Formal synthesis of P-chiral [16O,17O,18O]phosphoenol pyruvates by means of the α-hydroxyphosphonate-phosphate rearrangement

Article abstract of DOI:10.1080/10426507.2018.1452235

Transesterification of tris(hexafluoroisopropyl) phosphite with racemic 3-methyl-1-phenyl-butane-1,3-diol gave two isomeric hexafluoroisopropyl-substituted 1,2,3-dioxaphosphinanes. These cyclic phosphites were hydrolyzed rapidly and enantioselectively by water catalyzed by HCl. The respective metalated H-phosphonates were added to ethyl 3-chloropyruvate and underwent a stereospecific α-hydroxyphosphonate-phosphate rearrangement to protected phosphoenol pyruvates. This sequence with oxygen isotope-labeled enantiomers represents an alternative approach to P-chiral [¹⁶O,¹⁷O,¹⁸O]phosphoenol pyruvates.

Full text of DOI:10.1080/10426507.2018.1452235

Phosphorus, Sulfur, and Silicon and the Related Elements
ISSN: 1042-6507 (Print) 1563-5325 (Online) Journal homepage: http://www.tandfonline.com/loi/gpss20
Formal synthesis of P-chiral
[16O,17O,18O]phosphoenol pyruvates by means
of the α-hydroxyphosphonate-phosphate
rearrangement
Petra Malová Križková, Alexander Roller & Friedrich Hammerschmidt
To cite this article: Petra Malová Križková, Alexander Roller & Friedrich Hammerschmidt
(2018): Formal synthesis of P-chiral [16O,17O,18O]phosphoenol pyruvates by means of the α-
hydroxyphosphonate-phosphate rearrangement, Phosphorus, Sulfur, and Silicon and the Related
Elements, DOI: 10.1080/10426507.2018.1452235
To link to this article: https://doi.org/10.1080/10426507.2018.1452235
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PHOSPHORUS, SULFUR, AND SILICON
https://doi.org/./..
Formal synthesis of P-chiral [O,O,O]phosphoenol pyruvates by means of the
α-hydroxyphosphonate-phosphate rearrangement
a
Petra Malová Križkováa, Alexander Rollerb, and Friedrich Hammerschmidt
aFaculty of Chemistry, Institute of Organic Chemistry, University of Vienna, Vienna, Austria; bFaculty of Chemistry, Institute of Inorganic Chemistry,
University of Vienna, Vienna, Austria
ABSTRACT
ARTICLE HISTORY
Received  January 
Accepted  March 
Transesterification of tris(hexafluoroisopropyl) phosphite with racemic 3-methyl-1-phenyl-butane-
1,3-diol gave two isomeric hexafluoroisopropyl-substituted 1,2,3-dioxaphosphinanes. These cyclic
phosphites were hydrolyzed rapidly and enantioselectively by water catalyzed by HCl. The respec-
tive metalated H-phosphonates were added to ethyl 3-chloropyruvate and underwent a stereospe-
cific α-hydroxyphosphonate-phosphate rearrangement to protected phosphoenol pyruvates. This
sequence with oxygen isotope-labeled enantiomers represents an alternative approach to P-chiral
[16O,17O,18O]phosphoenol pyruvates.
KEYWORDS
α-Hydroxyphosphonate-
phosphate rearrangement;
cyclic H-phosphonate;
tris(hexafluoroisopropyl)
phosphite; phosphoenol
pyruvate; hydrolysis
GRAPHICAL ABSTRACT
Introduction
Phosphoenol pyruvate (PEP) is a high energy compound in liv-
ing cells produced in the glycolytic cycle. It is used by pyru-
vate kinase to regenerate ATP from ADP. The phosphorus atom
in PEP is a pro-pro-chiral center, which can be made chiral by
replacing one of its three equivalent 16O atoms by an 17O atom
and a second one by an 18O atom.[1,2] In the past many pro-
chiral, pro-pro-chiral phosphorus centers and even the pro-pro-
pro-chiral center of inorganic phosphate[3,4] have been made
chiral by virtue of the oxygen isotopes alone or in combination
Scheme . Synthesis of the enantiomers of P-chiral PEP 1.
with sulfur. These compounds have been used to unravel the
stereochemistry of enzymes forming and breaking P-O bonds.
