Clinical studies in nuclear medicine require an ever-increasing number of isotopically labeled compounds and preparationson a routine basis. These compounds are often labeled with cyclotron-produced short-lived radionuclides and, in conjunction withpositron computerized tomography (PCT), are used as tracers forestablishing the state ofvarious metabolic processes. The techniquedeveloped by Sokoloff et al. (I) permitted the autoradiographicmeasurement of local organ glucose metabolism in animals using[1-'4C]2-deoxy-D-glucose, and the desire to apply its basic principles to the in vivo, noninvasive determination of local organglucose metabolism in man by means of PCT has led to the synthesis of the fluorinated derivative 2-['8Fjfluoro-2 deoxy-D-glucose(2—5).Whilethis2-FDG(F-18)isnowroutinelyavailable(6,7)and studies utilizing it have yielded remarkable results (8,9), therelatively long half-life of fluorine-I 8 ( 109.72 mm) renders it inconvenient for serial studies. For this reason the labeling of 2-DGwith carbon-I I (T112 20.38 mm) increases the serial capabilitiesof this technique. The availability of [I-11C]2-deoxy-D-glucose([I-' ‘CJ2-DG)would allow the investigation of the transport andphosphorylation properties of these two glucose analogs in man(10). In addition, carbon-I I can be prepared efficiently in cyclotrons with low proton energy (e.g., 8 MeV) (1 1), in contrast toF-I8-labeled fluorine, obtained by the 20Ne(d,a)'8F nuclear reaction, necessary at the present time for the preparation of 2-FDG(F-I 8).

Received Feb. 3, 1982; revision accepted Mar. 29, 1982.For reprints contact: J. R. Barrio, PhD, UCLA School of Medicine,

Laboratory of Nuclear Medicine, Divisionof Biophysics,Los Angeles,CA 90024.

The actual synthesis of this and other short-lived radiolabeledcompounds that are to be used in a clinical nuclear medicine settingpresents unique problems from the standpoint of the radiochemist.In no other situation is the same combination of requirements tobe found: the process must be fast owing to the short half-lives ofthe radionuclides involved; it must also accommodate the sometimes complex nature of the procedures used, i.e., multistep organicsynthesis; and all of this is to be achieved safely in spite of thenecessarily high initial levels of radioactivity. This means that allsteps in the entire process, from cyclotron bombardment to sterilization, must be accomplished behind shielding adequate to resultin little or no exposure to the personnel involved.

We now report the successful application ofthe unit-operationsapproach to the design of remote semiautomated chemical processing systems (12). These are characterized by their simplicity,flexibility, reliability, and practicality. The approach has been usedsuccessfully to construct systems for the preparation of 2-[‘8F]2-fluoro-2-deoxy-D-glucose (6), [1-' ‘CJ2-deoxy-D-glucose(13,14), [1-―C]palmiticacid ([I-―C]PA)(15), and L-aminoacids tagged with carbon-Il and nitrogen-l3 (16—18).Here weillustrate its application by describing the construction and functionof the system designed for the preparation of sterile, pyrogen-free[I@1‘C]2-DGfor routine clinical studiesusing PCT.

MATERIALS AND METHODS

I1_HCj2@deoxy-D@giucose:synthesis. [‘‘C]Hydrogencyanide(21 ,22) was condensed on a DMCS-treated glass coil treated andcooled to —23°C(CC14-dry ice) (23); then, after warming, it wasswept with nitrogen into a solution ofsodium cyanide (4.9 mg, 0.1

Volume 23, Number 8 739

The Unit OperationsApproach Applied to the Synthesisof [1@11C]2-Deoxy-D--

Glucosefor RoutineClinical Applications

H. C. Padgett, J. R. Barrio, N. S. MacDonald, and M. E. Phelps

University of California, Los Angeles and School of Medicine and Laboratory of Biomedicaland EnvironmentalSciences, Los Angeles, California

An approachto the designof remotesemiautomatedchemicalprocessingsystemshasbeendevelopedandtestedin morethan750 productionruns.Thisdesignstresses the idea of unit operations, which allows maximum fiexiblifty while maintamingthe highestpossiblestandardsregardingsafety, reliability,efficiency,andpracticality.Applicationof thisapproachhasresultedinthe developmentof a simpie andreliableremotesemiautomatedsynthesissystemforthe routineproductionof [1-11cJ2-deoxy-c-glucose.Useofthissystemresultsinthe preparationofa sterlie, pyrogen-freeproductsuitablefor humaninjectionafter a synthesistime of 50mm, with radiochemicalpurityof greater than 98% and yields (25—30%)permittingquantitativemeasurementsusingpositroncomputedtomography.

