Radioiodination of Aryl-alkyl cyclic sulfates

Article abstract of DOI:10.3390/molecules171113266

Among the currently available positron emitters suitable for Positron Emission Tomography (PET), ¹²⁴I has the longest physical half-life (4.2 days). The long half-life and well-investigated behavior of iodine in vivo makes ¹²⁴I very

Full text of DOI:10.3390/molecules171113266

Molecules 2012, 17, 13266-13274; doi:10.3390/molecules171113266
OPEN ACCESS
molecules
ISSN 1420-3049
www.mdpi.com/journal/molecules
Communication
Radioiodination of Aryl-Alkyl Cyclic Sulfates
Chandra Mushti 1,* and Mikhail I. Papisov 2,*
1
Massachusetts General Hospital, Harvard Medical School and Shriners Hospital for Children,
Shriners Hospital for Children—Boston, Room 439, 51 Blossom St., Boston, MA 02114, USA
2
Massachusetts General Hospital, Harvard Medical School and Shriners Hospitals for Children,
Shriners Hospital for Children—Boston, Room 224, 51 Blossom St., Boston, MA 02114, USA
* Authors to whom correspondence should be addressed; E-Mails: cmushti@partners.org (C.M.);
papisov@helix.mgh.harvard.edu (M.I.P.); Tel.: +1-617-726-2000 (M.I.P).
Received: 7 September 2012; in revised form: 24 September 2012 / Accepted: 23 October 2012 /
Published: 7 November 2012
Abstract: Among the currently available positron emitters suitable for Positron Emission
Tomography (PET), 124I has the longest physical half-life (4.2 days). The long half-life and
well-investigated behavior of iodine in vivo makes 124I very attractive for pharmacological
studies. In this communication, we describe a simple yet effective method for the synthesis
of novel 124I labeled compounds intended for PET imaging of arylsulfatase activity in vivo.
Arylsulfatases have important biological functions, and genetic deficiencies of such
functions require pharmacological replacement, the efficacy of which must be properly and
non-invasively evaluated. These enzymes, even though their natural substrates are mostly
of aliphatic nature, hydrolyze phenolic sulfates to phenol and sulfuric acid. The availability
of [124I]iodinated substrates is expected to provide a PET-based method for measuring their
activity in vivo. The currently available methods of synthesis of iodinated arylsulfates
usually require either introducing of a protected sulfate ester early in the synthesis or
introduction of sulfate group at the end of synthesis in a separate step. The described
method gives the desired product in one step from an aryl-alkyl cyclic sulfate. When
treated with iodide, the source cyclic sulfate opens with substitution of iodide at the alkyl
center and gives the desired arylsulfate monoester.
Keywords: iodine; cyclic sulfate; Positron Emission Tomography; 124I; enzyme substrate

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1. Introduction
Arylsulfatases [1] are a group of lysosomal enzymes responsible for hydrolyzing the sulfate ester
group esters in various biological substrates. Substrates for arylsulfatases can be as simple as steroid
sulfates and as complex as glycosaminoglycans [1,2]. Deficiency or reduced activity of these enzymes
lead to the accumulation of sulfated molecules involved in important cellular functions involved in
hormone regulation, degradation, and signaling pathways [3–5].
Development of enzyme replacement therapies is presently hindered by the lack of suitable
non-invasive methods for assessment of the activity of the enzyme of interest in the organs and tissues.
Recently, we conducted several preclinical studies where radiolabeled enzyme molecules were studied
by Positron Emission Tomography (PET) to determine the kinetics of their transfer from the injection
point to the organs of interest [6,7]. Such studies give quantitative information on the delivery of the
enzyme molecules to the affected tissues and the initial deposition [8], but they don’t provide
information on the enzyme activity in these tissues. Development of non-invasive methods suitable for
direct measurement of enzyme activity in preclinical models, as well as in a clinical setting, would
shorten the drug development cycle and improve the assessment of the efficacy of the therapy.