However, P-chiral PEP has never been prepared by chemi- enantiomers of P-chiral phosphoenol pyruvate 1 as potassium
cal methods directly. Lowe et al. developed a method to cre- salts (Scheme 1).[7] In order to summarize the approach, it
ate a chiral phosphoryl group and attach it to alcohols and started with diol (R)-[18O1]2 of ee > 99%, which was converted
ADP.[5] Knowles et al. accessed it as an intermediate in a reac- to a roughly 1:1 cis/trans-mixture of cyclic H-phosphonates
tion sequence from P-chiral phosphonopyruvate by means of [17O,18O1]4 via phosphoramidites [18O1]3 formed in a ratio of
PEP phosphomutase catalyzing the intramolecular migration of at least 1:3.
the phosphorus atom from the carbon to the oxygen atom.[6]
The mixture of silylated H-phosphonates was allowed to
We have recently reported a direct chemical synthesis of the react with ethyl 3-chloropyruvate (Perkow reaction). The
CONTACT Friedrich Hammerschmidt
friedrich.hammerschmidt@univie.ac.at
Institute of Organic Chemistry, Währingerstrasse , A- Vienna, Austria.
Color versions of one or more of the figures in the article can be found online at www.tandfonline.com/GPSS.
Supplemental data for this article can be accessed on the publisher’s website at https://doi.org/./...
©  Taylor & Francis Group, LLC

2
P. M. KRIŽKOVÁ ET AL.
Scheme . Conversion of ,-diol ( )-2 to cyclic H-phosphonates ( )-4.
Scheme . Transesterification of tris(hexafluoroisopropyl) phosphite with ,-diol
)-2.
(
obtained cis/trans-mixture of protected phosphoenol pyruvates
[17O,18O1]5 was separated by flash chromatography and the
diastereomers were deprotected individually (only one is given
in Scheme 1) to give the enantiomers of P-chiral PEP 1 as potas-
sium salts.
We envisaged to improve the stereoselectivity for the forma-
tion of H-phosphonates [17O,18O1]4 from diol 2 and to shorten
the synthesis by building on the α-hydroxyphosphonate-
phosphate rearrangement[8,9] in place of the Perkow reaction to
generate the protected phosphoenol pyruvates.
the phenyl and hexafluoroisopropoxy group, are trans-arranged
(Figure 1).
Worth mentioning are the differences in bond lengths for the
P-O single bonds influenced by the fluorine atoms; P1-O1 =
˚
˚
1.60836(10) A; P1-O2 = 1.67053(10) A. The values for P2 are
almost the same. The O-C bond lengths vary also. Last but not
least the deviations of X-O-Y angles from 120° for O3 and O6
(both higher than 125°) should be noted.
Consequently, the isomer must be cis-( )-7. Their ratio of
the isomers is determined by the solvent and the reaction tem-
perature. It ranged from 3.7:1 for CH2Cl2 at –78°C with warm-
ing to room temperature in favor of trans-( )-7 to 0.8:1 in THF
at room temperature in favor of cis-( )-7. The trans-isomer is
assumed to be the thermodynamically more stable one.[10,11] To
test the thermal configurational stability of trans-( )-7, a small
sample was bulb-to bulb distilled (80°C / 1 mbar). This isomer is
configurationally stable at 80°C during the short period of time
needed for vacuum distillation, as the distillate was still homoge-
neous. Hydrolysis was significant during flash chromatography
on silica gel as evidenced by loss of product. The cis-isomer is
more labile than the trans-isomer. Hydrolysis of ( )-7 in THF
containing water was minimal, but addition of TMSCl releas-
ing HCl upon reaction with water increased it very much. Thus,
when a mixture of cis/trans-( )-3 was stirred in THF contain-
ing 3 equiv. of water and 0.5 equiv. of TMSCl at room tempera-
ture for 30 min, no starting material was detected in the reaction
mixture by TLC (Scheme 4).
