An algorithm for the deconvolution of mass spectrosopic patterns in isotope labeling studies. Evaluation for the hydrogen-deuterium exchange reaction in ketones

Article abstract of DOI:10.1021/jo070831o

(Graph Presented) An easy to use computerized algorithm for the determination of the amount of each labeled species differing in the number of incorporated isotope labels based on mass spectroscopic data is described and evaluated. Employing this algorithm, the microwave-assisted synthesis of various α-labeled deuterium ketones via hydrogen-deuterium exchange with deuterium oxide was optimized with respect to time, temperature, and degree of labeling. For thermally stable ketones the exchange of α-protons was achieved at 180°C within 40-200 min. Compared to reflux conditions, the microwave-assisted protocol led to a reduction of the required reaction time from 75-94 h to 40-200 min. The α-labeled deuterium ketones were reduced by biocatalytic hydrogen transfer to the corresponding enantiopure chiral alcohols and the deconvolution algorithm validated by regression analysis of a mixture of labeled and unlabeled ketones/alcohols.

Full text of DOI:10.1021/jo070831o

An Algorithm for the Deconvolution of Mass Spectrosopic Patterns
in Isotope Labeling Studies. Evaluation for the
Hydrogen-Deuterium Exchange Reaction in Ketones
Christian C. Gruber,† Gustav Oberdorfer,† Constance V. Voss,† Jennifer M. Kremsner,‡
C. Oliver Kappe,‡ and Wolfgang Kroutil*,†
Institute of Chemistry and Christian Doppler Laboratory for MicrowaVe Chemistry (CDLMC),
Karl-Franzens-UniVersity Graz, Heinrichstrasse 28, A-8010 Graz, Austria
wolfgang.kroutil@uni-graz.at
ReceiVed April 20, 2007
An easy to use computerized algorithm for the determination of the amount of each labeled species
differing in the number of incorporated isotope labels based on mass spectroscopic data is described and
evaluated. Employing this algorithm, the microwave-assisted synthesis of various R-labeled deuterium
ketones via hydrogen-deuterium exchange with deuterium oxide was optimized with respect to time,
temperature, and degree of labeling. For thermally stable ketones the exchange of R-protons was achieved
at 180 °C within 40-200 min. Compared to reflux conditions, the microwave-assisted protocol led to a
reduction of the required reaction time from 75-94 h to 40-200 min. The R-labeled deuterium ketones
were reduced by biocatalytic hydrogen transfer to the corresponding enantiopure chiral alcohols and the
deconvolution algorithm validated by regression analysis of a mixture of labeled and unlabeled ketones/
alcohols.
Introduction
mination of enantiomeric and diastereomeric excess by mass
spectrometry.3 Not surprisingly, there is therefore an ever
increasing demand for the synthesis of commercially unavail-
able, labeled starting materials and their accurate characteriza-
tion.
Because isotope labeling generally relies on the substitution
of atoms with the same chemical properties but different atomic
mass, mass spectrometry is often the analytical tool of choice.4
Thus, a significant amount of work has been carried out to date
on the detection of deuterium-labeled molecules with use of
mass spectrometry.5 For our research on alcohol dehydrogena-
ses, we required R-labeled ketones as well as â-labeled
secondary alcohols as starting materials and envisaged applying
a rapid microwave-assisted hydrogen/deuterium exchange pro-
cess for their efficient synthesis.6,7 In this context, however, we
noticed that for reaction optimization a reliable characterization
Isotope labeling, in particular 13C and deuterium marking, is
one of the most widespread methods for investigating molecular
structures and reaction mechanisms1 and additionally plays an
important role in deducing the metabolic pathways of drugs and
biologically active molecules.2 Deuterium-labeled compounds
recently also attracted attention in the high-throughput deter-
* To whom the correspondence should be addressed. Phone: +43-316-380-
5350. Fax: +43-316-380-9840.
† Institute of Chemistry.
‡ Christian Doppler Laboratory for Microwave Chemistry.
