JPH11332589A - Synthesis of (11-c)-l-methionine - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain the subject compound useful as a labeling agent, etc., for diagnosing cerebral functions according to a positron emission tomography, etc., by making a γ-cyano-α-aminobutyric acid syntheses act on [11 C]methanethiol. SOLUTION: [11C]Methanethiol represented by the formula<11> CH3 SH, [11 C]ethanethiol represented by the formula CH3 -<11> CH2 -SH, [11C]propanethiol represented by the formula CH3 CH2 -<11> CH2 SH, etc., are respectively converted into [11C]-L-methionine represented by formula I, [11C]-L-ethionine represented by formula II and [11C]-L-propionin represented by formula III according to an enzymic reaction with a γ-cyano-α-aminobutyric acid syntheses to thereby afford the objective compounds in a high radiochemical yield by simple operations without producing optical isomers. The compounds are useful as a labeling agent, etc., for diagnosing the cerebral functions or tumor tissues according to a positron emission tomography, etc.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present application relates to [11C] -L-methionine and [11C]-which are used as a labeling agent for diagnosing brain function or tumor tissue by positron emission tomography (PET) or the like.
The present invention relates to a method for synthesizing L-ethionine and [11C] -L-propionin.

[0002]

2. Description of the Related Art Conventionally, [11C] -L- has been used as a labeling agent for diagnosing tumor tissues or PET for structural and functional disorders of the brain that cause various neuropsychiatric disorders.
Methionine is used. As a method for synthesizing [11C] -L-methionine, for example,

[0003]

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[0004] A chemical synthesis method using a liquid phase as shown in the reaction formula (European Journal of Nuclear Medicine 1: 11-14,
1976) is known. The process of this method is as follows according to the description of the above-mentioned literature. a) [11C] trapped in 100 μL of acetone
Methyl iodide is added to a reaction vessel containing 300 μL of an aqueous solution of DL homocysteine or L-homocysteine thiolactone hydrochloride (2 mg) and 70 μL of an aqueous solution of 10 M sodium hydroxide. b) Seal the container and heat at 100 ° C. for 8 minutes. After cooling, acetone and unreacted [11C] methyl iodide in the reaction solution are distilled off under heating at 40 ° C. in a nitrogen stream. c) After a few minutes, add 20 μL of 10N hydrochloric acid, 1 mL of distilled water and 2 mL of physiological saline to the reaction vessel. 0.
After adjusting the pH of the reaction solution to 4 with 1N hydrochloric acid or sodium hydroxide aqueous solution, the reaction solution is filtered with a membrane filter.

[0005] The synthesis time in this method is within 25 minutes from the end of irradiation. The radiochemical yield of the obtained [11C] -L-methionine was 50 ± 11% based on [11C] methyl iodide, and the specific radioactivity was about 50 mCi / μm.
ol. This method has the advantages that the radiochemical yield is high, high specific radioactivity is not required as a drug, and removal of the raw material (homocysteine thiolactone) is not necessarily required.

Further, by improving the above method, the following equation is obtained.

[0007]

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[0008] On-column chemical synthesis method (Applied Radiation and Isotopes 44: 788-790, 1
993) is also known. The process of this method is as follows, for example. a) A column filled with a carrier holding L-homocysteine thiolactone and cooled to about −40 ° C. is collected by passing [11C] CH ₃ I under an arbitrary flow rate of He gas flow. b) Fill the column with 70% EtOH, close both ends, and heat at about 60 ° C. for about 3 minutes. c) Wash the reaction product from the column with HCl, then with local ethanol and collect in an evaporator flask. d) After removing the solvent, physiological saline is added to dissolve the residue, and the solution is passed through a membrane filter to obtain a drug for injection.

In this method, the synthesis is completed within 20 after the end of the irradiation. The radiochemical yield of the obtained [11C] -L-methionine was 9 relative to [11C] CH ₃ I.
8% or more (specific radioactivity is unknown). This method is superior to the above-mentioned synthesis method using a liquid phase, in that the operation is simplified because of on-column synthesis and the radiochemical yield is improved.