Results and discussion
For simplicity, we performed all experiments with racemic diol
( )-2 and derivatives thereof as the enantiomers will behave
similarly. It is known that 1,3-diols of type 2 react with phospho-
rus(III) compounds such as PCl3 in the presence of a tertiary
amine, (RO)3P, and (R2N)3P to give cyclic phosphorochlo-
ridites, phosphites, and phosphoramidites ( )-6, ( )-7, and
( )-3, respectively (Scheme 2). The position of X relative to Ph
in 3, 6 and 7 depends very much on X. For X = Cl and OR the
trans-isomer (Ph = equatorial, X = axial) is thermodynamically
more stable than the cis-isomer (Ph and X = equatorial). How-
ever, for X = NR2 it is the other way round.[10,11] The exchange
of X for OH with H2O and H217O in the labeled series, has to be
performed in the presence of an amine for X = Cl to neutralize
the acid and for X = OR, NR2 to speed up the reaction rate.
For 4 the cis-isomer is thermodynamically more stable than
the trans-isomer.[10] Importantly, H-phosphonates tautomerize
slowly in the presence of water under neutral conditions, but
rapidly under acidic conditions.[12] As phosphorochloridites are
unstable and difficult to purify, we decided to study cyclic phos-
phites 7 with X = OCH(CF3)2. It was hoped that its isomers can
be separated and that they are more amenable to hydrolysis than
analogs with ordinary alkoxy groups.
Concentration of the reaction mixture after addition of
1,1,1,3,3,3-hexamethyldisilazane for neutralization of the acid,
Tris(hexafluoroisopropyl) phosphite[13] was prepared and
handled under exclusion of moisture and oxygen, as it is easily
hydrolyzed to the H-phosphonate and oxidized to the respective
phosphate. Compared to trialkyl phosphites exchange of hex-
afluoroisopropoxy groups in the presence of Et3N proceeded
already easily below or at room temperature with diol ( )-
2 to give a mixture of two diastereomeric cyclic phosphites
7 (Scheme 3). When the reaction was performed in CH2Cl2
at room temperature, two isomeric compounds were formed.
They were separated by HPLC and the less polar isomer deliv-
ered crystals from hexane with a low melting point (37–38°C),
which were amenable to X-ray structure analysis. It proved
that the 1,3-substituents in the six-membered ring of ( )-7,
Figure . Crystal structure of cyclic phosphite trans-( )-7 drawn with % displace-
ment ellipsoids. The second molecule from the asymmetric unit was omitted for
clarity.

PHOSPHORUS, SULFUR, AND SILICON
3
Scheme . Acid-catalyzed hydrolysis of cyclic phosphites ( )-7.
followed by flash chromatography furnished a cis/trans-mixture
of H-phosphonates ( )-4 in 69% yield. Importantly, when
trans-( )-7 predominated in the starting material by 2:1,
H-phosphonate cis-( )-4 was formed preferentially (2.5:1). This
Scheme . Preparation of protected PEP by using the α-hydroxyphosphate-
result indicated at least a stereoselective inversion of configura- phosphate rearrangement.
tion upon hydrolysis. In order to study the behavior of individ-
substitution of chloride could be detected in the crude product
ual homogeneous samples of cis- and trans-isomer, they were
hydrolyzed under the same conditions as their mixture before.
As expected, cis-( )-7 gave preferentially trans-( )-4 and vice
versa, but both in admixture with a small amount of the respec-
tive isomer. It is not clear whether pseudorotation[14] interfered
with the hydrolytic process or whether partial epimerization of
phosphites[15] ( )-7 or H-phosphonates ( )-4 at phosphorus
under the acidic conditions caused the formation of the respec-
tive isomer. In any case, hydrolysis did not proceed stereospecif-
ically to deliver just one isomer as envisioned. However, it can be
used to prepare oxygen isotope-labeled H-phosphonates easily,
if H216O is replaced by H218O or H217O.
by 1H NMR spectroscopy.