(1) (a) Simon, H.; Palm, D. Angew. Chem., Int. Ed. 1966, 5, 920. (b)
Ozaki, A. Isotopic Studies of Heterogeneous Catalysis; Kodansha/Academic
Press: New York, 1977. (c) De Meer, K. M.; Roef, M. J. M.; Kulik, W.;
Jakobs, C. Isot. EnViron. Health Stud. 1999, 35, 19.
(2) (a) Guaragna, A.; De Nisco, M.; Pedatella, S.; Pinto, V.; Palumbo,
G. J. Labelled Compd. Radiopharm. 2006, 49, 675. (b) Miranker, A.;
Robinson, C. V.; Radford, S. E.; Alpin, R. T.; Dobson, C. M. Science 1993,
262, 896. (c) Synthesis and Applications of Isotopically Labelled Com-
pounds; Pleiss, U., Voges, R., Eds.; John Wiley & Sons: Chichester, UK,
2001; Vol. 7. (d) Buncel, E. Synthesis and Applications of Isotopically
Labelled Compounds; Kabalka, W., Ed.; Elsevier Science Ltd.: Amsterdam,
The Netherlands, 1992. (e) Vederas, J. C. Nat. Prod. Rep. 1987, 4, 277. (f)
Thomas, A. F. Deuterium Labeling in Organic Chemistry; Appleton-
Century-Crofts: New York, 1971.
(3) (a) Tsukamoto, M.; Kagan, H. B. AdV. Synth. Catal. 2002, 344, 453.
(b) Reetz, M. T. Angew. Chem., Int. Ed. 2001, 40, 284.
(4) Heumann, K. G. Int. J. Mass Spectrom. 1982, 45, 87.
(5) (a) Dienes, T.; Pastor, S. J.; Schurch, S.; Scott, J. R.; Yao, J.; Cui, S.
L.; Wilkins, C. L. Mass Spectrom. ReV. 1996, 15, 163-211. (b) Loaiza,
A.; Zaera, F. J. Am. Soc. Mass Spectrom. 2004, 15, 1366.
10.1021/jo070831o CCC: $37.00 © 2007 American Chemical Society
Published on Web 06/20/2007
5778
J. Org. Chem. 2007, 72, 5778-5783

DeconVolution of Mass Spectrosopic Patterns
SCHEME 1. Hydrogen-Deuterium Exchange in the
r-Positions of 3-Octanone
columns). The pattern results on the one hand from the natural
distribution of 13C and other atoms showing significant isotope
distributions (e.g., Cl) as well as from fragmentation. For these
theoretical considerations we have chosen a starting material
that can be labeled only in two positions. Since we deal with
low-resolution MS, the abundance pattern of the single deute-
rium labeled derivative d1 has to be the same as the unlabeled
one but shifted one mass unit upward. The d2-derivative pattern
is shifted two mass units to higher mass and so on. The pattern
of a mixture (the analyte) results from the sum of the fraction
of each derivative multiplied by the abundance at the corre-
sponding mass. For instance, in Table 1 at m/z of M + 2 the
abundance of the analyte (here z2) is proportional to c‚x(d0) +
b‚x(d1) + a‚x(d2), where x(di) is the fraction of the corresponding
derivative. Therefore, for the simple example in Table 1 five
equations (for each m/z M to M + 4 one equation) are obtained.
In general, considering the number of equations that are
obtained and the number of derivatives, the number of equations
is higher than the number of derivatives. Taking the example
of Table 1, we have three derivatives (d0-d2) but five equations
(M until M + 4, or z0 to z4). Therefore we deal with an
overdetermined linear equation system, which cannot be exactly
solved (for real data). The equations can be written in a
mathematical way, by using matrixes (eq 1), whereby A is the
matrix (for the example in Table 1, it is the one in the box, the
places without a value are set to 0), x is a vector containing the
fraction of each derivative {for the example [x(d0), x(d1), x(d2)]T,
T means transposed}, and b are the abundances of the analyte,
e.g., (z0, z1, z2, z3, z4)T.
of the labeling degree was required to be able to properly
optimize the synthesis of the desired labeled ketones. We
herewith present a novel algorithm for the exact determination
of the degree of labeling that is based on the evaluation of mass
spectroscopic data by computerized methods.