J Nucl Med 23: 739—744, 1982

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PADGETr, BARRIO, MAcDONALD,AND PHELPS

FIG. 1. System for synthesis of [1-11C]2-deoxy-i>glucose:A. Displacement reaction; B. solvent drying; C. desiccant; D. reduction;E. solventevaporation;F.silicagelcolumn;G.flow-throughgammadetector;H. hycfrolysis;J.purificationcolumn;K.sodiumchloridevial; L. sterilizationfilter; M.productvial.

_,+._ = three-way solenoid valve

@t::@—= syringeonexternalaccesstubing

with silica gel and equilibrated with the same solvent). The columnwas developed using petroleum ether/methyl acetate (7:3); thefirst 6 ml were directed into a waste container, and the protectedlabeled aldehyde (4) eluted in the subsequent 4 ml. The presenceofcarbon-li activity(asdeterminedbythe flow-throughgammadetector G) signaled the diversion of the desired product to flaskH, which was preheated to 80-85°C.The solvent was evaporatedand a suspension of cation-exchange resin (AG 50W-X8, 50—100mesh, hydrogen form, 1.0 g in 4.0 ml sterile water) was added tothe residue. After heating for 10 mm, the aqueous solution contaming [1-' 1C]2-deoxy-D-glucose (5) was transferred to the topofcolumn J (16.5 X 0.7 cm i.d.), which had been packed successively with ion-retardation resin (AG llA8, 50—100mesh, 5.5 cm),neutral alumina (5.5 cm), and ion-retardation resin (5.5 cm) andequilibrated with sterile water. The product was eluted with sterilewater (total volume = 10 ml) into a vial (K) containing sodiumchloride(90mg).Theresultingisotonicsolutionwassterilizedbypassage through membrane filter L (0.22 zm pore size) directlyinto sterile serum vial M. At this point, the pyrogen-free solutionwas ready for injection. Thin-layer radiochromatography showedthat the product El-' ‘C]2-DG(25—30mCi) had the same retentionfactor as [1-'4CJ2-DG (Rf = 0.30, silica gel, acetonitrile / H2O[95:5]), and was found to be >98% pure. By the use ofthe remotesemiautomated system described in this paper, each run resultedin radiation exposure of <1 mR to the radiochemist.

Unequivocal identification of the [1-' ‘C12-DClproduced wasdemonstrated by the efficient enzymatic conversion of [1-1‘C]2-DG to [1-' ‘C]2-DG-6-phosphate using yeast hexokinase

(26,27). Results of the conversion were monitored by HPLC(Ultrasil-NH2, 23-cm column,* 55%aqueous potassium phosphate[100 mM, pH 7.0], 45% methanol; flow rate 1.0 ml/min). Theprocess was tested by converting El-―C]2-DG(retention time 3.8mm) to [l-'4CJ2-DG-6-phosphate (retention time 8.1 mm).

To prepare [1-'‘C]2-DG-6-phosphate,yeast hexokinase(66units) was added to the reaction mixture (1.0 ml) containingTris-HC1buffer (100 mM, pH 8.0), magnesiumchloride(iOmM), adenosine triphosphate (10 mM), and El-' ‘C]2-DG(100zCi),andthe resultingsolutionwasleftat roomtemperaturefor30mm.Perchloricacid(4.7%,0.1ml) wasadded,followedbycentrifugation to precipitate and separate the protein. The supernatant was brought to pH 7.0 with 8M NaOH, and a sample(250 @l)wasinjected into the chromatograph. Carbon-l 1activityin the effluent was monitored by the flow-through radioactivitydetector.