We have investigated the pharmacokinetics of arylsulfatase A (ARSA) in rodents [7] and non-human
primates [9], which provided the pharmacokinetics data necessary for estimating the initial enzyme
deposition in the tissue as a function of administration modality. Presently, we are developing a PET-based
method enabling the assessment of enzyme activity, rather than measuring the amount of protein. The
method will utilize arylsulfatase substrates modified with a label that is detectable by imaging and,
upon enzymatic reaction, giving products that are retained in the tissues where the reaction has
occurred longer than the respective substrates would be retained. The products of the enzymatic
reaction will thus “label” the tissues where the enzyme activity is present. Considering the quantitative
character of PET, we focus on substrates labeled with positron emitting 124I. Iodine-124 is used here as
a positron emitter with a physical half-life of 4.2 days, which enables non-invasive quantitative
tracking of the administered radioactivity in vivo for several days [10]. The above approach suggests
developing a method for synthesizing radiolabeled arylsulfatase substrates that undergo enzymatic
hydrolysis without affecting the enzyme activity.
The natural substrates for ARSA are sulfosphingolipids [11] (Figure 1, 1). Such compounds form
micelles and thus are unlikely suitable as molecular probes for arylsulfatase activity in vivo because of
their incorporation in the lipid compartment in vivo, which is not compatible with the proposed
imaging approach. Synthetic substrates, e.g., 4-nitrocatechol sulfate and 4-methylumbelliferyl sulfate
(Figure 1, 2 and 3), are known to be hydrolyzed by ARSA (as well as other arylsulfatases) under
physiological conditions. They give colored and fluorescent products upon desulfation [12],
respectively, which enables their use in in vitro assays of arylsulfatase activity. While our approach
suggests using similar 124I-labeled structures that can be hydrolyzed by arylsulfatases, the modality of
their in vivo use suggests designing molecules which, after desulfation, can bind tissue components
(e.g., via hydrophobic interactions) and thus, “label” the tissues with 124I.

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Figure 1. Natural and known phenolic substrates of ARSA.
One of the major hurdles in designing compounds suitable for use as substrates for arylsulfatases
is the introduction of the sulfate ester. This group is sensitive to acidic conditions and is known to
undergo nucleophilic substitution when present on an alkyl scaffold. The currently available
procedures usually utilize either the introduction of sulfate ester at the end of the synthetic sequence, or
introducing a protected sulfate ester at the beginning, followed by removal of the protection at the end
of the sequence.
Although these methods are attractive for the synthesis of sulfate esters of stable compounds, they
are not optimal for radiochemical syntheses. The latter require handling of the radiolabeled precursors
through multiple chemical transformations, which not only generates radioactive waste but also leads
to the loss of radioactivity due to the radionuclide decay. For the PET imaging applications,
radiolabeling should be, ideally, the last stage of the process so that the radiolabeled product would be
used for the imaging immediately after the labeling and fast purification. Here, we report a new
method for synthesis of radioiodinated arylsulfatase substrates, where the introduction of the radiolabel
(124I) and formation of the sulfate ester are achieved in one step.
2. Results and Discussion
Cyclic sulfates [13] are diesters of sulfuric acid, commonly used in organic chemistry for mono
functionalization of vicinal diols. They are better leaving groups than the more commonly used leaving
groups tosylates and mesylates, and are fairly stable compounds [14]. We speculated that, if we could
prepare an asymmetrical cyclic sulfate involving a phenol and an aliphatic alcohol, a nucleophilic
reagent would react with the aliphatic side of the cyclic sulfate and generate a phenolic sulfate ester on
the other side.
