Patai S., Rappoport Z. 1997 The chemistry of functional groups. The chemistry of double-bonded functional groups / 0909 110

17. Syntheses and uses of isotopically labelled compounds

1009

Radioiodinated pregnenolone monoand polyiodinated benzoate esters 327a f and carbonate 327g have been obtained in pivalic acid with slightly lower yields.

O

R

(327a)	R = 2,3,5-triiodobenzoate, 95% yield after 1 h
(327b)	R = 3,4,5-triiodobenzoate, 99% yield after 1 h
(327c)	R = 2,5-diiodobenzoate, 81% yield after 1 h
(327d)	R = 2,-iodobenzoate, 82% yield after 1 h
(327e)	R = 4-iodobenzoate, 81% yield after 1 h
(327f)	R = 2,6-dimethyi-3-iodobenzoate, 63.5% yield after 1 h
(327g)	R = 4-(4-iodophenyl)butyryl-carbonate, 68% yield after 1 h

o-Iodohippuric acid (hippuran, 328), a scintigraphic agent for assessing kidney function, has been radioiodinated in pivalic acid 353similarly as had been done earlier354.

12 3 I

O

NH COOH

(328)

Other medically important compounds, such as the adrenal chemotherapeutic agent 5- iodotubercidin (329), the adenosine kinase inhibitor, 330, and the myocardial imaging agent 15-(4-iodophenyl)-pentadecanoic acid (331), have been radioiodinated in pivalic acid353 or in a melt of benzoic acid355, also in high yields. 3-Iodobenzoic acid and 3- bromobenzoic acid were radioiodinated by interhalogen exchange in low yield353 due to deactivation by the electron-withdrawing carbonyl group256. [U-125I]iodoundecanoic acid has been synthesized253 in 99% yield by 125I for Br exchange357 to give carrier-free iodoalkanoic acids.

The time and temperature dependencies of the radiochemical yield (%) have been investigated in the case of iopanoic acid (324c)353. The effect of medium (acetoamide, benzoic acid, pivalic acid) on the radioiodide exchange between Na125I and cyclohexyl iopanoate has been studied at 155 °C for 1 h. The yields were decreasing in the following order: 78% (in pivalic acid) >65% (in benzoic acid) >9% (in acetamide). By increasing the amount of Na125I and decreasing the amount of iopanoic acid, [125I]iopanoic acid with a specific activity of over 700 Ci/mmol in 60% radiochemical yield has been achieved353.

1010	Mieczysław Ziełinski´ and Marianna Kanska´
				NH2
					I
			CHCl2	N
	o-ClC6 H4 CHC6 H4 I-p				N
		(329)		N
				HOCH2 O
	p-IC6 H4 (CH2 )14 COOH
		(331)
				HO	OH
		12 5 I		(330)
	I = 12 3 I,

7. Synthesis of NCA( )-[123I]iodocyanopindolol (332)

332, a high-affinity ˇ-adrenergic antagonist, has been synthesized358 for SPECT

evaluation of cardiac	adrenergic receptor	density	by a modified chloramine-
T radioiodination of	( )cyanopindolol, 333,	with	Na[123I]iodide followed by a

novel reversed-phase HPLC purification, in 59% radiochemical yield (16 mCi). The radiosynthetic procedure will be automated to minimize radiation exposure. The commercially available ( )-[125I]iodocyanopindolol (specific activity 2175 Ci/mmol) has been used previously for intensive study of ˇ-adrenoceptors359,360.

HO

NH

O

R

CN

NH

(332)R = 12 3 I

(333)R = H

8.Synthesis of [131I]iodotamoxifen (336)

Tamoxifen, 334, employed for treatment of breast tumors which are estrogen receptor positive361, has been labelled362 with NCA 131I via an aryl diazonium salt intermediate, 335 (equation 176), for detection of breast tumors362.