The synthesis system: construction and use. The remote scm

-.4- = two-way solenoid valve

-4 = vent (open)

mmol) in N,N-dimethylformamide (DMF) (2.0 ml) contained ina bubbler-tube apparatus. After the trapping was completed, thesolution was transferred to reaction flask A (Fig. 1). To this wasadded a solution of i-O-trifluoromethanesulfonyl-2,3:4,5-di-O-isopropylidine-D-arabitol (2, Scheme 1) (0.36 g, 1.0 mmol) inDMF (1.0 ml) (24). The reaction mixture was agitated for 1 mmand then quenched by the addition of cold (5 °C)15% aqueoussodium chloride (7.5 ml) and diethyl ether (3.0 ml). After mixingand phase separation, the ethereal solution containing the desired[1-' ‘C]2-deoxy-3,4:5,6-di-O-isopropylidine-D-glucononitrile (3)was transferred to flask B and mixed with saturated aqueous sodium chloride (10 ml). The phases were separated and the etherealsolution was passed through a column (10 X 1.5 cm i.d.) containinganhydrous magnesiumsulfate (3.0 cm) and calcium sulfate (3.0cm)andintoa dry flask(D)cooledto 5 °C.Diisobutylaluminumhydride (25) (20% in hexane, 2.0 ml, 2.8 mmol) was added to flaskD, the mixture stirredfor 3 mm at 5 °C,and the reactionquenchedby the addition of 1%sulfuric acid/ethanol (1:1, 6.0 ml). Afterstirring for 10 mm, the organic phase, containing the resultingEl-I ‘C]2-deoxy-3,4:5,6-di-O-isopropylidine-D-glucose(4) alongwith trifluoromethanesulfonate (triflate) 2 and alcohol I, wastransferred to empty heated flask E, and, after solvent evaporationand removal of the hot bath, the residue was taken up in petroleumether 35—60°C/methyl acetate (7:3; 1.5 ml). The resulting solution was applied to thetop of the column F (20 X 0.7 cm, packed

3

CH2OH

H@ HO.v.@@'@@lIc/―

‘@0H

5

Scheme1. Synthesisof E1-11C12-deoxy-D-glucose.

740 THE JOURNAL OF NUCLEAR MEDICINE

CH2OH

2 0‘cH2-0S02CF3

@ ,@ -

2O@

@:cH2-―cN@Z[_j@'@

CH2O ‘@

1 2

CH 2@ 1CHO

IH]0

L@,

Fo@,

cH2O―\

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TECHNICAL NOTES

iautomated production system designed for the synthesis of [1-I ‘C]2-deoxy-D-glucose is shown schematically in Fig. 1 . The

synthetic process can be divided into five basic steps: (a) nitrileformation, (b) nitrile to aldehyde reduction, (c) aldehyde isolation,(d)deprotection(hydrolysis),and (e)sterilization.Thissyntheticprocedure is carried out using the five-unit system as shown.

The system consists principally of five flasks, two chromatography columns, a flow-through gamma detector, and a sterilizationunit, all connectedthrough 13two-and three-wayTeflon solenoidvalves and appropriate high-pressure Teflon tubing and fittingsto facilitatetheadditionofreagentsandsolventsandthedesiredmanipulation of the reaction mixture from one unit of the systemto the next.Theseitemsare accommodatedon a supportconstructed from standard 1.25-cm solid aluminum rods held togetherwith lattice connectors. The size of the system (50 X 30 X 45 cm)was dictated by the space required for the two vortex mixers, themagnetic stirrer, and the oil baths. The solenoid valves were secured to aluminum plates and mounted on the support rods. Theflasks and columns were mounted using standard equipmentclamps.Theactual transfersof solutionare achievedby the useofvacuumor pressurealongwithremoteoperationof the solenoidvalves.