The 1,3-diol ( )-2 was also present in the crude product
(cis-enol phosphate / trans-enol phosphate / 1,3-diol ( )-2 =
1:1:1) and was removed by flash chromatography. The yield of
cis- and trans-enol phosphate increased to acceptable 25% and
23%, respectively. Repetition of the experiment delivered the
same yields. To evaluate the stereoselectivity of the reaction, the
homogeneous H-phosphonates were similarly allowed to react
with n-BuLi and ethyl 3-chloropyruvate. H-Phosphonate cis-
( )-5 containing 1% of trans-isomer furnished less polar enol
phosphate trans-( )-5 in 60% yield [crude product: trans-( )-5
/ 1,3-diol ( )-2 = 1.0:0.30] in admixture with 1% of cis-( )-5.
However, homogeneous trans-H-phosphonate gave only the
more polar enol phosphate cis-( )-5 in 56% yield [crude prod-
uct: cis-( )-5 / 1,3-diol ( )-2 = 1.0:0.37 by 1H NMR]. There-
fore, lithiated H-phosphonates do not epimerize under the reac-
tion conditions and are added to the α-carbonyl group of ethyl
3-chloropyruvate with retention of configuration at the phos-
phorus atom. The subsequent rearrangement of the depro-
tonated α-hydroxyphosphonates follows a retentive course.
Satisfyingly, each H-phosphonate undergoes a stereospecific
reaction.
For convenience, the mixture of diastereomeric H-
phosphonates ( )-4 needed for the following experiments
was prepared by transesterification[8] of 1,3-diol ( )-2 with
bis(2,2,2-trifluoroethyl) phosphite and not by hydrolysis of
phosphites ( )-7. The first experiment was performed with
ethyl 3-bromopyruvate and DBU as base (pKa in CH3CN:
23.9[16]) in DMF at ambient temperature (Scheme 5). DBU
deprotonated the H-phosphonates (pKa in iPrOH: 17.4[17]),
which attacked the α-carbonyl group of pyruvate. The mixture
of the cis/trans-hydroxyphosphonates ( )-8 formed under-
went
a DBU-catalyzed α-hydroxyphosphonate-phosphate
rearrangement[8,9] with expulsion of Br-. The two enol phos-
phates could be separated by flash chromatography and their
spectra were identical to those of the literature.[7] Although the
Conclusion
yields for the cis- and trans isomer (21% and 9%) were low, it We have shown that cyclic cis- and trans-phosphites derived
was worth testing n-BuLi as base and ethyl 2-chloropyruvate[18] from 1,3-diol ( )-2 and tris(hexafluoroisopropyl) phos-
as the carbonyl compound at lower temperature. n-BuLi phite are formed and easily hydrolyzed to give cyclic H-
converted the mixture of H-phosphonates (cis/trans 1.2:1) phosphonates. These convert ethyl 3-chloropyruvate medi-
to the corresponding lithium salts at –78°C quantitatively. ated by n-BuLi in a stereospecific α-hydroxyphosphonate-
Addition of 3-chloropyruvate induced the formation of the phosphate rearrangement to protected PEP. When an 18O-
lithiated hydroxyphosphonates ( )-8, which rearranged upon labeled enantiomer of 1,3-diol 2 is utilized for the prepara-
slowly warming the reaction mixture to –25°C in the cooling tion of cyclic phosphites 7 and H217O for their hydrolysis,
bath. Neither H-phosphonate nor phosphonate derived from the enantiomers of cylic H-phosphonates cis- and trans-
3-chloropyruvate and lithiated phosphonate by nucleophilic [17O,18O1]4 will result. These can be transformed into protected

4
P. M. KRIŽKOVÁ ET AL.
(R)- and (S)-[16O,17O,18O]phosphoenol pyruvate by the α-
hydroxyphosphonate-phosphate rearrangement. Deprotection
by the literature protocol will afford the P-chiral phosphoenol
pyruvates. The presented sequence is a formal synthesis of (R)-
and (S)-[16O,17O,18O]phosphoenol pyruvate.