335	Ð	hexafluorophosphate (PF6 )	Na131I	336
			!		176
			MeCN, 82°C, 1 min

17. Syntheses and uses of isotopically labelled compounds

R

N

O	R = N2 +; (336) R = 13 1 I
(334) R = H; (335)
OH		= 13 1 I
	I

HO	HO
I	I
(337)	(338)
O	HO

HO	HO
I	I
(339)	(340)

1011

O

9. Synthesis of [131I]-6-iodo-androsten-5-enes and [131I]-6-iodo-pregnen-5-enes

The 6-iodo-steroids, [131I]-6-iodoandrost-5-en-3ˇ,17ˇ-diol, 337, [131I]-6-iodoandrost-5- en-3ˇ-ol, 17-one, 338, [131I]-6-iodo-preg-5-en-3ˇ-ol-20-one, 339 and [131I]-6-iodopregn-

5-en-3ˇ,20-diol, 340, have been synthesized363 366 by direct mercuration of the parent-5 steroids with mercuric acetate followed by conversion of the 6-acetoxymercury compound 341 into 342 by exchange with sodium chloride, and subsequent treatment of 342 with [131I]sodium iodide in the presence of chloramine-T in 20% H2O in EtOH

1012	Mieczysław Ziełinski´ and Marianna Kanska´

(equation 177).

+

HgOA c

HO

HO

+HgOA c

(177)

		342
HO
	R	OAc
(341)	R = HgOAc	(by-product, 6-acetoxycompound)
(342)	R = HgCl

The tissue distribution study of these NCA[131I]-6-iodo-steroids in liver, spleen, adrenal, stomach, tyroid and lung showed a very rapid adrenal uptake with a peak value (in rats) at 15 min post-injection or at even shorter time periods363.

10. Synthesis of 5-iodo-2-thiouracil (ITU) (343) labelled with 123I, 125I and 131I

ITU, 343, able to bind specifically to the pigment melanin during melanogenesis, is of potential value in the diagnosis and treatment of malignant melanoma367,368. ITU labelled with radioiodine has been synthesized369 for melanoma therapy experiments directly from 2-thiouracil using commercially available lodo-GenTM in 0.05M phosphate buffer pH 7.0 (equation 178). Iodo-GenTM (1,3,4,6-tetrachloro-3˛,6˛-diphenylglycoluril generates the reactive electrophilic IC species).

O		OH
R		R
HN	Iodo-genTM	N

(178)

S	NH	HS	N
2-thiouracil, R = H (TU),		(343) 5-iodo-2-thiouracil, R = I (I TU)
		I =12 3 I, 12 5I, 13 1I

17. Syntheses and uses of isotopically labelled compounds	1013
[125I]ITU has been applied in clinical trials for distribution and clearance	studies,

[123I]ITU for the whole body imaging studies and [131I]ITU to successfully treat the Harding Passey melanoma carried in Balb/c mice369.

In previous methods of electrophilic radioiodination the sulphur has been protected by a methoxymethyl group370 or a benzyl group371 which had to be removed.

VI. SYNTHESIS AND USES OF COMPOUNDS CONTAINING C=C, C=O, OR

CN GROUPS LABELLED WITH RADIOACTIVE SULPHUR

1. Synthesis of 5f3-[2-(7-chloroquinolin-2-yl)ethenyl]phenylg-8-dimethylcarbamyl-4,6- [6-35S]dithiooctanoic acid (344)

MK-0571, 344, potent antagonist of LTD4306, has been synthesized307 to evaluate the role of leucotriene D4 in human disease states. 344 has been obtained by exchange between diacid 345 and 3-[35S]mercaptopropanoic acid in trifluoroacetic acid (equation 179). 346 has been generated from 348 prepared as shown in equations 180 and 181.