Thefirstunit,wherethenucleophilicdisplacementbycyanideionoccurs,consistsof twoflasks,Aand B,and fourtwo-waysolenoid valves. These flasks are 25- X 150-mm culture tubes withground-glass joints (1 24/25) added. The flasks were sealed withinvertedserum stoppers(24 mm) through whichhad been insertedlengthsofTeflontubing.FlaskA hasfourtubes:one(3.0mmo.d.)to serve as a vent, one each for liquid inlet and outlet, and an cxtemalaccessfortheapplicationofvacuumand/or fortheadditionof reactants or solvents.The last three are I .5-mm o.d. tubes tominimize solution holdup. During the synthesis these lines areroutinelyflushedwithadditionalsmallamountsofsolventto remove residual traces of desired product. All the external accesstubesterminatewitha Luersyringefittingequippedwitha plasticthree-waystopcock.tFlaskBhasthreetubings(1.5mmo.d.),oneeach for the reaction mixture inlet and outlet, and an externalaccess. Flask A is empty and dry at the outset ofthe synthesis andultimately contains the reaction product, radiolabeled nitrile (3),and the excessofstarting materialsin DMF, diethylether, and 15%aqueous sodium chloride. Flask B initially contains saturatedaqueous sodium chloride to remove nearly all of the water and allof the residual DMF from the ethereal solution transferred fromflaskA.Here,inbothflasks,wheretheproduct-containingsolutionisobtained after an aqueous-organic solvent (lighter than water)extraction, the outlet tubing wasextended into the flasksuch that,under the conditions of use, the surface of the resulting loweraqueouslayerisjustbelowtheendofthistube.Thus,whenvacuumis applied through this tube (i.e., at flask Bor column C) only theupperorganiclayerwillbe transferred.BothflasksA and Brestontheheadsofvortexmixers,whichprovidetheagitationrequiredto mixtheorganicreactionmixturequicklyandthoroughlywjththe aqueoussolutions.

Thesecondunit,wherethereductionofthe carbon-II-labelednitrile (3) to the aldehyde (4) takes place, is composed of columnC, flask D, and a two-way solenoid valve. The top of the column,which contains the desiccant, was sealed with a silicone stopperthroughwhichhadbeeninsertedtwolengthsofTeflontubing,onefor the reaction-mixture inlet (1.5 mm o.d.), and the other forexternal access (3.0 mm o.d.) for the application of vacuum andpressure. This and all the columns function as follows. The reactionmixture (the diethyl ether solution in flask B) is introduced intocolumnC bycreating a slightvacuum in the columnusinga 60-nildisposable syringe connected to the external access tubing. Thesolenoid valve at the bottom of the column is closed and the valvein the inlet line is opened. After the transfer is complete, the valvepositions are reversed and mild pressure is applied through the

externaltubingusingthe samesyringe.This forcesthe solutionthroughthe columnand into flaskD, whichis a 25 X 150-mmculture tube with a ground-glassjoint (1 24/25) added. The flaskwas sealed with an invertedserum stopper (24 mm) through whichhad beeninsertedfourlengthsof Teflontubing:oneeachfor thereaction-mixture inlet, its outlet, and an external access (all 1.5mm o.d.) for the addition of first the reducing agent (diisobutylaluminum hydride) and then the quenching solution (1% sulfuricacid)/ethanol, 1:1), and a vent (3.0 mm o.d.). The reaction-mixtureinlet is connected to the bottom ofcolumn C through the two-waysolenoidvalve.The outlet tubing was extended into the flasksuchthat, undertheconditionsofuse(i.e.,afterthereductionreactionhas been quenched), only the organic solution (i.e., the upper layer)will be removed and transferred to flask E by the application ofvacuum at the terminus of this tube (i.e., flask E). Before the startofthesynthesis,thisdryflask(D),containinga magneticstirbar,wasflushedwithnitrogenandplacedinan ice-waterbathoveramagnetic stirrer. (See [1-' ‘C]2-Deoxy-D-Glucose:Synthesis)