We tested our hypothesis using a commercially available 2-hydroxyphenethyl alcohol (4), which,
when treated with thionyl chloride and pyridine at 0 °C in anhydrous dichloromethane, gave cyclic
sulfite 5 in excellent yield. This cyclic sulfite was then oxidized to the desired cyclic sulfate 6
quantitatively using catalytic RuCl3 and sodium periodate. Treatment of the cyclic sulfate with sodium
iodide in DMF gave the sulfate ester 7 in quantitative yield, as shown in Scheme 1. We note that
primary iodides such as the model compound (7) may be insufficiently stable in vivo for imaging
purposes. The method, however, is intended for developing secondary iodides (as described below)
and, eventually, [18F]fluorides that are significantly more stable.

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Scheme 1. Synthesis and ring opening of cyclic sulfate 6.
To extend the versatility of this method, we synthesized a cyclic sulfate with a long aliphatic side
chain as shown Scheme 2. The synthesis commenced from the amide 8, synthesized in approximately
50% yield according to a known three-step procedure [15] from commercially available 2-hydroxy-
phenylacetic acid. The amide 8 was then treated with the commercially available octylmagnesium
bromide solution to give the ketone derivative 9. Ketone 9 was reduced to alcohol 10 with sodium
borohydride. The subsequent removal of benzyl protection, using catalytic 10% Pd on C under a
hydrogen balloon, gave the substituted 2-hydroxyphenethyl alcohol 11. This compound was converted
to the desired cyclic sulfate 13 under the same conditions as described in Scheme 1.
Scheme 2. Synthesis and ring opening of cyclic sulfate 13.
Labeling of 6 with [124I]NaI was accomplished using the same procedure as described in Schemes 1.
The [124I] sodium iodide for this purpose was obtained from IBA Molecular (Richmond, VA, USA) as
a carrier free aqueous solution in 0.02 M NaOH, 20–50 mCi/mL. Water was removed by azeotroping
with anhydrous acetonitrile under an argon stream.
After the removal of water, a solution of cyclic sulfate in anhydrous DMF (100 μL, 1 mg/mL to 0.5 μCi
of NaI) was added to the solid [124I] sodium iodide and kept at ambient temperature for about 30 min.
The resulting product was then purified by reverse phase HPLC using UV and gamma radiation
detector. From our studies on 6, we found that the presence of significant amounts of water drastically
slows down the reaction.

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During the course of our studies on ring opening of cyclic sulfate with NaI, we found that adding an
aqueous solution of NaI to a DMF solution of the cyclic sulfate led to a significant slowdown of the
reaction; the latter was not complete after 24 h at ambient temperature. Similarly, the reaction was
slowed significantly when carried out in the polar protic solvents, e.g., ethanol. In acetone, the reaction
led to the formation of an uncharacterized non-polar product. Thus, without the need to be anhydrous,
DMF was found to be the best solvent for the reaction.
This methodology describes the synthesis of new aryl-alkyl cyclic sulfate, which can serve as
intermediates in organic synthesis. The method described here offers another significant advantage for
synthesizing substrates for arylsulfatase A because it is applicable not only to 124I, but it can also be
easily adopted by other PET-compatible radionuclides, such as F18 and C11.
3. Experimental
General
All reactions were performed under an atmosphere of argon in oven dried glassware using
anhydrous solvents on 4 Å molecular sieves obtained from Acros Organics/Thermo Fisher Scientific
(Pittsburgh, PA, USA). Solvents were always handled under argon atmosphere. The ambient
temperature was set at 21 °C.
Yields refer to chromatographically and spectroscopically homogenous materials, unless otherwise
stated. Reactions were monitored by thin-layer chromatography (TLC) carried out on 0.25 mm E.
Merck silica gel plates (60F-254), using UV light as a visualizing agent, and an aqueous solution of
basic potassium permanganate and heat as developing agents. Purifications were done on a Biotage
flash purification system using pre-packed Biotage SNAP columns.
