat, further administration of 99mTcO4 after 30 minutes and collection of blood 1 hour thereafter.
It was also confirmed that many kinds of ~its - shown in Example 5 sho~ed the same stabili~y.
Example 8 Toxicity of compositions for labeling of the invention:-In the same manner as in Example 1 or 2 but using the materials (other than water) in 5 times the amount,seven kinds of compositions for labeling were produced having 5 fold concentrations in comparison with the con~entrations of the compositions for labeling obtained in Examp1e 1 ~r 2. Each of the resulting so1utions was intravenously administered to groups consisting of lO
~prague Dawley strain ma1e rats, of lO S-D strain female rats, o~ lO ICR strain male mice and of lO ICR strain female mice at a dose of 0.5 ml per lO0 g of body weight (corresponding to an amount 1000 fold the intended dose to human beings) as well as to groups consisting of lO0 Hartley strain male guinea pigs, of lO Hartley strain female guinea pigs, of 10 male rabbits and of lO female rabbits at a dose of 0.25 ml per lO0 g Ot body w~ight (corresponding to an amount of abo~t 500 fold the inten-ded dose to human beings (per ~nit body weight)). As a control, the same vo1ume of a norm31 sa1ine so1ution was ;ntravenously administered t~ the separate groups of the same kinds of animals as above. ~hese animals were fed and observed for lO days, and the changes of the body weight wer~ recorded every d~y.
As the result of this toxicity test, no signifi-cant difference in the appearance of the animals and the change of the body weiyht was recognized between the yroups treated with the composition for labeling and the control groups. After 10 days of feeding and observation, all the animals were subjected to dissection, and observation of various organs was effected to examine histological abnormal-ities, whereby no abnormality was seen in any of the animals.
0 As shown from these results of the toxicity test, 1~3381Z
the composition for labeling of the invention produces no abnormality, even when administered to four kinds of animals (both male and female) in an amount of 500 to 1000 fold the designed dose to human beings per unit body ~eight. Thus, its toxicity is proven to be extremely low.
It should be understood by those skilled in the art that the above mentioned Examples are intended only to illustrate and explain the invention in detail and not to limit the scope of the invention.
Claims (3)
1. A non-radioactive composition for intracorporeal labeling of red blood cells with radioactive technetium, which comprises a lyophilized product containing (a) pyridoxal and/or at least one salt thereof, (b) at least one stannous salt, and (c) at least one .alpha.-amino acid as the essential components.
2. A kit for the preparation of a non-radioactive compo-sition which comprises (a) pyridoxal and/or at least one salt thereof, (b) at least one stannous salt, and (c) at least one .alpha.-amino acid as the essential components;
which kit comprises (A) an aqueous solution comprising at least one of the essential components (a), (b) and (c) or a lyophilized product therefrom, and (B) an aqueous solution comprising the remaining of those essential components or a lyophilized product therefrom.
which kit comprises (A) an aqueous solution comprising at least one of the essential components (a), (b) and (c) or a lyophilized product therefrom, and (B) an aqueous solution comprising the remaining of those essential components or a lyophilized product therefrom.