Thethirdunit,wheretheprotectedEl-'‘C]2-DG(4) isisolated,consists of flask E, a chromatography column F, a flow-throughgamma detector 0, four two-way solenoid valves, and one threeway. Flask E is a 40-ml conical centrifuge tube. This tube wassealed with a silicone stopper through which has been inserted fourlengths ofTeflon tubing, one each for an inlet and an outlet (1.5mmo.d.), and a vent and an external access (both 3.0 mm o.d.) for theintroduction of a nitrogen gas sweep and the addition of solvent.Flask E was immersed in a heated oil bath. This heat, along withthegentlenitrogengassweep,aidsintherapidevaporationofthesolvents transferred from flask D. The top ofthe chromatographycolumn wassealedwitha siliconestopper through whichhad beeninserted two lengths of Teflon tubing, one for the reaction-mixtureinlet (I .5 mm o.d.) and one for an external access (3.0 mm o.d.)to apply vacuum and pressure, or to add solvent (petroleumether/methyl acetate, 7:3). This solution is transferred to columnF and the column washed with the same solvent (10 ml). Thecolumn effluent passes through a flow-through gamma detector(G) to a three-way solenoid valve that, until activated, directs thesolvent flow to a waste container. This detector was fabricated bywinding four or five turns ofTeflon tubing (1.5 mm o.d.) arounda Geiger-Mueller tube and placing the unit inside a suitable leadcontainer. Thus the only radiation reaching the Geiger-Muellertubecomesfromthecolumneffluentpassingthroughthetubing;this activity is registered on an external monitor (see above) andsignals the diversion of the desired product to the next unit.

The fourth unit, wherethe hydroxyl-protectingketalsof radiolabeled 4 are hydrolyzed, is composed of an open flask H, achromatography column J, a vial K, and two two-way solenoidvalves. Into flask H (a 25-ml three-neck round-bottom flask) wereinserted three lengthsof Teflon tubing, one each for the inlet (1.5mm o.d), the aqueous [I-' ‘C]2-DGsolution (after hydrolysis)outlet,and the externalaccess(both3.0mmo.d.) forthe introductionofa nitrogengas sweepand the additionofthe resinand/orsolvent. The outlet tube was extended to the bottom of the flaskand secured. The top of the chromatography column was sealedwith a silicone stopper through which had been inserted two lengthsofTeflontubing,oneforthefinalproductsolutioninletandoneforan external access (both 3.0 mm o.d.) for applying vacuum orpressure. When the hydrolysis of 4 is completed, the solution istransferred to the top of column J and the column effluent is collected in vial K, whichis preloadedwithcrystallinesodiumchloride(90 mg). An outlet tubing (1.5 mm o.d.) was extended to the bottomofthisvialandsecured.FlaskH,whichcontaineda magneticstir bar, was immersed in an oil bath and placed on a hot-platestirrer preset such that the oil bath was maintained at 80—85°C.

The fifth unit, for the sterilization of the isotonic aqueous [I-‘‘CJ2-DG solution, consists ofa 20-ml syringe barrel, one three

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PADGETr, BARRIO, MAcDONALD, AND PHELPS

carbon-Il-labeled glucononithle (3) by nucbeophilic displacementon the triflate (2), is facilitated by the useofpolar aprotic solvents(34). There are several suitable solvents—includingdimethylsulfoxide, N,N-dimethylacetamide, and N,N-dimethylformamide(DMF)—allofwhich share the propertiesofrelative nonvolatility(high boiling point) and high water solubility. Looking ahead tothefollowingstepinthissynthesis,thediisobutylaluminumhydridereduction of nitrile (3) to aldehyde (4) requires an inert solventand anhydrous conditions.To meet these requirements as wellasthose imposed by the system design policy (i.e., simplicity,semiautomation, lowcost, and safety), we havechosen to removethe displacement-reactionsolventby extractionrather than byevaporation. In practice this was achieved as follows: the triflate(2),whichisusuallyfreshlypreparedeachday,wasallowedtoreact with cyanide ion in DMF for I mm. (Longer reaction timesresult in the formationof a sideproductwithoutincreasingtheyield of nitrile 3.) Then, I5% aqueous sodium chloride and diethylether were added, followed by thorough mixing. The etherealphase,containingthe desiredglucononitrile(3) and traces of waterand DMF, was washed with saturated aqueous sodium chlorideand further dried by passage over mixed anhydrous magnesiumand calcium sulfates. The DMF/I5% aqueous sodium chloridesolution was reextracted with an additional portion of diethyl ether(3 ml). At this point the combined ethereal extracts contained nodetectable DMF as determined by gas chromatography (3%OV-17, 6 ft X ‘/@in column at 100 °C;flame ionization detection;flow rate 35 ml/min; retention time 2.2 mm). The complete reduction of nitrile 3 with an excessof diisobutylaluminum hydridein hexane-diethylether then proceededsmoothlyin goodyield.Theexcess hydride reagent was subsequently decomposed to isobutylene and aluminum salts by the mixture of alcohol and diluteaqueous sulfuric acid.