Results and Discussion
Generation of Deuterium-Labeled Ketones. Our studies
commenced with the investigation of the hydrogen-deuterium
exchange reaction in simple ketones using deuterium oxide
(D2O) as the deuterium source and CD3COOD as a catalyst.
Deuterium exchange in D2O/CD3COOD6 of a rather simple
ketone like 3-octanone 1a employing controlled microwave
heating in sealed vessels or conventional reflux conditions
proceeded via various deuterated intermediates. Considering the
four acidic R-protons of 1a, five derivatives are possible, namely
d1, d2, d3, d4, and the unlabeled starting material d0.
In order to follow the degree of deuterium labeling in
3-octanone 1a during the reaction and to characterize the
obtained products, low-resolution MS analysis may be consid-
ered as an appropriate method. However, the mixture of
compounds labeled at the two different R-positions to varying
degrees led to a rather complex mass spectrum (Figure 1b).
We expected that the deconvolution of such a pattern is well
described in the literature. To our astonishment we found a rather
unsatisfying procedure in two standard reference books.8 Es-
sentially, this procedure proposes to take the abundance of the
lowest mass peak in the area of interest that can only be
attributed to one derivative and subtract the corresponding
abundance of this derivative from the abundance at higher mass
and then continue with the next higher substituted derivative,
and by using this iterative procedure the fraction of each
derivative is obtained. It is evident that this method is rather
inaccurate, since it relies completely on the accuracy of a single
abundance at a single mass (the first one), which leads to a
high inaccuracy. An additionally performed thorough search in
the primary literature did not lead to any more sophisitcated
algorithms for this specific deconvolution problem.
A‚x ) b
(1)
where A is the m × n matrix, m * n, m is the number of
columns, and n is the number of rows; x is a vector (1 × m),
containing the fraction of each derivative, i.e., [x(d0), x(d1), ...,
x(dm)]T; and b is a vector (1 × n), representing the abundances
of the analyte, i.e., (z0, z1, ..., zn)T.
Since we have an overdetermined system and we deal with
data of measurements which have a certain error, an algorithm
to minimize the error will definitely be superior to the rather
cumbersome literature procedure.8 Applying the least-squares
method to the overdetermined linear equation system, the
solution is found by applying the pseudoinverse matrix of A
which is (AT‚A)-1‚A.9 From a mathematical point of view, it
is required that the column vectors of A have to be linear
independent, which is fulfilled for our problem.
Therefore x of eq 1 can easily be obtained by the expression
given in eq 2.
Analyzing the problem in more detail, we listed the abundance
pattern of a hypothetical organic (carbon atom-based) starting
material at the different m/z ratios of interest (Table 1, two left
x ) (AT‚A)-1‚A‚b
(2)
Although this expression looks rather short, the determination
of x requires a number of mathematical operations. Standard
programs like Microsoft Excel can deal with these expressions.
However, to circumvent problems associated with Excel soft-
ware we prepared an Excel sheet for both PC and Macintosh
computer platforms.10 Since the same question arises when
dealing with 13C or 17O labeling, the identical sheet is also
applicable for these derivatives. In the case of tritium, 18O, or
(6) (a) Fodor-Csorba, K.; Galli, G.; Holly, S.; Gacs-Baitz, E. Tetrahedron
Lett. 2002, 43, 3789. (b) Jones, J. R.; Lockley, W. J. S.; Lu, S. Y.;
Thompson, S. P. Tetrahedron Lett. 2001, 42, 331. (c) Loupy, A.; Petit, H.;
Hamelin, J.; Texier Boullet, F.; Jacquault, P.; Mathe, D. Synthesis 1998,
1213. (d) Anto, S.; Getvoldsen, G. S.; Harding, J. R.; Jones, J. R.; Lu, S.