1
Products were characterized by H-NMR/13C-NMR on either a Varian 500 MHz, or 400 MHz
spectrometer in a deuterated solvent using tetramethylsilane (TMS) as an internal standard. Chemical
shifts are given in δ (ppm) and J-values in Hz. High-pressure liquid chromatography (HPLC) was
performed using Waters 1525 binary HPLC pump and Waters 2489 UV/Visible detector operating at
254 nm on a Waters symmetry C18 3.5 μm, 4.6/75 mm column. MeOH HPLC solvent was obtained
from Sigma-Aldrich (St. Louis, MO, USA).
4,5-Dihydrobenzo[d][1,3,2]dioxathiepine 2,2-dioxide (6): Anhydrous pyridine (3.4 mL, 41.62 mmol)
was added to a solution of the 2-phenethanol 4 (2.3 g, 16.65 mmol) in anhydrous DCM (70 mL), and the
mixture was cooled to 0 °C in an ice bath. Then, freshly distilled thionyl chloride (1.22 mL, 16.65 mmol)
was added to the above solution. The resultant reaction mixture was then stirred for 2 h with gradual
warming to the ambient temperature. At this point, TLC of the reaction mixture indicated complete
consumption of the starting material. The reaction mixture was then diluted with more DCM,
transferred to a separatory funnel, and washed with cold dilute HCl twice. Then the DCM layer was
washed with water (2×) and brine (1×), and, finally, the DCM layer was dried with anhydrous sodium
sulfate and concentrated to give the cyclic sulfite 5, 2.8 g, as an oily liquid (Rf: 0.58; 20%
EtOAc/hexane). The product was used in the subsequent step without purification.
Catalytic ruthenium chloride (0.02 mol %, 63 mg) was added to the crude cyclic sulfite 5 (2.8 g,
15.2 mmol), in acetonitrile (40 mL) and DCM (40 mL). Then, a solution of sodium periodate (4.9 g,

Molecules 2012, 17
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22.8 mmol) in water (60 mL) was slowly added dropwise to the above solution at room ambient
temperature. The reaction mixture was then stirred at room temperature until all of the starting material
was consumed (about 1 h). All of the volatile solvents were removed in vacuo and the aqueous layer
was extracted with EtOAc (3×). The combined ethyl acetate layer was washed with saturated sodium
bicarbonate solution (2×), water (2×), and brine (1×). Finally, the organic layer was dried over
anhydrous sodium sulfate, concentrated, and chromatographed (hexane/EtOAc 5–50%) over silica gel
to give pure cyclic sulfate (6), 2.9 g, 87%, as white solid. Rf: 0.28 (EtOAc/hexane). 1H-NMR (CDCl3;
500 MHz) δ 7.35 (t, 7.5 Hz, 1 H), 7.26 (dd, 13.5 Hz, 7,5 Hz, 2 H), 7.19 (d, 7.5 Hz, 1 H), 4.64 (m, 2 H),
13
3.26 (m, 2 H); C-NMR (CDCl3, 125 MHz) δ 149.9, 131.1, 130.2, 129.5, 127.7, 121.9, 72.6, 33.6;
HPLC: RT 9.1 min on Symmetry C18 column from Waters, flow 1 mL/min, linear gradient 5 to 95%
MeOH/Water.
Sodium 2-(2-Iodoethyl)phenyl sulfate (7): Solid sodium iodide (13 mg, 0.088 mmol) was added to a
solution of cyclic sulfate 6 (17.5 mg, 0.088 mmol) in the DMF (0.35 mL) at ambient temperature. The
reaction was monitored by the TLC to document the consumption of the starting material. Within
15 min, all of the starting material was consumed. The DMF was removed in vacuo and the residue
was dried thoroughly in vacuo at ambient temperature to give the pure product 7, yield 100%;
1H-NMR (DMF-d7, 400 MHz) δ7.21 (m, 3 H), δ 7.01 (dt, 1 Hz, 8 Hz, 1 H), 3.55 (m, 2 H), 3.26 (m, 2 H);
13C-NMR (100 MHz, DMF-d7) δ152.2, 133.2, 130.3, 127.7, 123.6, 122.3, 35.9, 6.5; HPLC: RT 6.1 min
on Symmetry C18 column from Waters, flow 1 mL/min, linear gradient 5 to 95% MeOH/Water.