[0010]

However, any of the above chemical synthesis methods has a problem that an optical isomer of the target compound is formed as a by-product. This application has been made in view of the circumstances as described above, and solves the problems of the conventional synthesis method, without generating optical isomers, with a high radiochemical yield by a simple operation. [11C]
It is an object of the present invention to provide a new synthetic method capable of obtaining -L-methionine.

[0011] This application also discloses that the radiochemical yield is high with simple operation without producing optical isomers.
Another object of the present invention is to provide a new synthetic method capable of obtaining [1C] -L-ethionine and [11C] -L-propionin. [11C] ethanethiol and [11C] which are novel synthetic intermediates in the above method
It also aims to provide propanethiol and a method for synthesizing it.

[0012]

According to the present invention, a first invention for solving the above-mentioned problems is represented by the following formula (1).

[0013]

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[11C] methanethiol represented by the formula:
By the enzymatic reaction by γ-cyano-α-aminobutyric acid synthase, the following formula (2)

[0015]

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The present invention also provides a method for synthesizing [11C] -L-methionine (claim 1), which is converted to [11C] -L-methionine represented by the formula: In the method of the first invention, the following equation (3)

[0017]

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Synthesizing [11C] methanethiol from [11C] methyl iodide represented by the following formula:
Is a preferred embodiment. In the method of the second aspect, the following equation (4) is used.

[0019]

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[11C] carbon dioxide represented by the formula [1]
In a preferred embodiment, 1C] methyl iodide is synthesized (claim 3). This application also provides, as a second invention, the following formula (5):

[0021]

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[11C] ethanethiol represented by the following formula:
By the enzymatic reaction by γ-cyano-α-aminobutyric acid synthase, the following formula (6)

[0023]

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The present invention provides a method for synthesizing [11C] -L-ethionine, which is characterized by converting into [11C] -L-ethionine. In the method of the second invention, the following equation (7)

[0025]

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Synthesizing [11C] ethanethiol from [11C] ethyl iodide represented by the following formula (5):
Is a preferred embodiment. In the method of claim 5, the following equation (8)

[0027]

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In a preferred embodiment, [11C] ethyl iodide is synthesized from methylmagnesium bromide represented by the following formula and [11C] carbon dioxide.
Further, this application provides a third method using the following equation (9).

[0029]

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[11C] propanethiol represented by the following formula (10) is obtained by an enzymatic reaction using γ-cyano-α-aminobutyric acid synthase.

[0031]

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The present invention provides a method for synthesizing [11C] -L-propionin, which is characterized in that it is converted to [11C] -L-propionin represented by the following formula (7). In the method of the third invention, the following equation (11)

[0033]

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In a preferred embodiment, [11C] propanethiol is synthesized from [11C] propyl iodide represented by the following formula (claim 8). In the method of claim 8, the following equation (12)

[0035]

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In a preferred embodiment, [11C] propyl iodide is synthesized from ethylmagnesium bromide represented by the formula and [11C] carbon dioxide. Furthermore, this application has the following formula (5)

[0037]

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[11C] ethanethiol represented by the following formula:
The following equation (9)

[0039]

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[11C] propanethiol represented by the formula:

[0041]

DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, embodiments of each invention of this application will be described in detail. The most preferred embodiment of the first invention of this application is the following formula:

[0042]

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The reaction formula can be exemplified as follows. In this reaction formula, [11C] methyl iodide is synthesized from [11C] carbon dioxide by a known method (Appl. Radiat. Iso).
t. 44 (8): 1119-1124, 1993). The synthesis of [11C] methanethiol from [11C] methyl iodide is also performed by a known method (J. Label.
Compds. Radiopharm. 37: 109, 1995).

The thus synthesized [11]
C] Methanethiol is converted to [11C] -L-methionine by an enzymatic reaction with γ-cyano-α-aminobutyric acid synthase. That is, this enzyme uses pyridoxal phosphate as a coenzyme to convert [11C] -L from O-acetyl-L-homoserine and [11C] methanethiol.
-Produces methionine. And this γ-cyano-α
By immobilizing -aminobutyric acid synthase on a column, [11C] -L-methionine can be synthesized by an on-column synthesis method, for example, as follows.