Y.; Russel, J. C. J. Chem. Soc., Perkin Trans. 2 2000, 2208.
(7) For a general review on controlled microwave synthesis, see: (a)
Kappe, C. O. Angew. Chem., Int. Ed. 2004, 43, 6250. (b) For a review on
microwave-assisted radiochemistry, see: Jones, J. R.; Lu, S.-Y. In
MicrowaVes in Organic Synthesis, 2nd ed.; Loupy, A., Ed.; Wiley-VCH:
Weinheim, Germany, 2006; Chapter 13, pp 435-462.
(8) (a) Hesse, M.; Meier, H.; Zeeh, B. Spectroscopic Methods in Organic
Chemistry; Thieme: New York, 1997; pp 256-267. (b) Budzikiewicz, H.
Massenspektrometrie; Wiley-VCH: Weinheim, Germany, 1998; pp 70-
71.
(9) Poole, D. Linear Algebra; Thomson Brooks/Cole: New York, 2006.
(10) The Excel sheets can be downloaded free of charge from: ftp://
biocatalysis.uni-graz/pub/IsoPat2/. When opening, the Excel-file macros
must be activated.
J. Org. Chem, Vol. 72, No. 15, 2007 5779

Gruber et al.
FIGURE 1. Mass spectrum of the molecule ion area of 3-octanone: (a) unlabeled 3-octanone 1a and (b) mixtures of d0-, d1-, ..., d4-3-octanone.
TABLE 1. Deconvolution Algorithm for a Compound To Be
Labeled in Two Positionsa
Apart from 3-octanone 1a, a variety of other ketone substrates
(Figure 2) were exposed to the deuteration conditions. For
example, hydrogen-deuterium exchange in ketone 2a led to a
complex mixture of all possible compounds d0-d5 after 40 min
at 180 °C by using microwave irradiation (Figure 4, 2a). By
increasing the reaction time the amount of the fully labeled
product d5-2a could be increased to 80%. Compounds d0-, d1-,
and d2-2a were only visible at 40 min, afterward the amount of
d4-2a slowly decreased, while that of d5-2a increased. The other
aliphatic ketones 1a and 3a showed a quite similar behavior:
after 200 min 85% and 82% respectively of the highest labeled
derivative d4 could be reached (Figure 4). For ketone substrates
1a-3a only protons in the R-position of the carbonyl moiety
were exchanged. For substrate 4a we observed in addition to
the exchange of the five R-protons an exchange of the two
benzylic protons leading to a sevenfold labeled derivative d7-
4a. Comparing the deuterium exchange for 4a under reflux
conditions (∼100 °C) to the microwave-assisted exchange
reaction at 180 °C, similarly deuterated product mixtures were
obtained under reflux after 75 h while under microwave
conditions only 1 h was required (Figure 4).
unlabeled d0
d1-derivative
d2-derivative
analyte
m/z
(abundance)
(abundance)
(abundance)
(abundance)
M
a
b
c
z0
z1
z2
z3
z4
M + 1
M + 2
M + 3
M + 4
a
b
c
a
b
c
a The unlabeled compound shows abundances (a, b, c) at m/z of M, M
+ 1, and M + 2. The mixture of the di-derivatives shows abundances (z0-
z4) at m/z from M to M + 4.
FIGURE 2. Substrates tested in hydrogen-deuterium exchange
reactions.