/ CH2Cl2, 5:1; TLC with heptane / CH2Cl2, 4:1; trans: Rf = 0.52,
cis: Rf = 0.38) allowed the separation of the isomers, but with
losses in yields, especially for the more polar cis-isomer.
trans-( )-7: IR (ATR): ν = 1375, 1271, 1211, 1191, 1167,
1
1105, 1020, 1003, 965 cm–1. H NMR (400.27 MHz, CDCl3):
δ = 7.39-7.28 (m, 5H), 5.51 (ddd, J = 12.0, 3.2, 2.1 Hz, 1H),
4.61 (septd, J = 6.4, 5.9 Hz, 1H), 2.20 (dd, J = 14.3, 12.0 Hz,
1H), 1.87 (ddd, J = 14.3, 3.2, 2.1 Hz, 1H), 1.64 (s, 3H), 1.35 (s,
3H). 13C NMR (150.93 MHz, CDCl3): δ = 140.7 (d, J = 3.0 Hz),
128.6 (2C), 128.2, 125.7 (2C), 121.3 (q with fine structure, J =
283.4 Hz, 2C), 78.0 (d, J = 9.1 Hz), 70.0 (septd, J = 33.4, 23.8 Hz),
68.7 (d, J = 3.0 Hz), 46.3 (d, J = 3.5 Hz), 32.5 (d, J = 4.5 Hz), 28.0.
31P NMR (162.03 MHz, CDCl3): δ = 132.0 (sept, J = 7.2 Hz).
Anal. Calcd for C14H15F6O3P (376.23): C, 44.69; H, 4.02. Found:
C, 44.78; H, 4.22.
cis-( )-7: IR (ATR): ν = 2917, 1375, 1290, 1228, 1197, 1107,
1057, 968 cm–1. 1H NMR (600.25 MHz, CD2Cl2): δ = 7.34-7.23
(m, 5H), 5.24 (td, J = 12.6, 3.3 Hz, 1H), 4.73 (septd, J = 7.4,
6.0 Hz, 1H), 2.69 (dd, J = 14.9, 12.6 Hz, 1H), 1.87 (td, J = 14.9,
3.3 Hz, 1H), 1.47 (s, 3H), 1.35 (s, 3H). 13C NMR (150.93 MHz,
CD2Cl2): δ = 141.4, 128.6 (2C), 128.1, 126.1 (2C), 121.5 (q with
fine structure, J = 284.5 Hz, 2C), 77.5 (d, J = 7.6 Hz), 74.0 (d, J
= 7.4 Hz), 70.4 (septd, J = 33.7, 27.2 Hz), 44.6 (d, J = 15.7 Hz),
31.2, 28.1 (d, J = 1.5 Hz). 31P NMR (242.99 MHz, CD2Cl2): δ =
132.3 (sept, J = 7.9 Hz).
Experimental
1H, 13C (J-modulated) and 31P NMR spectra were recorded
in CDCl3 and CD2Cl2 with a Bruker Avance AV III 400 (1H:
400.27 MHz, 13C: 100.65 MHz, 31P: 162.03 MHz) and with a
Bruker Avance AV III 600 (1H: 600.25 MHz, 13C: 150.93 MHz,
31P: 242.94 MHz) spectrometer at 25°C. Chemical shifts (δ) are
reported in parts per million (ppm) relative to CHCl3 / CDCl3
(δ 7.24, δ 77.0) and CHDCl2 / CD2Cl2 (δ 5.24, δ 53.5) and
H
C
H
C
external H3PO4 (85%; δ 0.0); coupling constants (J) are given
P
in Hz. Data for 1H NMR spectra are reported as follows: chem-
ical shift, multiplicity (s = singlet, d = doublet, t = triplet, q =
quartet, sept = septet, m = multiplet), coupling constants, and
integration. IR spectra were recorded with a Bruker VERTEX 70
IR Spectrometer in ATR mode. HPLC was performed either on
a Jasco System with a PU-980 pump, UV 975 and RI 930 detec-
tor (in case of analytical determination) or on a Dynamix Model
SD-1 with a UV-1 absorbance detector (in case of semiprepara-
tive separation using Nucleosil 50–5 column, Ø 4.6 cm × 25 cm).