1-(2-(Benzyloxy)phenyl)decan-2-one (9): A solution of amide 8 (570 mg, 2 mmol) in anhydrous THF
(8 mL) was cooled to −78 °C in a dry ice-acetone bath under an argon atmosphere. To this cooled
solution, a solution of octylmagnesium bromide (2 M, 1.25 mL, 2.5 mmol) in ether was added. After
the addition was complete, the reaction was allowed to proceed for 2 h with gradual warming to the
ambient temperature. The reaction was quenched by the addition of a saturated ammonium chloride
solution and the THF was removed in vacuo. The reaction mixture was transferred to a separatory
funnel and extracted with diethyl ether (3×). The combined organic layer was washed with water (1×)
and brine (1×), dried over anhydrous sodium sulfate, concentrated and chromatographed on silica gel
1
to give pure ketone 9, 415 mg, 61%, as viscous oil. Rf: 0.77 (15% EtOAc in hexane); H-NMR
(CDCl3, 500 MHz) δ 7.39 (m, 5 H), 7.34 (m, 1 H), 7.25 (m, 1 H), 7.16 (m, 1 H), 6.96 (m, 1 H), 5.06
(s, 2 H), 3.71 (s, 2 H), 2.4 (t, 7 Hz, 2 H), 1.52 (m, 2 H), 1.23 (bm, 10 H), 0.89 (t, 7.5 Hz, 3 H); 13C-NMR
(125 MHz, CDCl3) δ 206.8, 156.4, 136.9, 131.3, 128.5, 128.3, 127.8, 127.3, 124.1, 120.8, 111.6, 69.9,
44.7, 42.2, 31.8, 29.3, 29.1, 29.09, 23.7, 22.6, 14.0.
1-(2-(Benzyloxy)phenyl)decan-2-ol (10): To a solution of ketone 9 (415 mg, 1.23 mmol) in absolute
ethanol (5 mL), solid sodium borohydride (93 mg, 2.46 mmol) was added at ambient temperature. The
reaction mixture was stirred at ambient temperature for 30 min. At this point, the TLC indicated
complete consumption of the starting material, and the reaction was quenched by the addition of a
saturated ammonium chloride solution. All of the ethanol was removed in vacuo, and the residue was
transferred to a separatory funnel and extracted with EtOAc (3×). The combined organic layer was
washed with water (2×) and brine (1×). Finally, the organic layer was dried over anhydrous sodium

Molecules 2012, 17
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sulfate, concentrated, and chromatographed on silica gel to give pure alcohol 10, 393 mg, 94%, as
viscous oil. Rf: 0.48 (10% EtOAc in hexane); 1H-NMR (500 MHz, CDCl3) δ 7.41 (m, 5 H), 7.20 (m, 2 H),
6,94 (m, 2 H), 5.09 (s, 2 H), 3.89 (bs, 1 H), 3.00 (dd, 4 Hz, 14 Hz, 1 H), 2.68 (dd, 8 Hz, 14 Hz, 1 H),
13
1.93 (bs, 1 H), 1.50 (m, 2 H), 1.27 (bm, 12 H), 0.89 (t, 7 Hz, 3 H); C-NMR (125 MHz, CDCl3) δ
156.7, 136.9, 131.5, 128.5, 127.9, 127.7, 127.5, 127.2, 120.9, 111.7, 71.8, 70.0, 38.8, 37.2, 31.8, 29.7,
29.6, 29.2, 25.7, 22.6, 14.1.