For more complex substrates like R-chloroketones 5a and 6a,
optimization of the reaction parameters demonstrated that a
temperature <180 °C was required to avoid the formation of
side products due to the thermal instability of the educts. For
substrate 5a the main product after 180 min at 100 °C under
microwave conditions did not show the maximum degree of
labeling d4. The main product was the 3-fold labeled d3-5a,
probably due to the lower temperature chosen as a compromise
between the degree of labeling and the undesired thermal
degradation of 5a. Nevertheless, the unlabeled substrate d0-5a
was found in a concentration of <1%. In case of ω-chloro
acetophenone (6a), the amount of byproducts was approximately
20% when lowering the temperature to 150 °C (40 min). Under
these conditions the highest possible degree of labeling d2 was
reached with a purity of 98% (Table 2) and the corresponding
product could be isolated with 73% yield. Especially for
substrates with atoms possessing a significant natural isotope
distribution (for example, chlorine atoms), the resulting mass
patterns become rather complicated and would be tedious to
solve without the program (Figure 5b). The results of the
optimization of the reaction parameters are summarized in Table
2.
14C labeling, the matrix differs from the problem shown in Table
1 only as far as the mass of each derivative is shifted two mass
units upward. The setup of such a matrix is again rather easy
and was realized in an additional worksheet in the above-
mentioned Excel file. For more complex analysis a Mathematica
routine was developed. Further details on the Excel and
Mathematica programming are provided in the Supporting
Information.
Being now able to quickly analyze a mixture of derivatives
varying only in their degree of isotope labeling, the transforma-
tion of ketones 1a-6a (Figure 2) to their corresponding labeled
derivatives was optimized by following the degree of labeling
in D2O/CD3COOD at different reaction temperatures in a sealed
vessel microwave reactor (100-180 °C) or under reflux
employing traditional heating (∼100 °C).
During the deuteration reaction, samples were drawn and the
complex mass patterns of the resulting mixtures were measured
by MS (Figure 3a). By applying the above-described algorithm
for deconvolution the time course of each derivative could be
followed (Figure 3b).
5780 J. Org. Chem., Vol. 72, No. 15, 2007

DeconVolution of Mass Spectrosopic Patterns
FIGURE 3. Deconvolution of the mass spectroscopic raw data (a) leading to the time course (b) of each d0-d4-derivative (3-octanone 1a). Reaction
conditions: 0.8 mol/L substrate, CD3COOD, D2O, 200 min, 180 °C, single mode microwave irradiation.
FIGURE 4. Time course of di-derivatives during the deuterium exchange for ketones 2a, 3a, and 4a (Figure 2). Reaction conditions: 0.7-0.8
mol/L substrate, CD3COOD, D2O, 60 or 200 min, 180 °C, single mode microwave irradiation (for 4a additional reflux conditions, 100 °C, 96 h).
TABLE 2. Optimized Reaction Conditions for Microwave-Assisted
unlabeled “impurity” and thus is a good parameter to character-
ize a product mixture. Due to the additional properties we named
that parameter l.r. for labeled compound ratio (eq 3):
Hydrogen-Deuterium Exchange
optimized reaction
conditions
isolated yield
n
sub-
strate product
main
T
time
(min)
l.r.
(%)
purityb
(%)
(°C)a
mg
%
d
∑
i
l.r. ) i)1 100[% ]
(3)
1a
2a
3a
4a
4a
5a
6a
d4-1a
d5-2a
d4-3a
d5-4a
d5-4a
d3-5a
d2-6a
180, MW
180, MW
180, MW
180, MW
100, reflux 94 h
100, MW
150, MW
200 >99.9
200 >99.9
200 >99.9
40 >99.9
292
335
274
456
377
230
369
58
67
55
91
76
46
73
84
80
82
57
75
38c
98
n
d
∑
i
i)0
99.8
99.7
180
40 >99.9
where l.r. is the labeled compound ratio, di is the concentration/
amount of i-times labeled derivative, and n is the highest
possible degree of labeling. For example, a l.r. of 100%
corresponds to a mixture of labeled derivatives with no unlabeled
substrate, while an l.r. of 0% stands for the unlabeled starting
material.