			COOH
		S			= 3 5S
				S
Cl			CONMe2
	N	S

(344)

Cl

N

(SCH2 CH2 COOH)2

(345)

(179)

CF3 COOH, 1 h

H3 5SCH2 CH2 CONMe2

(346)

344 (45% yield) + 345 +

(SCH2 CH2 CONM2 )2

(347)

H

3 5S,CF

COOH

Ph3 C3 5SH

CH2

CHCONMe2

Ph3 C3 5SCH2 CH2 CONMe2

Ph3 COH

2

3

MeCN, DBU

(348)

(180)

1. n-BuLi, THF

Ph3 CH

2. [3 5S]sulphur

Ph3 C3 5SH

Ph3 C3 5SCH2 CH2 CONMe2

(181)

3. n-Bu3 P
4. CF3 COOH	(348)

1014	Mieczysław Ziełinski´ and Marianna Kanska´

2. Synthesis of pidotimod (349) labelled with 14C- and 35S-isotopes

The 14C-labelled 349, the new immunostimulating agent with a peptide-like struc-

ture, has been synthesized372 from 350 (equation 182). [35S]-Pidotimod-349 has been prepared372,373,374a as shown in equation 183, [14C]- and [35S]-349 have been needed to study the pharmacokinetics and metabolism of 349 in immunodeficiencies. 349 increases cell-immediate immune response by stimulation of IL-351 production374b,374c.

205 °C

HOOCCH2 CH2 CH

COOH

0.3 mm Hg

O

COOH

NH2

NH

(350) L-[U-14 C]

(351)

DCC

S

(182)

NH COOEt

S

S

1. OH−

N

N

O

2. 37% HCl

O

NH

NH

O

COOEt

O

COOH

[14 C]-(349)70.5% yield from 351

S

HCHO

HOOC

CHCH2 SH

RT, overnight

N

COOH

NH2 .HCl

(183)

H.HCl

Cl

O

O

Cl

DMF, Et3 N,RT,24h

NH

O

Cl

Cl

Cl

[3 5S]-349

VII. ISOTOPE EFFECT STUDIES

A. Isotope Effect Studies of Chemical Reactions

1. Oxygen-18, nitrogen-15 and ˇ-deuterium isotope effects in the transfer reactions of p-nitrophenyl acetate (PNPA) with various nucleophiles

The 18O, 15N and ˇ 2H isotope effects in the acyl transfer reactions of PNPA (equation 184) have been studied375, and the conclusion has been reached that these

17. Syntheses and uses of isotopically labelled compounds

1015

transfers are concerted with no stable tetrahedral intermediate of a significant lifetime.

18

O

18 O-

Nu

−

+ CD3 C

18

OC6 H4

15

k1

Nu

C

18 OC6 H4

15NO2

NO2

k-1

CD3

(184)

18 O

k2

where:

NuCCD3 + O2

15NC6 H4 18 O−, k2 >> k−1

Nu = hydroxide

15k: 1.0002

š

0.0001

(pK D 15.7)

18

18

klg: 1.0135 š 0.0007

(room temperature)

kcarbonyl: 1.0039 š 0.0003

(pH of 9.5)

Dk (for 3 deuterium atoms, CD3): 0.9562 š 0.0008

Phenolate (pK D 9.9)

15

k: 1.0010

š 0.0002

18

(ice bath temperature)

18

klg: 1.0199

š 0.0009

pH of 9.7)

kcarbonyl: 1.0043

š 0.0008

(CF3)2CHO

Dk(D3): 0.9583

š 0.0010

15k: 1.0010

š

0.0002

18

(pK of 9.3)

18

klg: 1.0210

š 0.0010

(RT, pH 9)

kcarbonyl: 1.0058

š 0.0006

D (D3): 0.9481

š

0.0030

mercaptoethanol

k

15

k: 1.0001

18

š 0.0003

(pK D 9.5)

18

klg: 1.0219

š 0.0009

(RT, pH 9)

kcarbonyl: 1.0119

š 0.0003

D (D3): 0.9780

š

0.0008

k

15

anion of HSCH2CH2COOMe

18

k: 1.0003

š 0.0001

(pK D 9.3)