The next step, the isolation of the carbon-i i-labeled 2-DGprecursor, was conveniently accomplished by column chromatography of the organic phase from the reduction reaction. Afterevaporation of the hexane, diethyl ether, and most of the ethanolsolvent, the residue contains unreacted triflate (2) in addition toalcohol (1) and protected aldehyde (4), as shown by thin-layerchromatography on silica gel using petroleum ether/methyl acetate (7:3) (Rr s: I 0.30; 4 0.50; 2 = 0.65). This was taken upin 1—2ml of petroleum ether/methyl acetate (7:3) and applied tothe top of the silicagel column. Developmentof the column usingthe same solvent mixture effectivelyseparates the three components: the triflate (2) elutes in the first 6 ml, and the desired radiolabeled aldehyde (4) elutes in the next 4 ml. Alcohol I remainson the column under these conditions. Thus all of the startingprotected D-arabitol,presentas the alcoholand unreacted triflate,is separated from the protected El-' ‘C]2-DGat this step. Elâ€â€˜C]2-DG preparations, free of D-arabitol for administration to

humans or animals, are nowconvenientlyobtained and are probably necessary for cerebral glucose metabolic-rate determinationsowing to the competitive nature of glucose-facilitated transportinto the brain (35).

The nextstep,the removalof the hydroxyl-protectingisopropylidine groups of aldehyde 4, presented potential problems frombothmechanicalandchemicalstandpoints.Useofstandardketalhydrolysis conditions, namely, aqueous acid at elevated temperature, could result in the acid-catalyzed decomposition of thedeprotected product, 2-deoxy-D-glucose, which is known to decomposethrough dehydrationin aqueousacid (36). Mechanically,this process could result in unnecessary manipulation and the inconvenientvolumesassociatedwithneutralizationand purification.These problems were eliminated by the use of a strongly acidicpolystyrene cation-exchange resin (37). Such resins are used forgeneral acid-catalyzed reactions, and are easily removed after useby filtration. Thus the protected aldehyde (4) is heated to 70-75°Cwith an aqueous suspensionof cation-exchange resin (hydrogen

way solenoid valve, a 0.22-aim membrane filter L, and a I0-mlsterile serum vial M. The openend of the syringebarrel wassealedwith a silicone stopper through which was inserted a length ofTeflon tubing (3.0 mm o.d.) for an external access. The Luer tipendofthesyringebarrelwasconnectedtothecommonportofthethree-way solenoid valve, and the membrane filter and serum vial,with vent needle, were connected to the normally closed port of thevalve. Sterilization of the product is then accomplished by creatingvacuum in the syringe barrel, which pulls the solution through thevalve and into the barrel. The solenoid valve is then activated andmild pressure is applied through the external access tubing to pushthe finalsolutionoutofthe syringebarrel,throughthemembranefilter, and into the sterile vial.

Before each run, all tubing, solenoid valves, and flasks areflushed with sterile water, acetone, and dry nitrogen gas. All disposable components (e.g., membrane filter, vials) are replaced,and all chromatography columns are repacked. These simpleprecautions ensure sterile, pyrogen-free preparations. Multipleruns per day, for example for serial studies, would require twoidentical processing systems to minimize exposure to the radiochemist. This requirement is easily met, since these systems aresimple, small, and inexpensive.We have found, however, that asingle system would meet the requirements of serial runs, if theywere separated by 2.5 to 3 hr, without unnecessarily increasingexposure to the chemist.