[11C] To methanethiol, an enzyme reaction volume substrate solution consisting of sodium phosphate buffer, O-acetyl-L-homoserine and pyridoxal phosphate was added, and this mixed solution was sent to an immobilized enzyme column. The eluate containing the target compound [11C] -L-methionine is collected in an eggplant flask or the like. Then, to wash the column,
The substrate solution of the enzyme reaction volume is again sent to the immobilized enzyme column, and the eluate is collected in an eggplant flask or the like. After collecting the eluate, the eluate is dried under reduced pressure using an evaporator or the like, and the residue in an eggplant flask or the like is dissolved in physiological saline for injection.

As γ-cyano-α-aminobutyric acid synthase, for example, Chromobacterium
u) γ-cyano-α-aminobutyric acid synthase (Biochemist) obtained by isolation from a bacterium belonging to the genus (C. violaceum)
ry, 12: 5369-5377, 1973) and B. stearothes found by the inventors of the present application.
rmophilus) CN3 strain (FERM BP-4773) and a heat-stable γ-cyano-α-aminobutyric acid synthase (Japanese Patent Application No. 7-171171) can be used. Alternatively, γ-cyano-α-amino acid obtained by expressing the gene encoding γ-cyano-α-aminobutyric acid synthetase (Japanese Patent Application No. 9-34612) found by the present inventors in a microorganism such as Escherichia coli. Aminobutyric acid synthase can also be used.

The most preferred embodiments of the second invention and the third invention of this application are as follows:

[0048]

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[0049]

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The reaction formulas can be exemplified as follows. In these reactions, except that [11C] ethyl iodide was synthesized from methylmagnesium bromide and [11C] carbon dioxide, and [11C] propyl iodide was synthesized from ethylmagnesium bromide and [11C] carbon dioxide. 1 By the same procedure as the method of the invention,
[11C] -L-ethionine and [11C] -L-propionin can be synthesized.

[0051]

EXAMPLES Hereinafter, the present invention of this application will be described in more detail and concretely with reference to examples, but the present invention is not limited to the following examples. The materials used in the following examples are as follows. [11C]
Is 1 using a small cyclotron CYPRIS370.
Manufactured with 4N (p, α) 11C.

Γ-cyano-α-aminobutyric acid synthase (E
C4.4.1.9) was isolated and purified from recombinant Escherichia coli into which the Bacillus stearothermophilus γ-cyano-α-aminobutyric acid synthase gene described in Japanese Patent Application No. 9-34612 was introduced. The specific activity of γ-cyano-α-aminobutyric acid synthase was determined by measuring L-methionine produced using O-acetyl-L-homoserine and methanethiol as substrates, and the specific activity was 26.7 mol / min / min. mg protein. Protein concentration is 14.4 mg / m
l. To maintain the activity of the enzyme, γ-cyano-
The enzyme solution prepared for α-aminobutyric acid synthase was stored in a freezer little by little.

The immobilized enzyme column was prepared according to the following procedure. Glutaraldehyde aqueous solution (2.5%, 10 m) was added to aminopropyl-CPG (0.2 g, pore size 1400 angstroms: sold by Funakoshi Co., Ltd.).
l) was added, and the mixture was reacted at room temperature for 1 hour, and then thoroughly washed with water. In addition, γ-cyano-α-aminobutyric acid synthase (1
0 mg) of sodium phosphate buffer (pH 7.5,
0.1 M, 10 ml) and react at 4 ° C. with stirring overnight. Thereafter, unreacted γ-cyano-α-aminobutyric acid synthase is removed to prepare immobilized γ-cyano-α-aminobutyric acid synthase.

A column (a cylinder having a hollow portion) having an inner diameter of 4.2 mm and a length of 7.0 mm (internal volume: 0.1 ml) was filled with the above-mentioned immobilized γ-cyano-α-aminobutyric acid synthase, [11C] -L-methionine synthesis reaction column. Liquid chromatography was performed using a liquid chromatograph equipped with a UV detector, in series with a NaI radioactivity detector, using the following columns.