18

klg: 1.0172

š 0.0004

(RT, pH 9)

kcarbonyl: 1.0117

š 0.0004

D (D3): 0.9765

š

0.0006

methoxyethylamine

k

15

k: 1.0011

18

š 0.0001

(RT, pH 9)

klg: 1.0330

š 0.0007

18kcarbonyl: 1.0064

š 0.0003

Dk(D3): 0.9682

š 0.0010

The RT (at pH 9) equilibrium isotope effect determinations are presented in the upper, lower and right-hand side of equation 185. They have been used to calculate the isotope effect for the equilibrium between p-nitrophenolate and PNPA in water since the PNPA undergoes hydrolysis during the equilibrium. The 18Keq isotope effect for deprotonation was found to be 1.01533. The earlier reported value376 was 1.018 š 0.002.

The computed 18Kcarbonyl oxygen isotope effects for the conversion of the carbonyl double bond of the reactant to the nearly single bond of a tetrahedral intermediate are

about 1.025 for the very late transition state377 379. The computed Dk values have been estimated to be 0.89 (inverse, also for a very late TS of the formation of tetrahedral intermediate). In the concerted mechanism (equation 186) the carbonyl -bond may not be

1016	Mieczysław Ziełinski´ and Marianna Kanska´

altered. In this study the 18kcarbonyl is in the range 1.0039 1.0058 for oxyanion reactions, indicating some tetrahedral character in the TS only. The magnitudes of the isotope effect

for the p-nitrophenyl leaving group are lower than those expected for an alkyl leaving group due to the contribution of the resonance structure which compensates to some extent the loss of the bond order to the carbonyl carbon in the TS. In the reaction with both phenolate nucleophiles the nitrogen-15, 15k, effects are nearly unity. The magnitudes of

18kcarbonyl effects in reactions with thiolates are twice those of the oxyanion reactions, indicating greater loss of carbonyl -bond and the TS structure shown below.

−

Keq (phenolate in

−

H2 O/phenolate in CHCl3 )

OC6 H4 NO2

OC6 H4 NO2

1 5Keq = 1 .001 0 ± 0.0002

(in H2 O)

(in CHCl3 )

(1 5N concentrates in the aq. phase)

1 8 Keq = 0.9977 + 0.0001

(1 8 O concentrates in CHCl3 )

1 5Keq = 0.9982 + 0.0003 = (1 / 1.001803)

Keq

phenolate in CHCl3

1 8 Keq = 0.9711 + 0.0001 = (1 / 1.02976)

PNPA in CHCl3

PNPA in H2 O

1 5Keq = 1.0016 + 0.0006

calculated

Keq

1 8

Keq = 1.0277 + 0.0007

values

phenolate in H2 O

O

O

Keq (PNPA in CHCl3 /PNPA in H2 O)

MeCOC6 H4 NO2

MeCOC6 H4 NO2

1 5Keq = 0.9992 + 0.0001

(in H2 O)

(in CHCl3 )

1 8 Keq = 1.0043 + 0.0005

(185)

−

−

O

O

(186)

CH3 CHO + HO−

CH3

C

OH

CH3 C

H

H

OH

		+	O−
	O	N
			O−
	δ −	O−	δ +
O2 N	O	C CH3 TS

SR

17. Syntheses and uses of isotopically labelled compounds

1017

The differences between hydrosulphide and hydroxide have been investigated theoretically in the case of carbonyl addition reactions to formaldehyde and formamide380. The data for the reaction of PNPA with methoxyethylamine indicate that the bond cleavage to the leaving group takes place in the rate-limiting step of this last reaction to a higher degree than in the case of other nucleophiles, and/or smaller charge delocalization which reduced the 18Klg values in the previous reactions more than one-half. The maximum 18O KIE (value of reduced partition function D 1.0408) for 12C-16O/12C-18O bond rupture equals 1.0665 at 25 °C. No temperature dependence of the above KIEs have been studied. The 1H/2H, 12C/13C, and 16O/18O KIE and equilibrium IE have been computed378.