Synthesis of Deuterium-Labeled Chiral Secondary Alco-
hols. We recently applied an alcohol dehydrogenase for
deuterium transfer to obtain chiral deuterium labeled secondary
a MW ) microwave. b Percentage of labeled main product to all other
labeled derivatives. c 15% d4-6a
In case of no side reactions, the conversion (the amount of
converted substrate over the total starting material) is equal the
sum of all labeled derivatives over the sum of labeled and
unlabeled derivatives. Additionally, this parameter also gives a
quantitative yield of labeled compounds or of the amount of
J. Org. Chem, Vol. 72, No. 15, 2007 5781

Gruber et al.
FIGURE 5. Mass pattern of (a) unlabeled compound 5a and (b) after deuterium exchange (D2O/CD3COOD) after 80 min at 100 °C under microwave
irradiation conditions.
FIGURE 6. Synthesis of deuterium labeled chiral secondary alcohols in a coupled substrate approach: (a) R-Labeled starting material leading to
enantiopure alcohols with a stable label and (b) unlabeled starting material, labeled hydride source for asymmetric reduction.
TABLE 3. Biocatalytic Reduction of r-Labeled Ketones to Chiral
alcohols, for which the deuterium was located at the stereocenter.
Secondary Alcohols
As a source of deuterium, d8-2-propanol was applied in a
yield
coupled substrate approach (Figure 6b).11 These labeled alcohols
catalyst
substratea
product
mg
%
ee (%) l.r. (%)
have the disadvantage that the label is removed during a
metabolic oxidation of the alcohol. In contrast, when preparing
labeled chiral alcohols from R-labeled ketones the label on the
carbon adjacent to the chiral center is stable even during
oxidation steps. Additionally, for monitoring these labeled
compounds for kinetic and mechanistic studies during metabo-
lism, no primary isotope effect due to cleavage of a C-D bond
can cause an unwanted distortion of the observed data for
oxidations. For the synthesis of such deuterium-labeled enan-
tiopure chiral alcohols the labeled ketones dn-1a-6a were
reduced to the corresponding enantiopure labeled alcohol by a
biocatalytic hydrogen transfer method (Figure 6a). By employing
the alcohol dehydrogenase from Rhodococcus ruber ADH-
“A”10,12 and the stereocomplementary alcohol dehydrogenase
from Lactobacillus breVis LB-ADH13 both enantiomers were
accessible in good yields and excellent enantiomeric excess
(Table 3).
ADH-“A”
d4-1a
d5-2a
d4-3a
d5-4a
d3-5a
d2-6a
d4-1a
d5-2a
d4-3a
d5-4a
d3-5a
d2-6a
d4-(S)-1b
d5-(S)-2b
d4-(S)-3b
d5-(S)-4b
d3-(R)-5bb
d2-(R)-6bb
d4-(R)-1b
d5-(R)-2b
d4-(R)-3b
d5-(R)-4b
d3-(S)-5bb
d2-(S)-6bb
73
79
61
61
92
74
56
74
27
98
84
87
71.6
77.5
59.8
59.8
90.2
72.5
54.9
72.5
26.5
96.1
82.4
85.3
>99
>99
>99
>99
>99
>99
>99
>99
>99
>99
>99
>99
>99.9
>99.9
>99.9
>99.9
99.7
>99.9
>99.9
>99.9
>99.9
>99.9
99.7
LB-ADH
>99.9
a 100 mg of 1a-6a. b Switch of CIP priority.
reactions except that the mass pattern was shifted one (2-
propanol) or two mass units (d8-2-propanol) to higher mass.
For validation of the MS-pattern deconvolution algorithm the
obtained labeled chiral alcohols as well as the labeled ketones
were mixed with unlabeled compounds. The mixtures were then
analyzed by MS. After deconvolution of the data the results
were plotted in a diagram with the increasing mixed amount of
labeled compound asthe x-axis and on the y-axis the labeled
ratio. In case the algorithm performs as outlined above, a linear
trend graph should result with a regression correlation coefficient
(R2) close to “1”. As an example, unlabeled 1a was admixed
with the product of the microwave-assisted deuterium exchange
reaction containing the d1-d4-derivatives of ketone 1a (Figure
The obtained alcohols were analyzed by GC-MS as well as
GC-FID on a chiral stationary phase. Its is worth mentioning
that the labeling pattern did not change during the reduction
(11) Edegger, K.; Gruber, C.C.; Poessl, T. M.; Wallner, S. R.; Lavandera,
I.; Faber, K.; Niehaus, F.; Eck, J.; Oehrlein, R.; Hafner, A.; Kroutil, W.