RESULTS AND DISCUSSION

The remote, semiautomated system used in this synthesis of[1-' ‘C]2-DGis an extension of the designs used in this laboratoryto construct similar systemsfor the routine preparation of 2-FDG(F-I8), [l-―CJPA, and carbon-Il and nitrogen-13 L-amino acids(6,13—18).Ourapproachto thedesignof thesesystemswasdictated by two requirements. First, the system must be composedof simple, interchangeable units, one for each type of operationbeing carried out (e.g., aqueous-organic extraction, column purification,sterilization,etc.) Second,theseunitsare to becomposedof common laboratory items whenever possible, connected byTeflon tubing through Teflon-body solenoid valves. Thus, anysystem initially constructed for the preparation of a specificcompoundmay easily be disassembledinto its components,whichcan then be recombined in a different way to obtain another synthesis system. In addition, this approach permits easy adaptationto microprocessor-controlledoperation to achieve authentic automated synthesis. In line with these objectives, each unit (or step)of the synthetic procedure has been simplified as much as possible.

The synthesisof [1-' ‘C]2-deoxy-D-glucosereportedin this paperis an adaptation of the procedure reported by Bayly and Turner(28) forthepreparationof [I-'4C]2-D-ribose,andconstitutesamodification of the approaches used in other reported synthesesof [l-―C]2-DG(24,29-32).

The requisite hydrogencyanide (C-l 1)was prepared, followingthe method of Banfi et al. (21,22), via the ‘4N(p,a)'‘Creactionby the bombardment of nitrogen gas with 22-MeV protons. Theinitially produced labeled carbon dioxide was reduced to methane(C-I 1)by passageovernickelcatalyst at 370 °Cin the presenceof hydrogen, and was then mixed with anhydrous gaseous ammonia and passed over platinum metal at 950 °Cto yield the desired labeled hydrogen cyanide. Using a literature procedure (24),the carbohydrate starting material, l-O-trifluoromethanesulfonyl-2,3:4,5-di-O-isopropylidine-D-arabitol (2, Scheme 1), wasgenerated from the corresponding alcohol (I) (33), and trifluoromethanesulfonic anhydride. This triflate, in pentane, may bestored at 0 °Cfor several hours, or overnight if necessary.

Thefirststepinthesynthesisof El-'‘C]2-DG,formationofthe

742 THE JOURNAL OF NUCLEAR MEDICINE

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NumberTypicalactivity (mCi)

FinalCompoundof runsStartproducts Reference

TECHNICAL NOTES

form) to give the desired El-' ‘C]2-DG(5) without any evidenceof concomitant acid-catalyzed decomposition. At lower temperatures the hydrolysis reaction rates are slow and consequentlyimpractical; at temperatures higher than 80 °C,decomposition(dehydration) products are observed.

Before the use of carbon-I I , all reactions and processing manipulations were fully optimized using both cold chemistry withstandard analytical techniques and carbon-i4-labeled intermediates, and all losses of activity were consistent with those expectedfor a multistep synthetic process.

El-' ‘C]2-Deoxy-D-glucoseis now available for serial studies inthe typical nuclear medicine clinical setting. Utilization of thedescribed remote semiautomated production system results in thepreparation of El-' ‘C]2-DGon a routine basis in 25—30%radiochemical yield, (based on the amount of hydrogencyanide(C-i 1)initially trapped in the sodium cyanide/DMF solution), after asynthesis time of “-‘50mm, and with a radiochemical purity of>98% as determined by thin-layer radiochromatography. Thus,starting with 600-720 mCi of H' ‘CN,25-30 mCi of El-' ‘C12-DGare obtained.

The specific activity of carbon-i 1 activity delivered and trappedin the reaction vessel,while being variable depending on the duration of bombardment, is “@.‘I.2Ci/mmol at 60 mm after EOB.This was determined by conversion ofthe radiolabeled cyanide intoDL-El‘‘C]leucine(18,22) and verification of its specificactivityby using the o-phthaldialdehyde (OPT) precolumn fluorescencederivatization procedure as previously described for L-amino acidslabeled with carbon-Il and nitrogen-i3 (16,17). From these datawethereforeobtain the specificactivityof El-'‘C]2-DGat the timeof injection, 60 mm after EOB (10 mm to allow for the conversionof ‘‘CO2to H' ‘CNand 50 mm for synthesis time).