2-(2-Hydroxydecyl)phenol (11): Catalytic 10% palladium on carbon (20 mg) was added to a solution
of alcohol 10 (393 mg, 1.15 mmol) in EtOAc (4 mL). Then the reaction flask was purged with
hydrogen (by applying vacuum and flushing hydrogen gas). This process was repeated three times, and
then the reaction proceeded overnight under a balloon of hydrogen gas. The reaction mixture was then
filtered through a pad of Celite, and the Celite pad was washed thoroughly with EtOAc. Finally,
EtOAc was removed in vacuo to give the desired product 11—a pale yellow oil (285 mg, 99%). Thus,
the product obtained was sufficiently pure for use in the next step; an analytically pure sample for
NMR was obtained by chromatographing a small sample on silica gel. Rf: 0.45 (25% EtOAc in
hexane); 1H-NMR (500 MHz, CDCl3) δ 8.19 (bs, 1 H), 7.15 (m, 1 H), 7.02 (m, 1 H), 6.91 (m, 1 H), 6.84
(m, 1 H), 4.00 (m, 1 H), 2.85 (dd, 2 Hz, 15 Hz, 1 H), 2.80 (dd, 7 Hz, 15 Hz, 1 H), 2.39 (bs, 1 H), 1.53
13
(m, 1 H), 1.28 (m, 12 H), 0.88 (t, 7 Hz, 3 H); C-NMR (125 MHz, CDCl3) δ 155.9, 131.5, 128.3,
125.9, 120.2, 117.2, 74.6, 40.3, 38.9, 36.9, 31.8, 29.5, 29.4, 25.6, 22.6, 14.1.
4-Octyl-4,5-dihydrobenzo[d][1,3,2]dioxathiepine 2,2-dioxide (13): Anhydrous pyridine (330 μL, 3.6
mmol) was added to a solution of the substituted 2-phenethanol 11 (340 mg, 1.44 mmol) in anhydrous
DCM (6 mL), and the mixture was cooled to 0 °C in an ice bath. Then, freshly distilled thionyl chloride
(115 μL, 1.58 mmol) was added to this solution. The reaction mixture was then removed from the ice
bath and stirred for 2 h while warming up to the ambient temperature. At this point, the TLC of the
reaction mixture indicated the complete consumption of the starting material. The reaction mixture was
diluted with more DCM, transferred to separatory funnel, and washed with cold dilute HCl (2×). Then,
the DCM layer was washed with water (2×) and brine (1×). Finally, the DCM layer was dried over
anhydrous sodium sulfate and concentrated to give the cyclic sulfite 12 (360 mg, 87%) as an oily
liquid. The product was then used in the next step without purification. The product formed two spots
on the TLC (Rf: 0.80 & 0.72; 10% EtOAc/hexane) due to the formation of diastereomers.
Catalytic ruthenium chloride (0.02 mol %; 5 mg) was added to the solution of the crude cyclic
sulfite 12 (350 mg, 1.20 mmol) in acetonitrile (4 mL) and DCM (4 mL). Then, a solution of sodium
periodate (380 mg, 1.77 mmol) in water (6 mL) was added to this solution dropwise at ambient
temperature. The reaction mixture was then stirred at ambient temperature until the starting material
was consumed (about 1 h). All of the volatile solvents were removed in vacuo and the aqueous layer
was extracted with EtOAc (3×). The combined ethyl acetate layer was washed with saturated sodium
bicarbonate solution (2×), water (2×), and brine (1×). Finally, the organic layer was dried over
anhydrous sodium sulfate, concentrated, and chromatographed (hexane/EtOAc 5–15%) on silica gel to
give pure cyclic sulfate 13 (288 mg, 78%), as a pale yellow, viscous oil. Rf: 0.43 (10% EtOAc in
1
hexane); H-NMR (500 MHz, CDCl3) δ 7.34 (m, 1 H), 7.25 (m, 2 H), 7.16 (m, 1 H), 4.80 (m, 1 H),
3.41 (dd, 9.5 Hz, 16 Hz, 1 H), 2.90 (d, 16 Hz, 1 H), 1.89 (m, 1 H), 1.73 (m, 1 H), 1.29 (m, 12 H), 0.88

Molecules 2012, 17
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13
(t, 7 Hz, 3 H); C-NMR (125 MHz, CDCl3) δ 150. 0, 131.2, 129.4, 127.6, 126.5, 86.2, 40.3, 38.9,
35.0, 31.8, 29.1, 29.0, 24.9, 22.6, 14.1; HPLC: RT 9.4 min, flow 1 mL/min, gradient 10 to 100%
MeOH/Water.
Sodium 2-(2-iododecyl)phenyl sulfate: Solid sodium iodide (15 mg, 0.1 mmol) was added to a solution
of cyclic sulfate (13, 26 mg, 0.083 mmol) in the DMF (0.5 mL) at ambient temperature. The reaction
was monitored by TLC to document the consumption of the starting material. Within 15 min, the latter
was consumed, and the DMF was removed in vacuo. The residue was dried thoroughly in vacuo at
1
ambient temperature to give a pure product (yield 100%); H-NMR (DMF-d7, 400 MHz) δ 7.56 (dd,
1.2 Hz, 8 Hz, 1 H), 7.20 (m, 2 H), 7.00 (dt, 1.2 Hz, 8 Hz, 1 H), 3.79 (bs, 1 H), 3.40 (dd, 7 Hz, 14 Hz, 1 H),
3.33 (dd, 8 Hz, 14 Hz, 1 H), 1.73 (m, 2 H), 1.26 (m, 12 H), 0.86 (t, 6.8 Hz, 3 H); 13C-NMR (DMF-d7,
100 MHz) δ 153.1, 132.5, 131.0, 127.7, 123.3, 122.1, 43.4, 40.9, 40.2, 32.2, 30.0, 29.8, 29.6, 29.1,
22.9, 14.2; HPLC: RT 8.1 min, flow 1 mL/min, gradient 10 to 100% MeOH/Water.
Procedure for Labeling 6 with 124I: 200 μL of anhydrous MeCN were added to a solution of [124I]NaI
(500 μCi) in 15 μL 0.02 M NaOH, and the solvents were evaporated under steady stream of argon.
After the solvent was completely removed, an extra 100 μL of MeCN was added, and the solvent was
removed again, as above. This procedure was repeated two more times to remove most of the water
from [124I]NaI. Then, a solution of cyclic sulfate (100 μL, ~1 mg/mL) in DMF was added to the
residue, and the iodination reaction was allowed to proceed for 30 min. Then, the reaction mixture was
analyzed by HPLC using dual detection (UV and gamma) to distinguish the desired product from the
unreacted [124I]iodide. Under the HPLC conditions (see above), the [124I]iodide had an RT of 1 min and
the desired compound eluted at 7.3 min, as observed on gamma-detector. Retention times for the
labeled compound on gamma-detector (7.3 min), and unlabeled compound on UV detector (6.1 min),
are different because the gamma-detector was externally connected.
4. Conclusions
We developed a very effective method for labeling synthetic arylsulfatase substrates with 124I. In
this method, the steps of introducing the sulfate ester and radioactive iodine converge into one stage.
This not only simplifies the synthesis, but also reduces the time between the radionuclide production
and use, which preserves the radioactivity. We expect that the method can be easily adopted for
labeling arylsulfates with other radionuclides suitable for PET, such as F18 and C11.
Supplementary Materials
Supplementary materials can be accessed at: http://www.mdpi.com/1420-3049/17/11/13266/s1.
Acknowledgments
This research was supported by Grant 85310 from Shriners Hospitals for Children and a grant from
Shire HGT. Authors thank M. Gagne for logistical support and E. Papisov for assistance with editing.

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Sample Availability: Samples of Compounds are available upon request.
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