Melting points were measured with a Leica Galen III Thermovar
instrument and are uncorrected. Small amounts of liquids were
added to reaction mixtures by using a μL syringe.
Test of thermal configurational stability of trans-( )-7
Flash (column) chromatography was performed with silica
gel 60 (230–400 mesh) and monitored by TLC, conducted on
glass-backed 0.25 mm thick silica gel 60 F254. Spots were visu-
alized by UV and / or dipping the plate into a solution of
(NH4)6Mo7O24 • 4 H2O (23.0 g) and Ce(SO4)2 • 4 H2O (1.0 g)
in 10% aqueous H2SO4 (500 mL), followed by heating with a
heat gun.
A small sample (50 mg) of trans-( )-7 was bulb to bulb distilled
1
(80°C / 1 mbar). The H and 31P NMR spectrum recorded in
toluene-d8 showed that the distilled sample was still homoge-
neous and did not contain cis-( )-7.
Hydrolysis of cyclic phosphites ( )-7 – cis- and trans-( )-
4,4-dimethyl-6-phenyl-1,3,2-dioxaphosphinane-2-oxides
[cis- and trans-( )-4]
cis- and trans-( )-2-[(1,1,1,3,3,3-Hexafluoropropan-2-
yl)oxy]-4,4-dimethyl-6-phenyl-1,3,2-dioxaphosphinane
[cis- and trans-( )-7]
A mixture of phosphites cis- and trans-( )-7 (0.75 g, 2.0 mmol,
A mixture of diol ( )-2 (0.36 g, 2 mmol), tris(1,1,1,3,3,3- trans/cis, 2:1), H2O (7.2 mmol, 0.13 mL) and TMSCl (0.11 g,
hexafluoro-2-propyl) phosphite[13] (1.117 g, 2.1 mmol, sensi- 1.0 mmol, 0.12 mL) in dry THF (5 mL) was stirred at
tive to oxygen and moisture!) and Et3N (1 mmol, 0.14 mL) room temperature under argon for 30 min. 1,1,1,3,3,3-
in dry CH2Cl2 (8 mL) was stirred under argon for 40 min at Hexamethyldisilazane (0.18 g, 1.1 mmol, 0.14 mL) was added
room temperature and then concentrated under reduced pres- (under argon!) and the solution was concentrated under
sure (trans/cis, 2:1, by 31P NMR). The crude product was puri- reduced pressure. The crude product was dried (45°C / 1 mbar)
fied by bulb to bulb distillation (65–100°C / 1.4 mbar) to yield a for 5 min and then purified by flash chromatography (EtOAc,
mixture of cyclic phosphites trans- and cis-( )-7 (0.669 g, 89%) Rf = 0.50) to yield a crystalline mixture (0.31 g, 69%; cis/trans,
1
as a colorless oil. The isomers were separated by semiprepara- 2.5:1 by H NMR) of cyclic H-phosphonates cis- and trans-4.
tive HPLC; injection of 1.8 mL of solution (25 mg of mixture The NMR spectra (1H, 31P) were identical to those of the
of cis/trans-( )-7/mL); 10% CH2Cl2 in hexanes (40 mL/min); literature.[8]
trans-( )-7: tr = 2.32 min, mp 37–38°C (hexanes); cis-( )-7: tr
Similarly, phosphite cis-( )-7 (0.15 g, 0.40 mmol) was con-
verted to H-phosphonate trans-( )-4 [0.07 g, 78%, contained
= 2.77 min, colorless oil.