3. The kit according to claim 2, which comprises (A) an aqueous solution comprising the essential components (a) and (b) or a lyophilized product therefrom, and (B) an aqueous solution comprising the essential component (c) or a lyophilized product therefrom.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP14395678A JPS5569517A (en) | 1978-11-20 | 1978-11-20 | Labelling preparation for labelling of erythrocytes with radio-active technetium |
| JP143956/1978 | 1978-11-20 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1133812A true CA1133812A (en) | 1982-10-19 |
Family
ID=15350954
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA340,133A Expired CA1133812A (en) | 1978-11-20 | 1979-11-19 | Composition for labeling of red blood cells with radioactive technetium |
Country Status (4)
| Country | Link |
|---|---|
| US (1) | US4313928A (en) |
| EP (1) | EP0011301A1 (en) |
| JP (1) | JPS5569517A (en) |
| CA (1) | CA1133812A (en) |
Families Citing this family (17)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4504463A (en) * | 1982-06-10 | 1985-03-12 | Mallinckrodt, Inc. | Process for making a lyophilized product for use in skeletal imaging |
| US4504462A (en) * | 1982-06-10 | 1985-03-12 | Mallinckrodt, Inc. | Process for making a lyophilized product for use in skeletal imaging |
| US4443426A (en) * | 1982-06-14 | 1984-04-17 | Yale University | Blood agent |
| US4574079A (en) * | 1983-05-27 | 1986-03-04 | Gavras Haralambos P | Radiolabeled angiotensin converting enzyme inhibitors for radiolabeling mammalian organ sites |
| DE3331159A1 (en) * | 1983-08-30 | 1985-03-14 | Hoechst Ag, 6230 Frankfurt | N- (4-AMINOBENZOYL) AMINODICARBONIC ACIDS FOR THE STABILIZATION OF TECHNETIUM-99M PREPARATIONS, STABILIZED INJECTION PREPARATIONS AND METHOD FOR THEIR PRODUCTION |
| EP0169232B1 (en) * | 1984-01-23 | 1990-04-25 | For Energiteknikk Institutt | Composition for technetium-99m labelling of proteinaceous material |
| US4755375A (en) * | 1984-01-27 | 1988-07-05 | Associated Universities, Inc. | Method and kit for the selective labeling of red blood cells in whole blood with TC-99M |
| JPS61103841A (en) * | 1984-10-26 | 1986-05-22 | Nippon Mejifuijitsukusu Kk | Stable stannous chloride composition to be fed radioactivity technetium mark |
| AR246956A1 (en) * | 1987-05-08 | 1994-10-31 | Salutar Inc | Dipyridoxyl phosphate nuclear magnetic resonance imagery contrast agents |
| US4992555A (en) * | 1987-05-08 | 1991-02-12 | Salutar, Inc. | Certain dipyridylphosphate chelatable compounds capable of forming metal ion chelates |
| US5130431A (en) * | 1987-05-08 | 1992-07-14 | Salutar, Inc. | Dipyriodoxyl phosphate chelating compound intermediates |
| US4992554A (en) * | 1987-05-08 | 1991-02-12 | Salutar, Inc. | Dipyridoxyl phosphate chelating compounds |
| US5091169A (en) * | 1987-05-08 | 1992-02-25 | Salutar, Inc. | Dipyridoxyl phosphate NMRI contrast agent compositions |
| ATE80022T1 (en) * | 1988-06-07 | 1992-09-15 | Daxor Corp | AUTOMATED MULTI-POINT BLOOD VOLUME ANALYZER. |
| US20100196272A1 (en) * | 2009-01-30 | 2010-08-05 | Neoprobe Corporation | Compositions for radiolabeling diethylenetriaminepentaacetic acid (dtpa)-dextran |
| US9002656B2 (en) | 2011-10-17 | 2015-04-07 | Daxor Corporation | Automated total body albumin analyzer |
| DE112020004313T5 (en) | 2019-09-11 | 2022-06-15 | Ngk Insulators, Ltd. | Method of making a group 13 nitride crystal layer and seed crystal substrate |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3988429A (en) * | 1975-04-16 | 1976-10-26 | The United States Of America As Represented By The United States Energy Research And Development Administration | Kit for the rapid preparation of 99m Tc red blood cells |
| JPS5252427A (en) * | 1975-10-24 | 1977-04-27 | Yoshimasa Okumura | Execution method of smoke preventive vertical wall |
| JPS53148541A (en) * | 1977-05-07 | 1978-12-25 | Nihon Mediphysics Co Ltd | Radioactive diagnostic agent labelled by technesiumm99m |
-
1978
- 1978-11-20 JP JP14395678A patent/JPS5569517A/en active Granted
-
1979
- 1979-11-16 EP EP79104536A patent/EP0011301A1/en not_active Withdrawn
- 1979-11-19 CA CA340,133A patent/CA1133812A/en not_active Expired
- 1979-11-19 US US06/095,789 patent/US4313928A/en not_active Expired - Lifetime
Also Published As
| Publication number | Publication date |
|---|---|
| JPS6126887B2 (en) | 1986-06-23 |
| JPS5569517A (en) | 1980-05-26 |
| US4313928A (en) | 1982-02-02 |
| EP0011301A1 (en) | 1980-05-28 |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| MKEX | Expiry |