Chem. Commun. 2006, 2402.
(12) Stampfer, W.; Kosjek, B.; Moitzi, C.; Kroutil, W.; Faber, K. Angew.
Chem., Int. Ed. 2002, 41, 1014.
(13) Wolberg, M.; Hummel, W.; Wandrey, C.; Mu¨ller, M. Angew. Chem.,
Int. Ed. 2000, 39, 4306.
5782 J. Org. Chem., Vol. 72, No. 15, 2007

DeconVolution of Mass Spectrosopic Patterns
FIGURE 7. Validation of the deconvolution algorithm: (a) raw MS data morphing from unlabeled to labeled patternand (b) calibration data with
linear regression.
7). Gratifyingly, after analysis as described above the R2
reactor was cooled to room temperature the product was extracted
with CCl4 (4 times), washed with a saturated NaDCO3 solution (4
mL), and dried (Na2SO4) and the organic solvent was removed by
evaporation. When required the product was purified by silica gel
column chromatography. The product was analyzed by GC-MS (see
the Supporting Information for more details). From the obtained
MS pattern the percentage of the various labeled derivatives was
coefficient was 0.99. For the other series the R2 values were
between 0.98 and 0.99. The R2 values close to “1” therefore
validate the deconvolution algorithm presented above.
Conclusion
analyzed with use of the above-described Excel sheet.
Typical Procedure for the Synthesis of Deuterium-Labeled
sec-Alcohols. Lyophilized cells of E. coli Tuner pet22b-ADH-“A”
(310 mg) were rehydrated in TRIS-HCl buffer (30 °C, 60 min, 120
rpm) in a 50 mL vessel. The reaction was started by addition of
labeled 3-octanone (100 mg, 0.78 mmol) and 2-propanol (1515 µL,
15 v/v %). After 24 h at 30 °C, 120 rpm ,the reaction was stopped
In the context of performing isotope labeling studies by
isotope exchange reactions, an error-minimizing algorithm based
on the computerized analysis of readily available low-resolution
mass spectroscopic patterns was developed. The algorithm
allows the exact determination of the relative amounts of each
labeled species differentiated in the number of incorporated
deuterium, oxygen, carbon, or other isotopes. By using this
methodology the preparation of R-labeled ketones employing
microwave technology was optimized. The latter were then
reduced in an asymmetric fashion to yield enantiopure alcohols
possessing a stable label. Confirmation of the algorithm was
achieved by analysis of the linear regression correlation coef-
ficient.
by extraction with ethyl acetate (3×) before the solvent was
evaporated. 1H NMR and MS data are provided in the Supporting
Information.
Acknowledgment. This study was financed by the Austrian
Science Fund (FWF Project P18537-B03). We also thank
Biotage AB (Uppsala, Sweden) for use of the Emrys Synthesizer
and Initiator EXP Eight microwave reactors.
Experimental Section
Supporting Information Available: General experimental
procedures, 1H NMR and MS spectroscopic data, program descrip-
tions, and source code. This material is available free of charge
via the Internet at http://pubs.acs.org.
Typical Procedure for the Synthesis of Labeled Ketones.
3-Octanone (500 mg, 3.9 mmol) was added to D2O (3.5 mL, 175
mmol, 99.9%) and CD3COOD (1 mL, 16 mmol) in a 10 mL sealed
microwave process vial. The exchange reaction was performed at
180 °C for 200 min in a single-mode microwave reactor. After the
JO070831O
J. Org. Chem, Vol. 72, No. 15, 2007 5783