Alternatively, a rapid flash chromatography (heptane / 10% of cis-( )-4] as colorless solid.
EtOAc, 7:1, trans: Rf = 0.69, cis: Rf = 0.58) gave a mixture with
Similarly, phosphite trans-( )-3 (0.42 g, 1.12 mmol) was
the same ratio of the isomers as in the crude product with mini- converted to H-phosphonate cis-( )-4 [0.23 g, 91%, contained
mal loss of product. A standard flash chromatography (heptane 6% of trans-( )-4] as colorless solid.

PHOSPHORUS, SULFUR, AND SILICON
5
Preparation of enol phosphates ( )-5 using DBU as base
References
Ethyl 3-bromopyruvate (0.14 g, 90 μL, 0.72 mmol) and DBU
(0.21 g, 0.20 mL, 2 equiv.) were added to a solution of H-
phosphonates cis- and trans-( )-4 (0.16 g, 0.71 mmol, cis/trans,
1.3:1) dissolved in dry DMF (4 mL) at 0°C under argon. The
dark colored reaction mixture was stirred for 30 min and con-
centrated under reduced pressure. The residue was diluted with
CH2Cl2 (5 mL), washed with water (2 × 3 mL), dried (Na2SO4)
and concentrated under reduced pressure. The residue was
flash chromatographed (hexane / EtOAc, 2:1, trans-( )-5: Rf =
0.34; cis-( )-5: Rf = 0.20) to yield enol phosphate trans-( )-5
(0.022 g, 9%) and cis-( )-5 (0.050 g, 21%) as colorless oils. The
NMR spectra of the two compounds were identical to the ones
reported in the literature.[7]
[1] Floss, H. G.; Tsai, M.-D.; Woodard, R. B. Stereochemistry of Biologi-
cal Reactions at Proprochiral Centers. Top. Stereochem. 1984, 15, 253–
321.
[2] Eckstein, F. Nucleoside Phosphorothioates. Annu. Rev. Biochem.
1995, 54, 367–402. doi:10.1146/annurev.bi.54.070185.002055.
[3] Tsai, M.-D.; Chang, T.-T. Chirality at a Pro-Pro-Prochiral Phos-
phorus Center. Stereochemical Course of the 5ꢀ-Nucleotidase-
Catalyzed Reaction. J. Am. Chem. Soc. 1980, 102, 5416–5419.
doi:10.1021/ja00536a061.
[4] Tsai, M.-D. Use of 31P(18O), 31P(17O), and 17O NMR Methods to
Study Enzyme Mechanisms Involving Phosphorus. Methods Enzy-
mol. 1982, 87, 235–279. doi:10.1016/S0076-6879(82)87017-1.
[5] (a) Cullis, P. M.; Lowe, G. Lowe, G.; Potter, B. V. L. The Stereochem-
istry of 2-Substituted-2-Oxo-4,5-Diphenyl-1,3,2-Dioxaphospholans
and the Related Chiral [16O,17O,18O]Phosphate Monoesters. J. Chem.
Soc., Chem. Commun. 1981, 245; (b) Lowe, G.; Potter, V. L. The Stere-
ochemical Course of Yeast Hexokinase-Catalysed Phosphoryl Trans-
fer by Using Adenosine 5ꢀ[gamma(S)-16O,17O,18O]Triphosphate as
Substrate. Biochem. J. 1981, 199, 227–233.
Preparation of enol phosphates ( )-5 using n-BuLi as base
n-BuLi (1.12 mL, 2.5 M solution in hexane; 2.8 mmol, 1.1
equiv.) was dropwise added to a solution of the mixture of H-
phosphonates ( )-4 (0.576 g, 2.55 mmol, cis/trans, 1.2:1) in dry
THF (7 mL) under argon at –78°C. After stirring for 3 min,
the solution of ethyl 3-chloropyruvate[18] (0.422 g, 2.8 mmol,
1.1 equiv.) in dry THF (3 mL) was added quickly. The reac-
tion mixture was allowed to warm slowly in the cooling bath.
When the temperature had risen to –25°C (within 2 h), AcOH
(five small drops) was added. The reaction mixture was concen-
trated under reduced pressure. CH2Cl2 (25 mL) and a saturated
aqueous solution of NaHCO3 (20 mL) were added. The organic
phase was separated and the aqueous one was extracted with
CH2Cl2 (2 × 10 mL). The combined organic layers were washed
with water (10 mL), dried (Na2SO4) and concentrated under
reduced pressure. After recording 1H and 31P NMR spectra of
the residue (cis-enol phosphate / trans-enol phosphate / 1,3-diol
( )-2, 1:1:1), it was flash chromatographed (CH2Cl2 / acetone,
20:1, trans: Rf = 0.50; cis: Rf = 0.33; 1,3-diol ( )-2: Rf = 0.16) to
yield enol phosphates cis-( )-5 (0.221 g, 25%) and trans-( )-
5 (0.200 g, 23%), both as colorless crystals, and 1,3-diol ( )-
2 (0.080 g, 18%) as gum. The NMR spectra of the three com-
pounds were identical to the ones reported in the literature.[7,19]
Similarly, H-phosphonate cis-( )-4 (0.440 g, 1.95 mmol,
contained 1% of trans-isomer) was converted to homogeneous
enol phosphate trans-( )-5 (0.395 g, 60%).
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[7] Malová Križková, P.; Prechelmacher, S.; Roller, A.; Ham-
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Chem. Eur. J. 2015, 21, 10200–10206. doi:10.1002/chem.201406661.
[9] Philippitsch, V.; Hammerschmidt, F. Rearrangement of Lithiated
S-alkyl O,O-Dialkyl Thiophosphates: Scope and Stereochemistry
of the Thiophosphate—Mercaptophosphonate Rearrangement. Org.
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[10] Verkade, J. G. Interplay of Steric and Electronic Influences in the
Chemistry of Monocyclic and Bicyclic Phosphorus Esters. Phospho-
rus Sulfur 1976, 2, 251–281. doi:10.1080/03086647608078956.
[11] Bentrude, W. G.; Tan, H. W. Conformations of Saturated Phosphorus
Heterocycles. Possible pπ -dπ Overlap Effects on the Apparent Con-
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=
[12] Janesko, B. G.; Fisher, H. C.; Bridle, M. J.; Montchamp, J.-L. P( O)H
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[13] Sakatsume, O.; Yamane, H.; Takaku, H.; Yamamoto, N. Use of
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Nucleic Acids Res. 1990, 18, 3327–3331. doi:10.1093/nar/18.11.3327.
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of Configuration in Acyclic Phosphite Triesters. Inorg. Chim. Acta
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Similarly, homogeneous H-phosphonate trans-( )-4
(0.534 g, 2.36 mmol) was converted to homogeneous enol
phosphate cis-( )-5 (0.447 g, 56%).
[16] Galezowski, W.; Jarczewski, A.; Stanczyk, M.; Brzezinski, B.; Bartl,
F.; Zundel, G. Homoconjugated Hydrogen Bonds with Ami-
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Pudovik, A. N. Properties of Dialkyl Phosphites and Their Reactivity
in the Reaction with Bezalacetophenone. Doklady Chem. 1987, 292,
21–25.
[18] Stekol, J. A. The Synthesis of S-Benzylthiopyruvic Acid and its Con-
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[19] Behnke, D.; Hennig, L.; Findeisen, M.; Welzel, P.; Mueller, V.; Thor-
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1081–1095. doi:10.1016/S0040-4020(00)00005-3.
Acknowledgements
The authors thank V. Philippitsch for performing the α-phosphonate-
phosphate rearrangement with DBU as base, S. Felsinger for recording
NMR spectra, E. Macoratti for the preparative HPLC separation, and J.
Theiner for combustion analyses.
Funding
Austrian Science Fund (P19869-N19).
ORCID
Friedrich Hammerschmidt http://orcid.org/0000-0003-2193-1405