Column (A): 4.6 × 150 mm C-18 (Crest-
Pak C18S) column. Column (B): 4.0 × 150 mm C-18 (Crownp
ak CR (+)) column. Mobile phase (C): 0.01 M ammonium formate. Mobile phase (D): methanol / 0.01 M ammonium formate (10:90, v / v).

Mobile phase (E): 0.01 N perchloric acid (pH
2.0). All other chemical reagents were used without purification. Example 1 [11C] -L-methionine was synthesized by the following steps. (1) Synthesis of [11C] methyl iodide [11C] Methyl iodide was synthesized by a known method (Appl.
Radiat. Isot. 44 (8): 1119-1124, 1993).

[11C] carbon dioxide obtained from the cyclotron was placed in a reaction flask cooled to −18 to −20 ° C. by a nitrogen gas stream in a THF solution of lithium aluminum hydride (0.1 M, 0.15 ml). Led to. After evaporating the THF, hydroiodic acid (57%,
0.5 ml) and heated to 120 ° C. to produce [1
[1C] Methyl iodide was distilled off and transferred to a [11C] methanethiol synthesis reaction vessel. (2) Synthesis of [11C] methanethiol [11C] methyl iodide obtained in (1) was converted into sodium hydrosulfide (2-5 mg) and DMF by a nitrogen stream.
(0.2-0.5 ml) and transferred to a [11C] methanethiol synthesis reaction vessel. During the transfer, the reaction vessel was heated at 80 to 120 ° C. for 4 minutes, and the produced [11C] methanethiol was cooled to −40 ° C. or lower with a mixture of ethanol and liquid nitrogen by a nitrogen stream, and acetone (0.degree.
8 ml) into a 10 ml [11C] methanethiol trap vial. The radiochemical purity of [11C] methanethiol trapped in acetone was measured by analytical liquid chromatography using column (A). The chromatography conditions were as follows.

Flow rate: 2 ml / min, solvent: (C), column temperature: room temperature, wavelength: 210 nm, retention time of [11C] methanethiol: 4 to 4.5 minutes. (3) Production of [11C] -L-methionine [11C] After completion of the methanethiol trap,
1C] Immobilized enzyme reaction consisting of sodium phosphate buffer (pH 7.5, 125 mM), O-acetyl-L-homoserine (16.7 mM), and pyridoxal phosphate (0.1 mM) in a vial for methanethiol trap Volume substrate solution (1.2 ml) was added. This mixed solution is sent to an immobilized enzyme column, and the target compound [11C]-
The eluate containing L-methionine was collected in an eggplant flask. Next, by sending the same amount of the substrate solution to the immobilized enzyme column, [11C]-
The L-methionine was washed, and the eluate was collected in the eggplant flask described above. After collecting the eluate, the residue was dried under reduced pressure using an evaporator, and the residue in the eggplant flask was dissolved in physiological saline for injection. The lysate was sterilized through a 0.22 μm pore filter and filled into sterile vials. This vial was used for human or animal administration.

[11C] -L obtained as the final product
-The radiochemical purity of methionine was measured by analytical liquid chromatography using the column (A), and as a result, was 99% or more, and the yield was 60 to 70%. In addition,
The chromatography conditions were as follows. Flow rate: 2 ml / min, solvent: (C), column temperature: room temperature,
Wavelength: 210 nm, retention time of [11C] -L-methionine: 1.5 to 2.0 minutes.

Further, the non-radioactive methionine sample [11
FIG. 1 shows the results obtained by adding C] to methionine.
The position of the UV detection signal and the position of the radiation detection signal were consistent, and the compound obtained thereby was [11C] -L-
It was confirmed to be methionine. The optical purity of [11C] methionine obtained as a final product by the above method was 99% or more as a result of measurement by analytical liquid chromatography using column (B). In addition, the conditions of the chromatography were as follows.

Flow rate: 0.8 ml / min, solvent: (E), column temperature: room temperature, wavelength: 210 nm, [11C] -L-
Retention time for methionine: 4.5-5.5 minutes. Example 2 [11C] -L-ethionine was synthesized by the following steps. (1) Synthesis of [11C] ethyl iodide [11C] Ethyl iodide was synthesized by a known method (Procee
ding of 12th International Symposium on Radiopha
rmacuetical Chemistry, 252-253, 1997)
The following procedure was used.