2. Deuterium, carbon-13 and oxygen-18 KIE in the hydrolyses of ˛- and ˇ- glucopyranosyl fluorides

The kinetic isotope effects klight/kheavy, for the hydrolysis at pH 6.0 of ˛-glucosyl fluoride at 80 °C and ˇ-glucosyl fluoride at 50 °C, isotopically substituted as shown in

structure 352, have been found381 to be as follows:

HO	D
C		18O	D
HO C		13 C
C		C	F
HO	HO	D

(352)

for ˛-D (C 1 ) site: 1.142 š 0.0075 and 1.086 š 0.0012, for ˇ-D (C 2 ) site: 1.067 š 0.0077 and 1.030 š 0.0083, for -D (C 5 ) site: 0.979 š 0.0032,

for anomeric [1-13C]: 1.032 š 0.0032 and 1.017 š 0.0022, for ring 18O: 0.984 š 0.0049 and 0.985 š 0.0049,

In the reaction of the ˛-glucosyl fluoride with azide ion, the ˛-D effect increased modestly to 1.169 š 0.0082 while the [1-13C] effect increased to 1.085 š 0.0082.

In the hydrolysis of glucosyl fluoride in 0.3 M sodium succinate buffer (pH 6.0, I D 1.0 M NaClO4) at 50 °C, the KIEs are as follows:

for ˛-D: 1.105 š 0.0053, for ˇ-D: 1.059 š 0.0010, for (5-D): 0.981 š 0.0074,

for anomeric [1-13C]: 1.064 š 0.0070, for ring [5-18O]: 0.988 š 0.0036.

BEBOVIB-IV-TS structure for ˛-glucosyl fluoride hydrolysis involving the ring in a flattened 4C1 chair conformation and water and fluoride ion about the anomeric centre has been presented381,382. The conformations based on this structure reproduced well the ˛-D, ˇ-D, 5-D and [5-18O] experimental KIE, but not the [1-13C] KIE (theor. KIE D 1.023 compared with 1.032 for exp. KIE).

The qualitative TS structure for succinate-catalyzed hydrolysis of ˇ-glucosyl fluoride, shown as 353, has been proposed381.

1018	Mieczysław Ziełinski´ and Marianna Kanska´
			O
	OH		O
			H
	O		F
HO
HO		O−	O	OH
			(353)

3. Deuterium IE study of oxime formation

The rate-determining step of the oxime formation from carbonyl compounds (equation 187) changes with increasing pH from formation of the addition intermediate to its dehydration to the products.

HO

H

C O + NH2 OH

k

a d d

kd

2

C NOH

+ H2 O

C

N

OH

	kd2 = kH [H+] +	ko +	kOH [OH−]	H
				(187)

At pD 6.9 (calculated from the relation pD D pHC0.4) the solvent DIE in the oxime formation from hydroxyamine with cyclohexanone and bicyclic ketones equals383 kH/kD D 1.50 š 0.05 and the reaction occurs via a rate-limiting dehydration of the addition intermediate being in equilibrium with the ketone and hydroxyamine. The TS of the acid catalyzed reaction (pH 4 to ca 8) is characterized by a moderate cleavage of the C O bond384 with the carbon atom nearer to sp3 than to sp2. In the pH-independent region (pH 8 to ca 10) the TS with more extensive C O cleavage and with the carbon atom nearer to sp2 than to sp3 has been assumed. The deuterium solvent IE of 1.50 in neutral solution also indicates that not the N D bond rupture but the C OD bond rupture is rate-determining.

Two mechanisms shown in equations 188 and 189 have been proposed385 (pH 10 to ca 13).

B: + H

kc

BH+ + H N C

+ OH−

N

C

OH

slow

H

(188)

fast

B + H N

C + H2 O

In the base-catalysed dehydration for oxime formation a significant development of the carbon nitrogen double bond in the TS is assumed.

4. Deuterium IE studies with malonamide

In the course of mechanistic studies of the nitrosation (equation 190), bromination and iodination of malonamide (MA), proceeding according to the mechanism involving