CONCLUSION

In this work we have demonstrated the successfulapplicationof the unit-operations approach to the designand construction ofthe remote semiautomated synthesis system for the routine preparation of [I-' ‘C]2-DG.The flexibility and reliability ofthe designapproach described here is attested by the record of over 750production runs for the preparation of 2-FDG(F-18), [1-' ‘C]2-DG, El-' ‘C]palmiticacid, L-amino acids labeled with carbon-i 1and nitrogen-l3, produced enzymatically from carbon dioxide(C-I I) and ammonia(N-l3), and L-[l-' ‘C]leucinepreparedusing the Bucherer-Strecker reaction (Table 1).

The ultimate goal of the work is to refine this approach toachieve complete automation of the synthetic process through totalmicroprocessorcontrol. These systems will then take the form ofan enclosedcompactboxnot too different from other sophisticatedbut functional laboratory and clinical instruments.

a Altex Scientific, Inc.

t Pharmaseal, Inc.

FOOTNOTES

ACKNOWLEDGMENTS

WethankJ. S. Cook,R. L. Birdsall,and L. J. McConnell,ofthe UCLA Medical Cyclotron staff, for the preparation of hydrogen cyanide(C-l 1), and Dr. A. Najafi for his initial work onthe synthesis of [1-' ‘C]2-DG.This work was supported in part byDepartment of Energy Contract Number DE-AMO3-76-SF00012,and National Institute of Health Grant Number 7R01-GM24839.

APPENDIX

All Teflon tubing and fittings were purchased from Altex Scientific,Inc.and the two-and three-wayTeflon-bodysolenoidvalveswereobtainedfrom FluorocarbonCo. The ion-exchangeresinsandchromatography columns were from Bio-Rad Laboratories. Thevortex mixers (Lab-Line No. 1290), magnetic stirrer (Lab-LineNo. 1250), and hot-plate stirrer (Thermoline SP-IOIOSB) werefrom VWR Scientific, Inc. The commercial flow-through radioactivity detector was Ortec Model 406-A. The special flow-throughgamma detector was constructed using a radiation monitor (Dosimeter Corp., MiniRAD Model 3006) modified to function withan external Geiger-Muellertube. Thin-layer radiochromatogramsweregenerated from developedTLC plates by a Technical Associates Model HY-3R2 radiochromatogram scanner. Nitrogen andhelium (99.999%, less than 3 ppm H2O, 2 ppm 02) were from theLiquid Carbonic Corp. Dimethyldichlorosilane (DMCS) andtrifluoromethanesulfonic anhydride were from Tridom ChemicalCo.Theanhydridewaspurifiedbeforeuse(19). Di-isobutylaluminum hydride was from Alfa Products. N,N-dimethylformamide(DMF) was purified by the literature procedure (20). [i-'4C]2-deoxy-D-glucose was purchased from New England Nuclear Corp.

E1-11ClPalmiticacid2-@1@F]FIuoro-2-deoxy-o-glucoseL-E13N]Ammnoacids (from 13NH3)L-E11ClAmmnoacidsandTCAdcycle

intermediates(from 11C02)L-E11ClAminoacids(fromH‘1CN)E1-11C]2-Deox@'-c-@Iucose

137 200—250l50@l80'@175—200200—250

50—7518—253_60c

25—30

15

6

1617

41610860

2817

700—800600—720

20—2525—30

1813, 14

a Ready for injection.

b Products of fluorination of 3,4,6-tri-O.acetyl-o-glucal.

C Variation reflects data from nine different amino acids.

d Tricarboxylic acid.

Volume 23, Number 8 743

TABLE1. SUMMARYOF REMOTESEMIAUTOMATEDRADIOPHARMACEUT1CALPRODUCTiON

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PADGETT, BARRIO, MACDONALD,AND PHELPS

Yeast hexokinase (E.C.2.7.l.l.) was obtained from SigmaChemical Co.

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744 THE JOURNAL OF NUCLEAR MEDICINE

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1982;23:739-744.J Nucl Med.   H. C. Padgett, J. R. Barrio, N. S. MacDonald and M. E. Phelps  C]2-Deoxy-d-Glucose for Routine Clinical Applications

11The Unit Operations Approach to the Applied to the Synthesis of [1-

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