The [11C] carbon dioxide obtained from the cyclotron was fed into a reaction flask with a THF solution of methylmagnesium bromide (1.0 mL) by a nitrogen gas stream.
M, 0.15 ml). After the reaction, lithium aluminum hydride (0.5 M, 0.25 ml) was added to the reaction solution, and the mixture was heated. After evaporating the THF, add hydroiodic acid (0.5 ml) and heat to 120 ° C., [11C]
A mixture of ethyl iodide and [11C] methyl iodide was obtained. After the mixture was distilled and trapped in a stainless steel tube cooled with liquid argon, the liquid phase was
The mixture was led to a Chromosolve W-HP gas chromatography column of V101, and these mixtures were separated and purified at room temperature in a helium stream (100 mL / min). [11]
[C] Ethyl iodide was transferred to a reactor for [11C] ethanethiol synthesis. (2) Synthesis of [11C] ethanethiol [11C] ethanethiol was synthesized from [11C] ethyl iodide obtained in (1) in the same manner as in Example 1 (2).

The obtained [11C] ethanethiol was analyzed in the same manner as in Example 1 except that (C) was used as the solvent.
The analysis was performed under the same conditions as in (2) (retention time: 8.5 to 9.0).
Minutes). (3) Synthesis of [11C] -L-ethionine From [11C] ethanethiol obtained in (2) above,
[11C] -L-ethionine was synthesized in the same manner as in Example 1 (3).

The obtained [11C] -L-ethionine was analyzed under the same conditions as in Example 1 (3) (retention time: 2.5 to 3.0 minutes). As a result, [11
The radiochemical purity of [C] -L-ethionine was 99% or more, and the yield was 50 to 60%. FIG. 2 shows the results obtained by adding a non-radioactive L-ethionine sample to [11C] -L-ethionine. The position of the UV detection signal coincided with the position of the radiation detection signal, confirming that the resulting compound was [11C] -L-ethionine.

The optical purity of [11C] -L-ethionine obtained as a final product by the above method was measured by analytical liquid chromatography using column (B). The chromatography conditions were as follows. Flow rate: 0.8 ml / min, solvent: (E), column temperature: room temperature, wavelength: 210 nm, retention time: 9.0-9.5 minutes. Example 3 [11C] -L-propionin was synthesized by the following steps. (1) Synthesis of [11C] propyl iodide [11C] Synthesis of propyl iodide was performed by a known method (Proc
eeding of 12th International Symposium on Radiop
harmacuetical Chemistry, 252-253, 1997).

[11C] carbon dioxide obtained from the cyclotron was mixed with a THF solution of ethylmagnesium bromide (1.0 mL) in a reaction flask by a nitrogen gas stream.
M, 0.15 ml). After the reaction, lithium aluminum hydride (0.5 M, 0.25 ml) was added to the reaction solution, and the mixture was heated. After evaporating the THF, add hydroiodic acid (0.5 ml) and heat to 120 ° C., [11C]
A mixture of propyl iodide and [11C] methyl iodide was obtained. After distilling this mixture and trapping it in a stainless steel tube cooled with liquid argon, the liquid phase was led to a Chromosolve W-HP gas chromatography column having OV101 of 10%, and was placed in a helium gas stream at room temperature (100 mL / min).
These mixtures were separated and purified. [11]
[C] Propyl iodide was transferred to a [11C] propanethiol synthesis reactor. (2) Synthesis of [11C] propanethiol From [11C] propyl iodide obtained in (1) above,
[11C] propanethiol was synthesized in the same manner as in Example 1 (2).

The analysis of the obtained [11C] propanethiol was conducted in the same manner as in Example 1 except that (D) was used as the solvent.
Analysis was performed under the same conditions as in (2) (retention time: 9.0 to 10.
0 minutes). (3) Synthesis of [11C] -L-propionin [11C] -L from [11C] propanethiol obtained in (2) above in the same manner as in Example 1 (3).
-Propionin was synthesized.

The obtained [11C] -L-propionin was analyzed under the same conditions as in Example 1 (3) (retention time: 3.5 to 4.0 minutes). As a result, [11
The radiochemical purity of [C] -L-propionin was 99% or more, and the yield was 40 to 50%. FIG. 3 shows the results obtained by adding a non-radioactive L-propionin sample to [11C] -L-propionin. The position of the UV detection signal coincided with the position of the radiation detection signal, confirming that the obtained compound was [11C] -L-propionin.

The optical purity of [11C] -L-propionin obtained as a final product by the above method was measured by analytical liquid chromatography using column (B). The chromatography conditions were as follows. Flow rate: 0.8 ml / min, solvent: (E), column temperature: room temperature, wavelength: 210 nm, retention time: 20.0 to 21.0
Minutes.

[0070]

As described in detail above, according to the present invention, [11C] -L-methionine having a high radiochemical yield can be obtained by a simple operation without producing optical isomers.
There is provided a new synthetic method capable of obtaining [11C] -L-ethionine and [11C] -L-propionin. Thereby, for example, it becomes possible to improve the accuracy of PET diagnosis of tumor tissue and brain.

[Brief description of the drawings]

FIG. 1 shows the results obtained by adding a non-radioactive L-methionine sample to [11C] -L-methionine obtained in Example 1, and (A) a UV detection signal of the non-radioactive L-methionine sample; (B) Radiation detection signal of [11C] -L-methionine.

FIG. 2 shows the results obtained by adding a non-radioactive L-ethionine sample to [11C] -L-ethionine obtained in Example 2, and (A) a UV detection signal of the non-radioactive L-ethionine sample; (B) Radiation detection signal of [11C] -L-ethionine.

FIG. 3 shows the results obtained by adding a non-radioactive L-propionin sample to [11C] -L-propionin obtained in Example 3; (A) UV of a non-radioactive L-propionin sample;
Detection signal, (B) Radiation detection signal of [11C] -L-propionin.

 ──────────────────────────────────────────────────続 き Continuing on the front page (72) Inventor Yuji Furuya 20-9 Jusankenya, Kanbe-cho, Shenzhen-gun, Hiroshima Prefecture

Claims (11)

[Claims] 1. The following formula (1) [11C] methanethiol represented by the formula:
By the enzymatic reaction by α-aminobutyric acid synthase, the following formula (2) A method for synthesizing [11C] -L-methionine, which comprises converting into [11C] -L-methionine represented by the following formula: 2. The following formula (3): The method for synthesizing [11C] -L-methionine according to claim 1, wherein [11C] methanethiol is synthesized from [11C] methyl iodide represented by the formula: 3. The following formula (4): The method for synthesizing [11C] -L-methionine according to claim 2, wherein [11C] methyl iodide is synthesized from [11C] carbon dioxide represented by the formula: 4. The following formula (5): [11C] ethanethiol represented by the formula:
By the enzymatic reaction of α-aminobutyric acid synthase, the following formula (6) is obtained. A method for synthesizing [11C] -L-ethionine, which comprises converting into [11C] -L-ethionine represented by the formula: 5. The following formula (7): The method for synthesizing [11C] -L-ethionine according to claim 4, wherein [11C] ethanethiol is synthesized from [11C] ethyl iodide represented by the formula: 6. The following formula (8): The method for synthesizing [11C] -L-ethionine according to claim 5, wherein [11C] ethyl iodide is synthesized from methyl magnesium bromide represented by the formula and [11C] carbon dioxide. 7. The following formula (9): [11C] propanethiol represented by the following formula (10) is obtained by an enzymatic reaction using γ-cyano-α-aminobutyric acid synthase. [11C] -L-propionin; [11C] -L-propionin. 8. The following formula (11): [11C] -L- according to claim 7, wherein [11C] propanethiol is synthesized from [11C] propyl iodide represented by the formula:
A method for synthesizing propionin. 9. The following formula (12) The method for synthesizing [11C] -L-propionin according to claim 8, wherein [11C] propyl iodide is synthesized from ethylmagnesium bromide represented by the formula and [11C] carbon dioxide. 10. The following formula (5): [11C] ethanethiol represented by the formula: 11. The following formula (9): [11C] propanethiol represented by the formula: