Synthesis and characterization of enantiomerically pure cis- and trans-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-oxides as acetylcholine mimetics and inhibitors of acetylcholinesterase

Article abstract of DOI:10.1002/hlca.201100507

The title compounds, the P(3)-axially and P(3)-equatorially substituted cis- and trans-configured 7-benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-oxides (=7-benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphabicyclo[4.4.0]decane 3-oxides=5-benzyl-2-fluorohexahydro-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-oxides) were prepared (ee>99%) and fully characterized (Schemes 2 and 4). The absolute configurations were established from that of their precursors, the enantiomerically pure cis- and trans-1-benzyl-3-hydroxypiperidine-2-methanols which were unambiguously assigned. Being configuratively fixed and conformationally constrained phosphorus analogues of acetylcholine, they mimic rotamers of acetylcholine and are suitable probes for the investigation of molecular interactions with acetylcholinesterase. As determined by kinetic methods, the compounds are irreversible inhibitors of the enzyme displaying significant stereoselectivity. Copyright

Full text of DOI:10.1002/hlca.201100507

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Helvetica Chimica Acta – Vol. 95 (2012)
Synthesis and Characterization of Enantiomerically Pure cis- and trans-3-
Fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-Oxides as Acetylcholine Mimetics
and Inhibitors of Acetylcholinesterase
by Michael Wꢀchter and Peter Rꢁedi*
Organisch-chemisches Institut der Universitꢀt Zꢁrich, Winterthurerstrasse 190, CH-8057 Zꢁrich
(phone: þ 41-44-8215579)
The title compounds, the P(3)-axially and P(3)-equatorially substituted cis- and trans-configured
7-benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-oxides (¼ 7-benzyl-3-fluoro-2,4-dioxa-7-aza-3-
phosphabicyclo[4.4.0]decane 3-oxides ¼ 5-benzyl-2-fluorohexahydro-4H-1,3,2-dioxaphosphorino[5,4-b]-
pyridine 2-oxides) were prepared (ee > 99%) and fully characterized (Schemes 2 and 4). The absolute
configurations were established from that of their precursors, the enantiomerically pure cis- and trans-1-
benzyl-3-hydroxypiperidine-2-methanols which were unambiguously assigned. Being configuratively
fixed and conformationally constrained phosphorus analogues of acetylcholine, they mimic rotamers of
acetylcholine and are suitable probes for the investigation of molecular interactions with acetylcholi-
nesterase. As determined by kinetic methods, the compounds are irreversible inhibitors of the enzyme
displaying significant stereoselectivity.
1. Introduction. – In preceding reports, we have presented the synthesis and
characterization of the enantiomerically pure P(3)-axially and P(3)-equatorially
substituted cis- and trans-configured 3-fluoro-2,4-dioxa-9-aza-3-phospha- (type I) [1],
3-fluoro-2,4-dioxa-8-aza-3-phospha- (type II) [2], and 3-fluoro-2,4-dioxa-3-phospha-
decalin 3-oxides (type IV) [3] (Scheme 1). Continuing our investigations on the
inhibition of serine hydrolases (chymotrypsin, acetylcholinesterase) by decalin-type
organophosphates, we discuss the preparation and characterization of the isomeric
enantiomerically pure 7-benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-oxides
(¼ 7-benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphabicyclo[4.4.0]decane 3-oxides ¼ 5-ben-
zyl-2-fluorohexahydro-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-oxides 13 and 14
(type III, cf. Schemes 1 and 4)) [4]. The compounds are mimetics of rotamers of
acetylcholine (¼2-(acetyloxy)-N,N,N-trimethylethanaminium; ACh) and as such are
considered to be suitable probes for the investigation of molecular interactions with
acetylcholinesterase (AChE) [5] and the stereochemical course of the inhibition
reaction by 31P-NMR spectroscopy [4][6 – 8].
2. Synthesis and Characterization of the Precursors. – 2.1. Preparation of the
Enantiomerically Pure (þ)- and (ꢀ)-trans- and (þ)- and (ꢀ)-cis-1-Benzyl-3-hydroxy-
piperidine-2-methanols ((þ)- and (ꢀ)-4, and (þ)- and (ꢀ)-5, resp.). Following the
protocol for the preparation of the racemic compounds [9], 3-hydroxypyridine-2-
methanol (1) was converted by a six-step reaction sequence followed by chromato-
graphic separation (SiO2) of the diastereoisomers to the key intermediates (ꢁ)-trans-
ꢂ 2012 Verlag Helvetica Chimica Acta AG, Zꢁrich

Helvetica Chimica Acta – Vol. 95 (2012)
717
Scheme 1
and (ꢁ)-cis-[3-(acetyloxy)-1-benzylpiperidin-2-yl]methyl 2,2-dimethylpropanoates
((ꢁ)-2 and (ꢁ)-3; Scheme 2). Preparative HPLC (Chiralcelꢃ OD) afforded the
optically active diesters (þ)-2 (k’ ¼ 1.68, ee > 99%), (ꢀ)-2 (k’ ¼ 2.08, ee > 99%), (þ)-3
(k’ ¼ 0.74, ee > 99%), and (ꢀ)-3 (k’ ¼ 1.01, ee > 99%). Hydrolysis of the respective
diesters gave the 1-benzyl-3-hydroxypiperidine-2-methanols (þ)-4 ([a]D ¼ þ43.5, ee >
99%), (ꢀ)-4 ([a]D ¼ ꢀ43.7, ee > 99%), (þ)-5 ([a]D ¼ þ21.3, ee > 99%), and (ꢀ)-5
([a]D ¼ ꢀ21.5, ee > 99%) (Scheme 2)1)
2.2. The Absolute Configurations of the Diols (þ)- and (ꢀ)-4, and (þ)- and (ꢀ)-5.
The absolute configurations of the diols (þ)- and (ꢀ)-4, and (þ)- and (ꢀ)-5 were
1
inferred by the high-field H-NMR application of the Mosher method [10] and its
extension to the bis-MTPA derivatives 6 – 9 as recently reported [11]. Esterification of
(ꢀ)- and (þ)-4 with (þ)-(S)-MTPA-Cl (¼(þ)-(S)-a-methoxy-a-(trifluoromethyl)ben-
zeneacetyl chloride) afforded the bis-(R)-esters 6a and 7a, and the corresponding bis-
(S)-esters 6b and 7b were isolated after reaction of (ꢀ)- and (þ)-4 with (ꢀ)-(R)-
MTPA-Cl. The same procedure was performed with (ꢀ)- and (þ)-5 to yield the bis-(R)-
esters 8a and 9a, and the bis-(S)-esters 8b and 9b (Scheme 2).
1
)
The [a]D values were determined in EtOH (c ¼ between 0.55 and 1.28), ee > 99%.

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Helvetica Chimica Acta – Vol. 95 (2012)
Scheme 21)
a) H2, Rh/Alox, 60 bar, H2O, pH 3, 358. b) BnBr, K2CO3, EtOH, reflux. c) tBuCOCl, pyridine, ꢀ208.
d) (COCl)2, DMSO, CH2Cl2, ꢀ608. e) NaBH4, EtOH, ꢀ158 (cis/trans 1:1). f) Ac2O, pyridine, r.t. g) CC
(SiO2, hexane/AcOEt) (a) – g) as in Scheme 4 in [9]). h) Prep. HPLC (Chiralcelꢃ OD, hexane/(ꢁ)-2-
BuOH). i) Prep. HPLC (Chiralcelꢃ OD, hexane/iPrOH). j) KOH, EtOH/H2O, r.t. k) (ꢀ)-(R)- or (þ)-
(S)-MTPA-Cl, resp., Et3N, N,N-dimethylpyridin-4-amine (DMAP), CH2Cl2, r.t. (note: (R)-MTPA-Cl
yields the (S)-MTPA ester and vice versa). l) CC (SiO2, hexane/Et2O).
1
In the trans-series, the analysis of the H-NMR data led to the assignment of the
absolute configurations at C(3) as (3R) and (3S) for (þ)-4 and (ꢀ)-4, respectively. The
results were self-contained, and the Dd values (¼d(S) – d(R)) of the diagnostically
relevant signals were consistent: for the couple 6a/6b, we observed Dd(HꢀC(2)) ¼
ꢀ0.16, Dd(HeqꢀC(4)) ¼ þ0.15, and Dd(CH2(5)) ¼ ꢀ0.05, and the inverse was found
for the couple 7a/7b (see Exper. Part)2). The fact that Dd(HꢀC(3)) ¼ 0 for the couples
6a/6b and 7a/7b is indicative for the ideal conformation of the MTPA moiety3) and the
reliability of the experiment. As a consequence, the absolute configurations of the
2
)
)
Since 6a ¼ ent-7b and 6b ¼ ent-7a, 6a (2’R,2R,3S) and 7b (2’S,2S,3R), as well as 6b (2’S,2R,3S) and
7a (2’R,2S,3R) have identical NMR spectra.
The theoretical prerequisite for the success and reliability of the H-NMR Mosher experiments is
1
3
that the MTPA moiety adopts an idealized conformation where the H-atom at the decisive
stereogenic center C(3), the C¼O, and the CF3 groups lie in the same plane in a relative syn-
arrangement [10]. Therefore, an essential quality factor for the experiment is the equal chemical
shift (Dd ¼ 0) for HꢀC(3) in both the (R)- and the (S)-MTPA derivatives.

Helvetica Chimica Acta – Vol. 95 (2012)
719
trans-3-hydroxypiperidine-2-methanols (þ)-4 and (ꢀ)-4 were assigned as (2S,3R), and
(2R,3S), respectively.
In the conformationally flexible cis-series 8a/8b and 9a/9b, the reliability of the
interpretation was a priori reduced due to Dd(HꢀC(3)) ¼ꢁ 0.072 for the couples 8a/8b
and 9a/9b. According to the 1H-NMR spectra, the MTPAꢀO group at C(3) is axial, and
the magnitudes of the respective vicinal couplings suggested that the conformation is
not an ideal chair4). However, the Dd values of the diagnostically relevant signals were
as consistent as in the trans-series: Dd(HꢀC(2)) ¼ þ0.04, Dd(CH2(4)) ¼ ꢀ0.09, and
Dd(CH2(5)) ¼ ꢀ0.07 for the couple 8a/8b, and vice versa for 9a/9b (see Exper. Part).
The low quality factor (Dd(HꢀC(3)) seems to reflect the sterical impact of the
MTPAꢀOCH2 moiety at C(2) and the conformational uncertainty. However, taking the
consistency of the Dd values into account, the absolute configurations of the cis-3-
hydroxypiperidine-2-methanols (þ)-5 and (ꢀ)-5 were inferred as (3S) and (3R), and as
a consequence, as (2S,3S), and (2R,3R), respectively5).
Further support for this assignment was provided by a modified Mosher approach:
Esterification of (ꢁ)-10 with (þ)-(S)-MTPA-Cl and preparative HPLC separation
(Chiralcelꢃ OD) afforded the 2-[(pivaloyloxy)methyl]-substituted (R)-MTPA esters
11 and 12 (Scheme 3). The 1H-NMR analysis of 11 and 12 showed the MTPAꢀO group
at C(3) to be equatorial6), and the quality factor (Dd(HꢀC(3)) ¼ þ0.022) was indica-
tive for a nearly idealized conformation of the MTPA moiety and the enhanced reliabil-
ity of the experiment. As depicted in Fig. 1, the arrangement of the bulky substituents
in 12 only resulted in an small paramagnetic shift of HꢀC(2) (Dd ¼ ꢀ0.01), whereas
the H-atoms in the sphere of influence of the (R)-MTPA phenyl group in 11 were
shielded with respect to 12, in particular CH2(4) (Dd(HaxꢀC(4))¼ þ0.07, Dd(HeqꢀC(4)) ¼
þ0.03)7). The identity of the parent diols was verified by hydrolysis of the
diastereoisomers 11 and 12 that afforded (ꢀ)-5 and (þ)-5, respectively (Scheme 3).
Hence, the absolute configurations were in accord with the previous assignment.
Fig. 1. Shielding effects of the (R)-MTPA phenyl group in 11 and 12
3
3
3
4
5
)
HꢀC(2) is equatorial, HꢀC(3) is axial (9a: J(2,3) ¼ 4.0, J(3,4ax) ¼ 7.8, and J(3,4eq) ¼ 4.0 Hz; 9b:
3J(2,3) ¼ 4.0, J(3,4ax) ¼ 7.6, and J(3,4eq) ¼ 4.0 Hz).
3
3
)
Subtle differential reasoning, including the shielding effects exerted on HaxꢀC(6), HeqꢀC(6) and
PhCH2 (HA, HB), suggests the most probable conformation. Although it deviates from the ideal
arrangement [10], the Mosher experiment can be interpreted in terms of the absolute configuration
as indicated [4].
3
3
3
6
7
)
HꢀC(2) is equatorial, HꢀC(3) is axial (11: J(2,3) ¼ 3.3, and J(3,4ax) ¼ 6.6, and J(3,4eq) ¼ 3.3 Hz;
3
3
3
12: J(2,3) ¼ 3.3, and J(3,4ax) ¼ 6.9, and J(3,4eq) ¼ 3.3 Hz).
)
For the reason of simplicity, chair conformations are depicted for 11 and 12 although the vicinal
couplings suggest that they deviate from ideal chairs. However, this imprecision does not affect the
basic argumentation.

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Scheme 3
a) (þ)-(S)-MTPA-Cl, Et3N, N,N-dimethylpyridin-4-amine (DMAP), CH2Cl2, r.t. b) Prep. HPLC
(Chiralcelꢃ OD, hexane/iPrOH). c) KOH, EtOH/H2O.
Conclusive evidence for the absolute configuration of (þ)- and (ꢀ)-4, and (þ)- and
(ꢀ)-5 was provided by the X-ray crystallographic analysis (Sect. 3.3) of the 7-aza-3-
phosphadecalins (ꢀ)-13a (Fig. 2) and (þ)-14a (Fig. 3). This is the first account of the
full characterization of the isomeric 1-benzyl-3-hydroxypiperidine-2-methanols 4 and 58).
3. Synthesis and Characterization of the 7-Benzyl-3-fluoro-2,4-dioxa-7-aza-3-
phosphadecalins. – 3.1. 2,4-Dioxa-7-aza-3-phosphadecalin 3-Oxides 13 and 14. Apply-
ing our reliable protocol [9], the enantiomerically pure trans-7-benzyl-3-fluoro-2,4-
dioxa-7-aza-3-phosphadecalins 13 (Scheme 4) were prepared from (þ)- or (ꢀ)-4 by
reaction with POCl2F and chromatographic separation of the resulting P(3)-epimer
mixture (axial/equatorial ca. 1.5 :1) into the pure axial epimers (þ)- and (ꢀ)-13a and
equatorial epimers (þ)- and (ꢀ)-13b. Similarly, starting from (þ)- or (ꢀ)-5, the cis-7-
benzyl-3-fluoro-2,4-dioxa-9-aza-3-phosphadecalins (þ)- and (ꢀ)-14a, and (þ)- and
(ꢀ)-14b were obtained (Scheme 4)9). The NMR data of the 7-aza-3-phosphadecalins
8
)
The (2S,3R)-configuration has been assigned to the trans-configured 1-benzyl- and 1-methyl-3-
hydroxypiperidine-2-methanols [12] but no chiroptical data were reported. In connection with
investigations on 3-hydroxypipecolic acid analogues, piperidine alkaloids, and aza-carbohydrates,
the closely related 3-hydroxypiperidine-2-methanols [13], 1-butyl-3-hydroxypiperidine-2-methanols
[14], and tert-butyl 3-hydroxy-2-(hydroxymethyl)piperidine-1-carboxylates [15] have been de-
scribed. In the cis-series, the signs of the optical rotation are consistent and equal to those of (þ)-
and (ꢀ)-5 ((2S,3S): [a]D > 0; (2R,3R): [a]D < 0), whereas they differ in the trans-series. The latter
fact might be due to conformational impacts as tert-butyl (2S,3R)-3-hydroxy-2-(hydroxymethyl)pi-
peridine-1-carboxylate adopts the unexpected diaxial arrangement of the substituents [16].
However, the only report that presents all 4 stereoisomers [14] contains several inconsistencies:
The enantiomeric trans-1-butyl-3-hydroxypiperidine-2-methanols are both dextrorotatory ((2R,3S):
[a]D ¼ þ2.7; (2S,3R): [a]D ¼ þ13.3), and the cis-1-butyl-3-hydroxypiperidine-2-methanols have
inverted signs ((2R,3R): [a]D > 0; (2S,3S): [a]D < 0) with respect to all other accounts [13][15]. See
also our discussions on configurational and chiroptical inconsistencies in the 1-substituted 4-
hydroxypiperidine-3-methanols [11] and the 3-hydroxypiperidine-4-methanols [17].
9
)
The [a]D values were determined in acetone (c ¼ between 0.26 and 0.38), ee > 99%.

Helvetica Chimica Acta – Vol. 95 (2012)
Scheme 49)
721
a) Cl2P(O)F, Et3N, Et2O, < 08. b) CC (SiO2, hexane/Et2O). c) Cl2P(O)F, Et3N, CH2Cl2, < 08. d) CC
(SiO2, hexane/AcOEt).
13 and 14 (see Exper. Part) exhibited the same essential features as the type-I [1], type-
II [2], and type-IV [3] congeners (Scheme 1). In particular, the 31P-NMR spectra
confirmed the relative configuration at the P-atom, the double-chair conformations of
the axial epimers 13a and 14a, and distorted conformations of the 2,4-dioxa-3-phospha
moiety in the equatorial epimers 13b and 14b10). Due to the strongly electronegative F-
10
)
Generally, the 31P-NMR resonance of the axial epimer is shifted upfield with respect to the
equatorial one, and the chemical-shift difference (Dd ¼ deq – dax) is > 0, and its magnitude is
inversely proportional to the electronegativity of the substituent at the P-atom. However, the cyclic
phosphorofluoridates of the cis-series of the type I – IV compounds (Scheme 1, L ¼ F) displayed
Dd < 0, a fact that is only explained by significant conformational changes. The magnitude of the
1
3J(P,H) in the H-coupled 31P-NMR is indicative of the conformation of the 2,4-dioxa-3-phospha
moiety: Diagnostically relevant values for the axial epimers were 3J(P,HeqꢀC(5)) ꢂ 25 Hz and
3
3J(P,HaxꢀC(5)) ꢂ 0, whereas the equatorial ones displayed 3J(P,HaxꢀC(5)) ꢂ J(P,HeqꢀC(5)) ꢂ 10 –
15 Hz. Hence, the axial epimers exhibited a d-type and the equatorial ones a m-type splitting
pattern (see [13]).

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Helvetica Chimica Acta – Vol. 95 (2012)
substituent, the chemical shift difference (Dd ¼ deq – dax) was small in the trans-couple
13a/13b (Dd ¼ þ0.9 ppm) and negative in the cis-couple 14a/14b (Dd ¼ ꢀ0.3 ppm) as
discussed earlier [18].
3.2. Conformations of 13 and 14 in Solution. As previously presented [1][2][6 – 8]
and directly evidenced [18], stereoelectronic (anomeric) effects predominantly
determine the conformation of the 3-substituted 2,4-dioxa-3-phosphadecalin 3-oxides.
In the 3-axially substituted 3-fluoro-3-phosphadecalins 13a and 14a, both the steric and
the stereoelectronic effects act in the same direction. According to the vicinal couplings
(3J(P,HeqꢀC(5)) ¼ 25.3 Hz (13a) and 25.7 Hz, (14a))10), these compounds adopt the
double-chair conformation C-111)12) (Scheme 5).
Scheme 5
11
)
)
The short terms for the conformations (IUPAC convention) were introduced in [18]: C ¼ chair, B ¼
boat, E ¼ envelope, TB ¼ twist-boat (see also [1][2]).
12
There is no evidence for the completely inverted C-2 conformation in 14a. We assume that the
anomeric preference prevents this interconversion.

Helvetica Chimica Acta – Vol. 95 (2012)
723
Generally, the steric and the stereoelectronic effects are opposite in the P(3)-
equatorially substituted compounds 13b and 14b. Although the chair conformation is
sterically favored, the anomeric preference of the F-substituent to move into an axial
position results in nonchair conformations such as boat or twist-boats (i.e., B, TB-1, and
TB-2)11) of the 2,4-dioxa-3-phospha moiety (Scheme 5)13). Based on our detailed
conformational studies on the type-III inhibitors [4][18][19], in particular, according to
the vicinal coupling data (3J(P,HaxꢀC(5)) ¼ 14.5, 3J(P,HeqꢀC(5) ¼ 8.5, and
3
3J(P,HꢀC(1)) ꢂ 1 Hz) and the virtual invariability of J(P,HeqꢀC(5)) in low-temper-
ature 31P-NMR experiments [4][18][19], we conclude that the conformation of the
trans-configured epimers (þ)- and (ꢀ)-13b is an equilibrium mixture of TB-2
(predominant) and C-1 (Scheme 5)14).
In the cis-configured equatorial epimers (þ)- and (ꢀ)-14b, the flexibility of the decalin
system combined with the anomeric preference of the P(3)-equatorial F-substituent
render the situation significantly more complex, and additional conformations must be
envisaged. In particular, the bicyclic system can undergo complete ring inversion to yield
the prominent C-2 arrangement (Scheme 5), which seems to be favored by both the
anomeric and the steric effects. According to crystal structures (see [18], and Figs. 2 and
3), the piperidine moiety adopts the chair conformation with the N-benzyl group in the
equatorial position. Although its steric impact is not reliably predictable, it certainly
affects the equilibrium population of the conformers, and it would interfere with a
complete ring inversion to C-2. The diagnostically relevant 3J(P,HꢀC(1)) ¼ 12.8,
3
3J(P,HaxꢀC(5)) ¼ 14.0, and J(P,HeqꢀC(5)) ¼ 10.9 Hz (see Exper. Part) and the interpre-
tation of our modified Karplus equation [18] clearly evidenced that the vicinal couplings
are averaged, and none of the conformations depicted in Scheme 5 can be excluded, nor
any can be prominently assigned15). Hence, 14b is in solution a complex mixture of the
coexisting conformers C-1/C-2, and/or TB-1/TB-2, and also B and envelope E.
A full account of our detailed conformational studies by variable temperature 31P-
NMR experiments and X-ray analyses on the 3-fluoro-2,4-dioxa-3-phosphadecalin 3-
oxides (type IV) and the 7-benzyl-3-fluoro-2,4-dioxa-7-aza congeners (type III) will be
presented in a following report [19].
3.3. Crystallographic Analyses of (ꢀ)-13a and (þ)-14a16). The structures of (ꢀ)-13a
and (þ)-14a were solved and refined successfully. Both compounds in the crystal were
13
)
)
In such systems, the resulting minimum-energy conformations represent a balance between the
anomeric effect favoring the axial orientation in the twist-boat and the 1,3-steric and eclipsing
interactions favoring the chair conformation. This fact explains the unusual stabilization of nonchair
conformations.
Since 3J(P,HꢀC(1)) ꢂ 1, conformations B and TB-1 are excluded. At room temperature, the 31P-
NMR spectra were well resolved and coalescence phenomena were not observed, i.e., the
interconversion C-1 > TB-2 is fast on the NMR time scale. By lowering the temperature, 13b tends
to adopt the thermodynamically significantly stabilized TB-2 conformation, i.e., at sufficiently low
temperatures, TB-2 will freeze out.
14
15
16
)
)
The most striking argument for the exclusion of a prominent conformation is that none of the
3J(P,H) ꢂ 0 [18].
The full data sets are summarized in Table 2 (see Exper. Part). CCDC-857940 ((ꢀ)-13a) and CCDC-
857941 ((þ)-14a) contain supplementary crystallographic data. These can be obtained free of
charge from the Cambridge Crystallographic Data Centre via www.code.cam.ac.uk/data_request/cif.

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Helvetica Chimica Acta – Vol. 95 (2012)
enantiomerically pure, and the absolute configurations of the molecules were
determined independently by the diffraction experiments. The refinement of the
absolute structure parameter confidently confirmed that the refined coordinates
(Table 2, see Exper. Part) represent the true enantiomorphs with the expected
(1S,3S,6R)-configuration for (ꢀ)-13a (Fig. 2), and the (1S,3S,6S)-configuration for (þ)-
14a (Fig. 3). These results independently corroborated the configurational assignments
of the precursors (ꢀ)-(2R,3S)-1-benzyl-3-hydroxypiperidine-2-methanol ((ꢀ)-4) and
(ꢀ)-(2S,3S)-1-benzyl-3-hydroxypiperidine-2-methanol ((þ)-5) by means of the high-
1
field H-NMR Mosher method and our unusual extension to bis-MTPA derivatives
[11]. As discussed above, the six-membered ring containing the P-atom has an
undistorted chair conformation with the F-atom in the axial position, and the N-benzyl
group occupies the sterically favored equatorial position.
Since all attempts to obtain suitable crystals of the equatorial epimers (þ)- or (ꢀ)-
13b and/or (þ)- or (ꢀ)-14b were not successful, no X-ray crystallographic analysis of an
optically active equatorial type-III compound is available17).
4. Enzyme Kinetics. – 4.1. General. The inhibitory potency and the mode of action
of the enantiomerically pure 3-fluoro-7-aza-3-phosphadecalins 13 and 14 was
Fig. 2. Molecular Structure of (ꢀ)-13a ((1S,3S,6R)). Trivial atom numbering; 50% probability ellipsoids.
17
)
In contrast to this finding, the racemic equatorial epimers (ꢁ)-13b and (ꢁ)-14b could be suitably
crystallized [9], and their X-ray crystallographic analyses provided the direct evidence of the
anomeric effect [18].

Helvetica Chimica Acta – Vol. 95 (2012)
725
Fig. 3. Molecular Structure of (þ)-14a ((1S,3S,6S)). Trivial atom numbering; 50% probability ellipsoids.
determined according to the general considerations and procedure explicitely
described in the precedent reports [1][2]. In particular, the data acquisition was based
on the Ellman assay [20], and the mathematical evaluation was performed according to
[21]. For irreversible inhibition, the simplified overall processes were considered that
directly yield the covalently phosphorylated enzyme E – I (Scheme 6, mechanism a) or
involve a preceding reversible step, resulting in the formation of an associative
enzyme – inhibitor complex E · I*, followed by the irreversible phosphorylation step
(Scheme 6, mechanism b) [1][2].
4.2. Data Analysis and Results. The determination of the apparent rate constants
(kobs) from the progress curves (A ¼ f(t), see Eqn. 1 in the Exper. Part) and the
mathematical evaluation of the dependence kobs ¼ f([I]) to evaluate the irreversible

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Helvetica Chimica Acta – Vol. 95 (2012)
Scheme 6
inhibition parameters (KD, ka, ki, and kp, resp.) were performed according to [21]18).
The mathematical equations (Eqns. 1 – 3) underlying Scheme 6 and further details are
summarized in the Exper. Part. For all the P(3)-axially substituted 3-fluoro-7-aza-3-
phosphadecalins ((ꢁ)-13a, (þ)- and (ꢀ)-13a, and (þ)- and (ꢀ)-14a), the secondary plot
(kobs ¼ f([I])) exhibited a linear dependence, and inhibition mechanism a was assigned
to these compounds. In the case of (þ)- and (ꢀ)-13b and (þ)- and (ꢀ)-14b, the
secondary plot depended hyperbolically upon [I], and mechanism b was assigned to the
the P(3)-equatorially substituted congeners. The experimental results are summarized
in Table 1.
All the investigated 3-fluoro-7-aza-3-phosphadecalins (type III) inhibited AChE
irreversibly. Compared to diisopropyl phosphorofluoridate (P(O)F(OiPr)2) that was
used as the standard reference, they were moderate inhibitors. But with respect to the
related 9-aza- (type I) [1] and 8-aza-3-phosphadecalins (type II) [2], they were
significantly stronger. With the exception of (þ)- und (ꢀ)-14a that did not differ in the
inhibition behavior, the type-III compounds displayed pronounced diastereoselectivity,
the (3R)-configured isomers being roughly twice as potent than the P(3)-epimers.
Furthermore, the data of 13 again confirmed our finding that the inhibitory activity of a
racemic inhibitor is approximately the arithmetic mean of the enantiomers [4]19).
18
)
Recently, we have presented a novel, integrated approach with a reappraisal of kinetic mechanisms
and diagnostic methods that enables distinct as well as subtle differentiations of generalized
inhihibition mechanisms [22]. Although the assay data of the (þ)- and (ꢀ)-7-benzyl-3-fluoro-2,4-
dioxa-7-aza-3-phosphadecalin 3-oxides 13 and 14 were evaluated by the novel method, too [4], we
present the results of the simplified procedure [20] for reasons of conformity and direct comparison
with the preceding reports in this series [1][2].
19
)
Although apparently obvious, it cannot be concluded a priori that the inhibition constants are
simply additive.

Helvetica Chimica Acta – Vol. 95 (2012)
727
Table 1. Kinetic Data of the Inhibition of AChE with the Enantiomerically Pure 7-Aza-3-fluoro-2,4-dioxa-
3-phosphadecalins (þ)- and (ꢀ)-13 and (þ)- and (ꢀ)-14. P(O)F(OiPr)2 as reference.
Kinetic Parameters
Mechanisma)
Kinetic Parameters
Mechanisma)
(þ)-13a ka ¼ 11.7 ꢁ 0.4 mꢀ1sꢀ1
(ꢀ)-13a ka ¼ 6.0 ꢁ 0.2 mꢀ1sꢀ1
(ꢁ)-13a ka ¼ 8.6 ꢁ 0.2 mꢀ1sꢀ1
(þ)-13b ki ¼ 56.7 ꢁ 1.2 mꢀ1sꢀ1
KD ¼ 103 ꢁ 2 mm
a
a
a
b
(þ)-14a
(ꢀ)-14a
(þ)-14b
ka ¼ 5.1 ꢁ 0.2 mꢀ1sꢀ1
ka ¼ 5.1 ꢁ 0.3 mꢀ1sꢀ1
ki ¼ 123.3 ꢁ 3.3 mꢀ1sꢀ1
KD ¼ 100 ꢁ 5 mm
a
a
b
kp ¼ 0.0075 ꢁ 0.0001 sꢀ1
ki ¼ 48.3 ꢁ 0.5 mꢀ1sꢀ1
KD ¼ 57 ꢁ 2 mm
kp ¼ 0.0059 ꢁ 0.0002 sꢀ1
(ꢀ)-13b ki ¼ 105.0 ꢁ 2.3 mꢀ1sꢀ1
KD ¼ 97 ꢁ 2 mm
(ꢀ)-14b
b
a
b
kp ¼ 0.070 ꢁ 0.002 sꢀ1
kp ¼ 0.010 ꢁ 0.001 sꢀ1
P(O)F(OiPr)2 ka ¼ 245.0 ꢁ 7.3 mꢀ1sꢀ1
a) see Scheme 6.
5. Remarks. – Besides the main object of our research project, the investigation of
the stereochemical course of the inhibition reaction by 31P-NMR spectroscopy [4][6 –
8], kinetic studies are a valuable tool to study molecular interactions of acetylcholine
(ACh) with AChE. Being mimetics of rotamers of ACh, the 7-benzyl-3-fluoro-2,4-
dioxa-8-aza-3-phosphadecalin 3-oxides 13 and 14 (type-III inhibitors) are supposed to
be suitable probes to impart knowledge on the physiologically active (recognition)
conformation of ACh in the course of the hydrolysis. This enzymatic process is a nearly
diffusion controlled, highly complex reaction cascade where all steps are governed by
conformational changes of both the enzyme and the substrate20). Meanwhile, we have
evaluated the kinetic data of the 3-fluoro substituted 2,4-dioxa-3-phosphadecalins of
the types I – IV (Scheme 1), but our experimental results still do not allow reliable
conclusions with respect to the effective inhibition mechanism (active site, peripheral
or other conformatively induced irreversible binding at another nucleophilic site).
Considering the fact that our compounds differ from the natural substrate and have
additional structural features that significantly could influence both the steric demand
and the basicity of the compound, the situation is even more complicated, and
conclusions of evidencing a recognition conformation from the inhibitory data would
remain rather speculative. However, it is not clear to what extent this fact is due to the
simplified kinetic approach (see Scheme 6) and the respective mathematical treatment
[21]. A complete re-interpretation of the kinetic data of the types I – IV according to
our integrated approach [22]18) and the full comparison of the respective data sets and
their interpretation is in process and will be presented in a following, concluding report
[23].
The authors are indebted to PD Dr. A. Linden, head of the X-ray department of our institute for the
high-quality X-ray crystallographic analyses. The financial support of the project by the Swiss National
Science Foundation is gratefully acknowledged.
20
)
For a brief discussion of the recognition conformation of ACh, see [1].

728
Helvetica Chimica Acta – Vol. 95 (2012)
Experimental Part
1. General. See [1][7][9]. For the particular precautions in preparing and handling the organo-
phosphates, see [9]. Determination of ee: based on the integration of the peak areas of the anal. HPLC
separations of the precursor diols with optimized resolution (Rs > 4). NMR Assignments: based on
extensive 2D-NMR (see [9]) and selective 1H-decoupling experiments.
2. (þ)-(2S,3R)- and (ꢀ)-(2R,3S)-, and (þ)-(2S,3S)- and (ꢀ)-(2R,3R)-3-Hydroxy-1-(phenylmethyl)-
piperidine-2-methanol ((þ)- and (ꢀ)-4, and (þ)- and (ꢀ)-5, resp.). 2.1. [3-(Acetyloxy)-1-(phenyl-
methyl)piperidin-2-yl]methyl 2,2-Dimethylpropanoates 2 and 3. The racemic precursors, the (ꢁ)-trans-
and (ꢁ)-cis-diesters (ꢁ)-2 and (ꢁ)-3, resp., were prepared and characterized as described earlier [9].
Prep. HPLC of (ꢁ)-2 (Chiralcelꢃ OD, hexane/(ꢁ)-2-BuOH 200 :1; a ¼ 1.24, RS ¼ 4.95) afforded
enantiomerically pure (þ)-2 (k’ ¼ 1.68) and (ꢀ)-2 (k’ ¼ 2.08) as yellowish crystals. Prep. HPLC of (ꢁ)-3
(Chiralcelꢃ OD, hexane/iPrOH 100 :1; a ¼ 1.36, RS ¼ 6.4) furnished enantiomerically pure (þ)-3 (k’ ¼
0.74) and (ꢀ)-3 (k’ ¼ 1.01) as yellowish viscous oils. Colorless products were obtained by subjecting
(þ)- and (ꢀ)-2 and (þ)- and (ꢀ)-3 to CC (SiO2, (hexane/AcOEt 4 :1).
(þ)-(2S,3R)-[3-(Acetyloxy)-1-(phenylmethyl)piperidin-2-yl]methyl 2,2-Dimethylpropanoate ((þ)-
2)21): Colorless crystals. M.p. 44 – 468. Rf (hexane/AcOEt 4 :1) 0.25. [a]2D5 ¼ þ12.4 (c ¼ 1.15, EtOH,
ee > 99%).
(ꢀ)-(2R,3S)-[3-(Acetyloxy)-1-(phenylmethyl)piperidin-2-yl]methyl 2,2-Dimethylpropanoate ((ꢀ)-
2): [a]2D5 ¼ ꢀ12.2 (c ¼ 1.11, EtOH, ee > 99%). All other data identical with those of (þ)-2.
(þ)-(2R,3R)-[3-(Acetyloxy)-1-(phenylmethyl)piperidin-2-yl]methyl 2,2-Dimethylpropanoate ((þ)-
3)21): Colorless viscous oil. Rf (hexane/AcOEt 4 :1) 0.20. [a]2D5 ¼ þ9.1 (c ¼ 1.10, EtOH, ee > 99%).
(ꢀ)-(2S,3S)-[3-(Acetyloxy)-1-(phenylmethyl)piperidin-2-yl]methyl 2,2-Dimethylpropanoate ((ꢀ)-3):
[a]2D5 ¼ ꢀ9.0 (c ¼ 1.28, EtOH, ee > 99%). All other data identical with those of (þ)-3.
2.2. 3-Hydroxy-1-(phenylmethyl)piperidine-2-methanols 4 and 5. Saponification of the diester (þ)- or
(ꢀ)-2 or (þ)- or (ꢀ)-3 (each 500 mg, 1.44 mmol) in EtOH (10 ml) with KOH (600 mg) in H2O (15 ml) at
r.t. (3 h) and usual workup afforded the crude diol (þ)- or (ꢀ)-4, or (ꢀ)- or (þ)-5, resp., as yellowish
viscous oils. After boiling in EtOH over charcoal, the pure compounds were obtained as colorless crystals
(310 – 318 mg, 97 – 100%).
(þ)-(2S,3R)-3-Hydroxy-1-(phenylmethyl)piperidine-2-methanol ((þ)-4)22): Colorless prisms. M.p.
81 – 848. Rf (AcOEt) ca. 0.20 (tailing). [a]2D5 ¼ þ43.5 (c ¼ 1.05, EtOH, ee > 99%).
(ꢀ)-(2R,3S)-3-Hydroxy-1-(phenylmethyl)piperidine-2-methanol ((ꢀ)-4): [a]2D5 ¼ ꢀ43.7 (c ¼ 1.08,
EtOH, ee > 99%). All other data identical with those of (þ)-4.
(þ)-(2S,3S)-3-Hydroxy-1-(phenylmethyl)piperidine-2-methanol ((þ)-5)22): Colorless prisms. M.p.
75 – 778. Rf (AcOEt) ca. 0.18 (tailing). [a]2D5 ¼ þ21.3 (c ¼ 1.02, EtOH, ee > 99%).
(ꢀ)-(2R,3R)-3-Hydroxy-1-(phenylmethyl)piperidine-2-methanol ((ꢀ)-5): [a]2D5 ¼ ꢀ21.5 (c ¼ 0.55,
EtOH, ee > 99%). All other data identical with those of (þ)-5.
3. (R)- and (S)-MTPA Derivatives for the Determination of the Absolute Configuration. 3.1. Bis-
MTPA Esters 6a, 6b, 7a, 7b, 8a, 8b, 9a, and 9b. Each diol (ꢁ)- or (þ)-4, or (ꢀ)- or (þ)-5 (each 17.5 mg,
0.08 mmol) was dissolved in anh. CH2Cl2 (3 ml), and Et3N (46 ml, 0.32 mmol) and N,N-dimethylpyridin-
4-amine (DMAP; 2 mg) were added under Ar. The mixture was treated with (þ)-(S)-MTPA-Cl (34 ml,
0.18 mmol). After stirring for 1 h at r.t. under Ar, the mixture was subjected to CC (SiO2, hexane/Et2O
1:9): (R)-MTPA diester 6a, 7a, 8a, or 9a. Analogously, (ꢀ)- or (þ)-4, or (ꢀ)- or (þ)-5 was treated with
(ꢀ)-(R)-MTPA-Cl to yield the (S)-MTPA diester 6b, 7b, 8b, or 9b. All MTPA derivatives were isolated in
pure form as colorless, viscous oils: 6a (47 mg, 90%), 6b (48 mg, 92%), 7a (49 mg, 94%), 7b (48 mg,
92%), 8a (50 mg, 95%), 8b (51 mg, 98%), 9a (48 mg, 92%), and 9b (50 mg, 95%).
(2S,3R)-1-(Phenylmethyl)-2-{[(2R)-3,3,3-trifluoro-2-methoxy-1-oxo-2-phenylpropoxy]methyl}pi-
peridin-3-yl (aR)-a-Methoxy-a-(trifluoromethyl)benzeneacetate (Bis-(R)-MTPA ester; 7a) from (þ)-4:
1
Rf (AcOEt) 0.69. H-NMR (500 MHz, CDCl3): 7.56 – 7.50 (m, 4 arom. H); 7.42 – 7.33 (m, 6 arom. H);
7.26 – 7.12 (m, 5 arom. H); 5.05 (M of ABXM, ddd, 3J(3,2) ¼ 6.8, 3J(3,4ax) ¼ 7.6, 3J(3,4eq) ¼ 4.2,
21
)
)
The spectral data were identical with those of (ꢁ)-2 and (ꢁ)-3, resp., see [9].
The spectral data were identical with those of (ꢁ)-4 and (ꢁ)-5, resp., see [9].
22

Helvetica Chimica Acta – Vol. 95 (2012)
729
3
2
HꢀC(3)); 4.65, 4.46 (AB of ABXM, 2J ¼ 12.2, J ¼ 4.3, 3.8, CH2(OMTPA)); 3.89, 3.32 (AB, J ¼ 13.2,
PhCH2); 3.54, 3.51 (2q, 5J(Me,F) ¼ 1.1, MeO); 2.85 (X of ABXM, 3J(2,3) ¼ 6.8, 3J(2,CH2) ¼ 4.3, 3.8,
HꢀC(2)); 2.63 (ddd, 2J ¼ 10.5, 3J(6eq,5eq) ¼ 6.6, 3J(6eq,5ax) ¼ 3.5, HeqꢀC(6)); 2.13 (ddd, 2J ¼ 11.8,
3J(6ax,5ax) ¼ 8.2, 3J(6ax,5eq) ¼ 3.3, HaxꢀC(6)); 2.05 (ddt, 2J ¼ 11.3, 3J(4eq,3) ¼ 3J(4eq,5ax) ¼ 4.2,
3J(4eq,5eq) ¼ 8.0, HeqꢀC(4)); 1.60 (m, w1/2 ꢂ 15, HeqꢀC(5)); 1.51 (m, w1/2 ꢂ 20, HaxꢀC(4)); 1.41 (m, w1/2
ꢂ 20, HaxꢀC(5)). 13C-NMR (75 MHz, CDCl3): 166.6, 165.7 (CO of MTPA); 138.5 (C(1’)), 132.1, 132.0
(C(1’’), C(1’’’)); 129.7 (C(4’’), C(4’’’)); 129.6 (C(2’), C(6’)); 128.5 (C(2’’), C(2’’’), C(6’’), C(6’’’)); 128.2
(C(3’), C(5’)); 127.4 (C(3’’), C(3’’’), C(5’’), C(5’’’)); 127.3 (C(4’)); 123.4, 123.3 (2q, 1J(C,F) ¼ 289, CF3);
84.8, 84.7 (2q, 2J(C,F) ¼ 27.8, PhC(OMe)(CF3)CO); 72.3 (C(3)); 62.1 (C(2)); 61.6 (PhCH2); 57.9 (C(8));
55.5, 55.3 (MeO); 48.8 (C(6)); 27.4 (C(4)); 20.9 (C(3)). 19F{1H}-NMR (376.5 MHz, CDCl3): ꢀ 70.46, ꢀ
71.65 (2s, CF3).
(2S,3R)-1-(Phenylmethyl)-2-{[(2S)-3,3,3-trifluoro-2-methoxy-1-oxo-2-phenylpropoxy]methyl}piper-
idin-3-yl (aS)-a-Methoxy-a-(trifluoromethyl)benzeneacetate (Bis-(S)-MTPA ester; 7b) from (þ)-4: Rf
(AcOEt): 0.69. 1H-NMR (500 MHz, CDCl3): 7.60 – 7.47 (m, 4 arom. H); 7.40 – 7.31 (m, 6 arom. H); 7.26 –
7.18 (m, 3 arom. H); 7.16 – 7.12 (m, 2 arom. H); 5.05 (M of ABXM, td, 3J(3,2) ¼ 3J(3,4ax) ¼ 8.3,
2
3
3J(3,4eq) ¼ 4.2, HꢀC(3)); 4.70, 4.05 (AB of ABXM, J ¼ 12.3, J ¼ 3.2, 3.0, CH2(OMTPA)); 3.98, 3.17
(AB, 2J ¼ 13.4, PhCH2); 3.55 (br. s, 2 MeO); 2.72 (m, br. q-like, w1/2 ꢂ 18, HeqꢀC(6)); 2.69 (X of ABXM,
not resolved, HꢀC(2)); 2.20 (m, td-like, w1/2 ꢂ 20, HeqꢀC(4)); 2.03 (m, td-like, w1/2 ꢂ 25, HaxꢀC(6)); 1.64
(qd-like, w1/2 ꢂ 18, HeqꢀC(5)); 1.56 – 1.41 (m, HaxꢀC(4), HaxꢀC(5)). 13C-NMR (75 MHz, CDCl3): 166.4,
165.4 (CO of MTPA); 138.4 (C(1’)), 132.5, 132.2 (C(1’’), C(1’’’)); 129.7 (C(4’’), C(4’’’)); 128.7 (C(2’),
C(6’)); 128.5 (C(2’’), C(2’’’), C(6’’), C(6’’’)); 128.3 (C(3’), C(5’)); 127.4 (C(3’’), C(3’’’), C(5’’), C(5’’’));
1
127.1 (C(4’)); 123.4 (q, J(C,F) ¼ 289, CF3); 84.8, 84.4 (2q, 2J(C,F) ¼ 27.8, PhC(OMe)(CF3)CO); 72.3
(C(3)); 63.1 (C(2)); 61.6 (PhCH2); 57.9 (C(8)); 55.6, 55.2 (MeO); 50.1 (C(6)); 28.7 (C(4)); 22.7 (C(3)).
19F{1H}-NMR (376.5 MHz, CDCl3): ꢀ 71.52, ꢀ 71.58 (2s, CF3)).
Dd(1H) ¼ d(S) – d(R) (in Hz): HꢀC(2), ꢀ 80; HꢀC(3), 0; HaxꢀC(4), ꢀ 10; HeqꢀC(4), þ 75;
HaxꢀC(5), þ 35; HeqꢀC(5), þ 20, HaxꢀC(6), ꢀ 50; HeqꢀC(6), þ 45 ! (3R)-configuration.
(R)- and (S)-MTPA Esters 6a and 6b from (ꢀ)-4. Being enantiomeric compounds, 6a and 7b (6a ¼
ent-7b), as well as 6b and 7a (6b ¼ ent-7a), exhibited identical NMR spectra, only the sign of Dd was
inverted: Dd(1H) ¼ d(S) – d(R) (in Hz): HꢀC(2), þ 80; HꢀC(3), 0; HaxꢀC(4), þ 10; HeqꢀC(4), ꢀ 75;
HaxꢀC(5), ꢀ 35; HeqꢀC(5), ꢀ 20; HaxꢀC(6), þ 50, HeqꢀC(6), ꢀ 45 ! (3S)-configuration.
(2S,3S)-1-(Phenylmethyl)-2-{[(2R)-3,3,3-trifluoro-2-methoxy-1-oxo-2-phenylpropoxy]methyl}piper-
idin-3-yl (aR)-a-Methoxy-a-(trifluoromethyl)benzeneacetate (Bis-(R)-MTPA ester; 9a) from (þ)-5: Rf
(AcOEt): 0.70. 1H-NMR (500 MHz, CDCl3): 7.55 – 7.48 (m, 4 arom. H); 7.41 – 7.36 (m, 6 arom. H); 7.28 –
7.18 (m, 5 arom. H); 5.30 (M of ABXM, dt, 3J(3,2) ¼ 3J(3,4eq) ¼ 4.0, 3J(3,4ax) ¼ 7.8, HꢀC(3)); 4.49, 4.22
(AB of ABXM, 2J ¼ 11.8, 3J ¼ 6.4, 4.2, CH2(OMTPA)); 3.65, 3.42 (AB, 2J ¼ 14.0, PhCH2); 3.54, 3.48 (2q,
3
3
2
5J(Me,F) ¼ 1.1, MeO); 3.17 (X of ABXM, J(2,3) ¼ 4.0, J(2,CH2) ¼ 6.4, 4.2, HꢀC(2)); 2.56 (ddd, J ¼
11.5, 3J(6eq,5eq) ¼ 7.7, 3J(6eq,5ax) ¼ 3.6, HeqꢀC(6)); 2.31 (ddd, 2J ¼ 11.5, 3J(6ax,5ax) ¼ 7.1,
3J(6ax,5eq) ¼ 4.0, HaxꢀC(6)); 1.80 (m, w1/2 ꢂ 20, CH2(4)); 1.58 (m, w1/2 ꢂ 70, CH2(5)). 13C-NMR
(75 MHz, CDCl3): 166.3, 165.7 (CO of MTPA); 139.0 (C(1’)); 132.3, 132.0 (C(1’’), C(1’’’)); 129.7
(C(4’’), C(4’’’)); 128.5 (C(2’), C(6’)); 128.2 (C(2’’), C(2’’’), C(6’’’), C(6’’)); 128.1 (C(3’), C(5’)); 127.3, 127.2
(C(3’’), C(3’’’), C(5’’), C(5’’’)); 127.0 (C(4’)); 123.4, 123.3 (2q, 1J(C,F) ¼ 289, CF3); 84.8, 84.7 (2q,
2J(C,F) ¼ 27.8, PhC(OMe)(CF3)CO); 72.8 (C(3)); 63.2 (C(2)); 60.7 (PhCH2); 57.9 (C(8)); 55.4 (MeO);
47.5 (C(6)); 27.1 (C(4)); 21.1 (C(3)). 19F{1H}-NMR (376.5 MHz, CDCl3): ꢀ 71.45, ꢀ 71.46 (2s, CF3).
(2S,3S)-1-(Phenylmethyl)-2-{[(2S)-3,3,3-trifluoro-2-methoxy-1-oxo-2-phenylpropoxy]methyl}piperi-
din-3-yl (aS)-a-Methoxy-a-(trifluoromethyl)benzeneacetate (Bis-(S)-MTPA ester; 9b) from (þ)-5: Rf
(AcOEt): 0.70. 1H-NMR (500 MHz, CDCl3): 7.54 – 7.47 (m, 4 arom. H); 7.41 – 7.34 (m, 6 arom. H); 7.28 –
7.15 (m, 5 arom. H); 5.23 (M of ABXM, dt, 3J(3,2) ¼ 3J(3,4eq) ¼ 4.0, 3J(3,4ax) ¼ 7.6, HꢀC(3)); 4.66, 4.23
(AB of ABXM, 2J ¼ 11.7, 3J ¼ 6.5, 4.6, CH2(OMTPA)); 3.58, 3.43 (AB, 2J ¼ 14.1, PhCH2); 3.50, 3.49 (2q,
3
3
2
5J(Me,F) ¼ 0.9, MeO); 3.21 (X of ABXM, J(2,3) ¼ 4.0, J(2,CH2) ¼ 6.5, 4.6, HꢀC(2)); 2.56 (ddd, J ¼
11.9, 3J(6eq,5eq) ¼ 7.6, 3J(6eq,5ax) ¼ 3.3, HeqꢀC(6)); 2.32 (ddd, 2J ¼ 11.9, 3J(6ax,5ax) ¼ 6.6,
3J(6ax,5eq) ¼ 3.8, HaxꢀC(6)); 1.71 (m, w1/2 ꢂ 25, CH2(4)); 1.52 (m, w1/2 ꢂ 70, CH2(5)). 13C-NMR
(75 MHz, CDCl3): 166.4, 165.8 (CO of MTPA); 139.1 (C(1’)); 132.1, 132.0 (C(1’’), C(1’’’)); 129.7
(C(4’’), C(4’’’)); 128.5 (C(2’), C(6’)); 128.2 (C(2’’), C(2’’’), C(6’’), C(6’’’)); 128.1 (C(3’), C(5’)); 127.4, 127.2

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Helvetica Chimica Acta – Vol. 95 (2012)
(C(3’’), C(3’’’), C(5’’), C(5’’’)); 126.9 (C(4’)); 123.4, 123.3 (2q, 1J(C,F) ¼ 289, CF3); 84.7, 84.6 (2q,
2J(C,F) ¼ 27.8, PhC(OMe)(CF3)CO); 72.8 (C(3)); 63.0 (C(2)); 60.8 (PhCH2); 57.9 (C(8)); 55.5, 55.3
(MeO); 47.8 (C(6)); 26.8 (C(4)); 21.0 (C(3)). 19F{1H}-NMR (376.5 MHz, CDCl3): ꢀ 71.48, ꢀ 71.60 (2s,
CF3).
Dd(1H) ¼ d(S) – d(R) (in Hz): HꢀC(2), þ 20; HꢀC(3), ꢀ 35; HaxꢀC(4) ꢂ HeqꢀC(4), ꢀ 45;
HaxꢀC(5) ꢂ HeqꢀC(5), ꢀ 30; HaxꢀC(6), þ 5; HeqꢀC(6), 0 ! (3S)-configuration.
(R)- and (S)-MTPA Esters 8a and 8b from (ꢀ)-5. Being enantiomeric compounds, 8a and 9b (8a ¼
ent-9b) as well as 8b and 9a (8b ¼ ent-9a) exhibited identical NMR spectra, only the sign of Dd was
inverted: Dd(1H) ¼ d(S) ꢀ d(R) (in Hz): HꢀC(2), ꢀ 20; HꢀC(3), þ 35; HaxꢀC(4) ꢂ HeqꢀC(4), þ 45;
HaxꢀC(5) ꢂ HeqꢀC(5), þ 30; HaxꢀC(6), ꢀ 5; HeqꢀC(6), 0 ! (3R)-configuration.
3.2. [(Pivaloyloxy)methyl]-Substituted MTPA Esters 11 and 12. The soln. of (ꢁ)-10 (77 mg.
0.25 mmol, prepared according to [9]) in anh. CH2Cl2 (5 ml), Et3N (44 ml, 0.31 mmol), and N,N-
dimethylpyridin-4-amine (DMAP; 2 mg) was treated with (þ)-(S)-MTPA-Cl (55 ml, 0.30 mmol). After
stirring for 2 h at r.t. under Ar, the mixture was subjected to CC (SiO2, hexane/Et2O 1:9) to afford the
mixture of the diastereoisomeric (R)-MTPA esters 11/12 (120 mg, 92%). Prep. HPLC (Chiralcelꢃ OD,
hexane/iPrOH 150 :1; a ¼ 1.16, RS ¼ 1.6) afforded the diasteroisomers 11 and 12 (de > 90%) as slightly
yellowish oils. Saponification (cf. Sect. 2.2) of 11 yielded (ꢀ)-5, and from 12 we obtained (þ)-5
(< 95%).
(2R,3R)-1-Benzyl-2-[(2,2-dimethyl-1-oxopropoxy)methyl]piperidin-3-yl (aR)-a-Methoxy-a-(tri-
fluoromethyl)benzeneacetate ((R)-MTPA ester; 11): Rf (AcOEt) 0.67. k’ ¼ 1.19 (de ¼ 90%). 1H-NMR
(400 MHz, CDCl3): 7.51 – 7.46 (m, 2 arom. H); 7.35 – 7.31 (m, 3 arom. H); 7.24 – 7.13 (m, 5 arom. H); 5.26
(M of ABXM, dt, 3J(3,4ax) ¼ 6.6, 3J(3,2) ¼ 3J(3,4eq) ¼ 3.3, HꢀC(3)); 4.39, 3.92 (AB of ABXM, 2J ¼ 11.5,
3J ¼ 6.3, 3J ¼ 5.6, CH2(OMTPA)); 3.73 3.39 (AB, 2J ¼ 14.1, PhCH2); 3.48 (q, 5J(Me,F) ¼ 1.0, MeO); 2.98
(X of ABXM, 3J(2,3) ¼ 3.3, 3J(2,CH2) ¼ 6.3, 5.6, HꢀC(2)); 2.59 (ddd, 2J ¼ 11.7, 3J(6eq,5eq) ¼ 6.2,
3J(6eq,5ax) ¼ 3.8, HeqꢀC(6)); 2.16 (ddd, 2J ¼ 11.7, 3J(6ax,5ax) ¼ 8.4, 3J(6ax,5eq) ¼ 3.5, HaxꢀC(6)); 1.77 (m,
w1/2 ꢂ 18, HeqꢀC(4)); 1.62 (m, w1/2 ꢂ 22, HaxꢀC(4)); 1.61 – 1.33 (m, CH2(5)); 1.12 (s, Me3C). 13C-NMR
(100 MHz, CDCl3): 178.0 (COCMe3); 165.9 (CO of MTPA); 139.3 (C(1’)); 132.0 (C(1’’)); 129.5 (C(4’’));
128.4 (C(2’), C(6’)); 128.2 (C(2’’), C(6’’)); 128.1 (C(3’), C(5’)); 127.3, 127.2 (C(3’’), C(5’’)); 126.8 (C(4’));
123.3 (q, 1J(C,F) ¼ 289, CF3); 84.6 (q, 2J(C,F) ¼ 27.6, PhC(OMe)(CF3)CO); 72.5 (C(3)); 61.8 (C(2)); 61.1
(PhCH2); 58.0 (C(8)); 55.3 (MeO); 49.2 (C(6)); 38.6 (Me3C); 27.3 (C(4)); 27.1 (Me3C); 20.7 (C(5)).
19F{1H}-NMR (376.5 MHz, CDCl3): ꢀ 71.65 (s, CF3).
(2S,3S)-1-Benzyl-2-[(2,2-dimethyl-1-oxopropoxy)methyl]piperidin-3-yl (aR)-a-Methoxy-a-(tri-
1
fluoromethyl)benzeneacetate ((R)-MTPA ester; 12). Rf (AcOEt) 0.67. k’ ¼ 1.38 (de ¼ 83%). H-NMR
(400 MHz, CDCl3): 7.52 – 7.47 (m, 2 arom. H); 7.36 – 7.30 (m, 3 arom. H); 7.25 – 7.13 (m, 5 arom. H); 5.28
(M of ABXM, dt, 3J(3,4ax) ¼ 6.9, 3J(3,2) ¼ 3J(3,4eq) ¼ 3.3, HꢀC(3)); 4.34, 3.78 (AB of ABXM, 2J ¼ 11.6,
3J ¼ 6.4, 3J ¼ 5.2, CH2(OMTPA)); 3.72, 3.39 (AB, 2J ¼ 14.1, PhCH2); 3.51 (q, 5J(Me,F) ¼ 1.0, MeO); 2.97
(X of ABXM, 3J(2,3) ¼ 3.3, 3J(2,CH2) ¼ 6.4, 5.2, HꢀC(2)); 2.60 (ddd, 2J ¼ 11.6, 3J(6eq,5eq) ¼ 6.4,
3J(6eq,5ax) ¼ 3.4, HeqꢀC(6)); 2.18 (ddd, 2J ¼ 11.6, 3J(6ax,5ax) ¼ 7.9, 3J(6ax,5eq) ¼ 3.4, HaxꢀC(6)); 1.81 (m,
w1/2 ꢂ 18, HeqꢀC(4)); 1.69 (m, w1/2 ꢂ 22, HaxꢀC(4)); 1.62 – 1.41 (m, CH2(5)); 1.12 (s, Me3C). 13C-NMR
(100 MHz, CDCl3): 178.1 (COCMe3), 165.9 (CO of MTPA); 139.4 (C(1’)); 132.2 (C(1’’)); 129.6 (C(4’’));
128.4 (C(2’), C(6’)); 128.2 (C(2’’), C(6’’)); 128.1 (C(3’), C(5’)); 127.3 (C(3’’), C(5’’)); 126.9 (C(4’)); 123.4
(q, 1J(C,F) ¼ 289, CF3); 84.1 (q, 2J(C,F) ¼ 27.8, PhC(OMe)(CF3)CO); 72.6 (C(3)); 61.5 (C(2)); 61.3
(PhCH2); 58.2 (C(8)); 55.4 (MeO); 48.9 (C(6)); 38.6 (Me3C); 27.5 (C(4)); 27.1 (Me3C); 21.2 (C(5)).
19F{1H}-NMR (376.5 MHz, CDCl3): ꢀ 71.64 (s, CF3).
Dd(1H) ¼ d(12) – d(11) (in Hz): HꢀC(2), ꢀ 4; HꢀC(3), þ 8; HaxꢀC(4), þ 28; HeqꢀC(4), þ 16;
CH2(5), þ 20; HaxꢀC(6), þ 8; HeqꢀC(6), þ 4 ! (2R,3R)-configuration of 11, (2S,3S)-configuration of
12.
4. trans- and cis-5-Benzyl-2-fluoro-1,3-dioxa-5-aza-2-phosphadecalin 2-Oxides (¼2-Fluorohexahy-
dro-5-(phenylmethyl)-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-Oxides) (þ)- and (ꢀ)-13a, (þ)- and
(ꢀ)-13b, (þ)- and (ꢀ)-14a, and (þ)- and (ꢀ)-14b, resp. To a cooled soln. (<08) of (þ)-4 (100 mg,
0.45 mmol) in anh. Et2O (3 ml) and anh. Et3N (155 ml, 1.1 mmol) in a glove box under N2, a cooled soln.
(<08) of POCl2F [24] (49 ml, 76.8 mg, 0.56 mmol, 1.2 equiv.) in anh. Et2O (1 ml) was added with a syringe.
The mixture was stirred for 2 min at 08and then withdrawn and subjected to a fast CC (SiO2 60 (15 –

Helvetica Chimica Acta – Vol. 95 (2012)
731
40 mm, Merck No. 115111), pH 5.6 [7], Et2O). The eluate was gently concentrated (N2 stream, < 308),
and the residue (þ)-13a/(þ)-13b (125 mg; ax/eq ca. 1.5 :1) was subjected to CC (SiO2, pH 5.6 [7], hexane/
Et2O 1 :2): (þ)-13a (62 mg, 49%) and (þ)-13b (27 mg, 22%). Applying the same procedure,
phosphorylation of (ꢀ)-4 (97 mg, 0.44 mmol) afforded (ꢀ)-13a (70 mg, 57%) and (ꢀ)-13b (21 mg,
16%); the axial epimers were less polar. Due to pronounced epimerization of (þ)- and (ꢀ)-13b during
chromatography, the contact time with SiO2 had to be minimized (<10 min). Similarly, the cis-configured
compounds were obtained after phosphorylation and CC (SiO2, pH 5.6 [7], hexane/AcOEt 1:1): from
(þ)-5 (101 mg, 0.46 mmol), (þ)-14a (50 mg, 39%) and (þ)-14b (59 mg, 46%); from (ꢀ)-5 (105 mg,
0.47 mmol); (ꢀ)-14a (48 mg, 36%) and (ꢀ)-14b (54 mg, 40%); the equatorial epimers were less polar.
(þ)-(1R,3R,6S)-7-Benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-Oxide (¼(þ)-(2R,4aS,
8aR)-2-Fluorohexahydro-5-(phenylmethyl)-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-Oxide; (þ)-
13a): Colorless prisms. M.p. 82.5 – 848 ((ꢁ)-13a: m.p. 116.5 – 118.58 [9]). Rf (hexane/Et2O 1:2) 0.26.
[a]2D5 ¼ þ70.7 (c ¼ 0.29, acetone). IR (KBr): identical with that of (ꢁ)-13a [9]. 1H-NMR (400 MHz,
(D6)acetone): 7.36 – 7.22 (m, PhCH2); 4.92 (A of ABX-P, 2J ¼ 10.8, 3J(5eq,P) ¼ 25.3, 3J(5eq,6) ¼ 4.2,
HeqꢀC(5)); 4.44 (ddd, 3J(1,6) ¼ 10.5, 3J(1,10ax) ¼ 9.0, 3J(1,10eq) ¼ 4.8, HꢀC(1)); 4.35 (B of ABX-P, 2J ¼
3
2
3J(5ax,6) ¼ 10.8, J(5ax,P) ꢂ 1, HaxꢀC(5)); 3.96, 3.37 (AB, J ¼ 13.8, PhCH2); 2.86 (m, d-like, w1/2 ꢂ 15,
2J ꢂ 12, HeqꢀC(8)); 2.72 (X of ABX-P, J(6,1) ¼ 10.5, J(6,5ax) ¼ 10.8, J(6,5eq) ¼ 4.2, HꢀC(6)); 2.18 –
2.07 (m, HeqꢀC(10)); 2.15 (td, 2J ¼ 3J(8ax,9ax) ¼ 12.0, 3J(8ax,9eq) ¼ 3.0, HaxꢀC(8)); 1.73 – 1.69 (m,
HeqꢀC(9)); 1.68 – 1.55 (m, HaxꢀC(9), HaxꢀC(10)). 13C-NMR (100 MHz, (D6)acetone): 139.7 (C(1’));
129.4 (C(3’), C(5’)); 129.3 (C(2’), C(6’)); 128.1 (C(4’)); 83.2 (dd, 2J(1,P) ¼ 7.2, 3J(1,F) ¼ 1.8, C(1)); 73.5 (d,
2J(5,P) ¼ 8.0, C(5)); 62.2 (d, 3J(6,P) ¼ 4.8, C(6)); 58.3 (PhCH2); 53.3 (C(8)); 31.9 (d, 3J(10,P) ¼ 9.5,
3
3
3
4
1
C(10)); 23.7 (d, J(9,P) ¼ 2.4, C(9)). 31P-NMR (161.9 MHz, (D6)acetone): ꢀ 15.0 (ddd, J(P,F) ¼ 1004,
3J(P,HeqꢀC(5)) ¼ 25.3, 5J(P,HaxꢀC(5)) ꢂ 1). 19F-NMR (376.5 MHz, (D6)acetone): ꢀ 86.4 (d, 1J(F,P) ¼
1004). EI-MS: 285 (14, Mþ), 194 (9, [M ꢀ PhCH2]þ), 186 (14), 171 (19), 160 (23), 91 (100, PhCHþ2 ),
77 (11, Phþ).
(ꢀ)-(1S,3S,6R)-7-Benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-Oxide (¼(ꢀ)-(2S,4aR,8aS)-
2-Fluorohexahydro-5-(phenylmethyl)-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-Oxide; (ꢀ)-13a):
[a]2D5 ¼ ꢀ72.8 (c ¼ 0.29, acetone). All other data: identical with those of (þ)-13a.
(þ)-(1R,3S,6S)-7-Benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-Oxide (¼(þ)-(2S,4aS,8aR)-
2-Fluorohexahydro-5-(phenylmethyl)-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-Oxide; (þ)-13b):
Colorless plates. M.p. 68.5 – 718 ((ꢁ)-13b: m.p. 86.0 – 90.58 [9]). Rf (hexane/Et2O 1:2) 0.19. [a]D25
¼
þ91.9 (c ¼ 0.27, acetone). IR (KBr): identical with that of (ꢁ)-13b [9]. 1H-NMR (400 MHz,
(D6)acetone): 7.36 – 7.22 (m, PhCH2); 4.97 (A of ABX-P, 2J ¼ 10.2, 3J(5eq,P) ¼ 8.5, 3J(5eq,6) ¼ 5.5,
3
HeqꢀC(5))23); 4.55 (sept.-like, 3J(1,10ax) ꢂ J(1,6) ꢂ 10, 3J(1,10eq) ¼ 4.5, 4J(1,F) ¼ 3.5, 3J(1,P) ꢂ 1,
3
2
4
HꢀC(1)); 4.49 (B of ABX-P, J(5ax,P) ¼ 14.5, J ¼ 3J(5ax,6) ¼ 10.3, J(5ax,F) ¼ 3.2, HaxꢀC(5))23); 3.86,
3.28 (AB, 2J ¼ 13.6, PhCH2); 2.85 (X of ABX, 3J(6,1) ꢂ 10, 3J(6,5ax) ¼ 10.3, 3J(6,5eq) ¼ 5.5, 5J(6,F) ¼ 3.8,
HꢀC(6)); 2.78 (m, d-like, 2J ꢂ 12, HeqꢀC(8)); 2.21 (m, w1/2 ꢂ 20, HeqꢀC(10)); 2.08 (td, 2J ¼ 3J(8ax,9ax) ¼
11.9, 3J(8ax,9eq) ¼ 2.8, HaxꢀC(8)); 1.72 (m, d-like, w1/2 ꢂ 18, 2J ꢂ 11, 5J(9eq,P) ꢂ 2, HeqꢀC(9)); 1.63 – 1.50
(m, HaxꢀC(9), HaxꢀC(10)). 13C-NMR (100.6 MHz, (D6)acetone): 139.4 (C(1’)); 129.6 (C(3’), C(5’));
129.3 (C(2’), C(6’)); 128.1 (C(4’)); 82.1 (dd, 2J(1,P) ¼ 6.3, 3J(1,F) ¼ 1.5, C(1)); 74.1 (d, 2J(5,P) ¼ 78.4,
3
3
C(5)); 61.2 (d, J(6,P) ¼ 10.6, C(6)); 58.4 (PhCH2); 52.8 (C(8)); 32.0 (d, J(10,P) ¼ 7.6, C(10)); 23.5 (d,
4J(9,P) ¼ 2.3, C(9)). 31P-NMR (161.9 MHz, (D6)acetone): ꢀ 14.1 (dbr.ddd, 1J(P,F) ¼ 988,
3J(P,HaxꢀC(5)) ¼ 14.5, 3J(P,HeqꢀC(5)) ¼ 8.5, 3J(P,1) ꢂ 1, 5J(P,9eq) ꢂ 2)23). 19F-NMR (376.5 MHz,
4
5
(D6)acetone): ꢀ 71.2 (dq, 1J(F,P) ¼ 997, 4J(F,HꢀC(1)) ꢂ J(F,HaxꢀC(5)) ꢂ J(F,HꢀC(6)) ꢂ 3). EI-MS:
285 (10, Mþ), 194 (10, [M ꢀ PhCH2]þ), 186 (12), 171 (16), 160 (22), 91 (100, [PhCH2]þ), 77 (4) Phþ).
23
)
The descriptors ꢄaxꢅ and ꢄeqꢅ for the H-atoms of CH2(5) are based on their relative positions in the
chair conformation of the 2,4-dioxa-3-phospha moiety. As discussed (Scheme 5), the conformation is
rather a twist-boat (TB-2) than a chair in the P(3)-equatorially substituted compounds. For reasons
of simplicity, the notation ꢄaxꢅ and ꢄeqꢅ is maintained. HaxꢀC(5) is always cis to HꢀC(1) and
HeqꢀC(5) trans to HꢀC(1), see [18].

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Helvetica Chimica Acta – Vol. 95 (2012)
(ꢀ)-(1S,3R,6R)-7-Benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-Oxide (¼(ꢀ)-(2R,4aR,8aS)-
2-Fluorohexahydro-5-(phenylmethyl)-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-Oxide; (ꢀ)-13b):
[a]2D5 ¼ ꢀ93.8 (c ¼ 0.26, acetone). All other data: identical with those of (þ)-13b.
(þ)-(1S,3S,6S)-7-Benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-Oxide (¼(þ)-(2S,4aS,8aS)-2-
Fluorohexahydro-5-(phenylmethyl)-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-Oxide; (þ)-14a): Col-
orless prisms. M.p. 101.5 – 1048 ((ꢁ)-14a: m.p. 107 – 110.58 [9]). Rf (hexane/AcOEt 1:1) 0.24. [a]D25
¼
þ58.5 (c ¼ 0.38, acetone). IR (KBr): identical with that of (ꢁ)-14a [9]. 1H-NMR (400 MHz,
(D6)acetone): 7.43 – 7.22 (m, PhCH2); 5.05 (A of ABX-P, 2J ¼ 13.0, 3J(5eq,P) ¼ 25.7, 3J(5eq,6) ¼ 1.6,
HeqꢀC(5)); 5.00 (m, w1/2 ꢂ 8, HꢀC(1)); 4.54 (B of ABX-P, 2J ¼ 13.0, 3J(5ax,6) ¼ 3J(5ax,P) ¼ 1.5,
2
2
HaxꢀC(5)); 4.25, 3.27 (AB, J ¼ 13.8, PhCH2); 2.87 (m, quint.-like, w1/2 ꢂ 15, J ¼ 11.6, HeqꢀC(8)); 2.67
(X of ABX-P, 3J(6,1) ¼ 3J(6,5ax) ¼ 3J(6,5eq) ¼ 4J(6,P) ¼ 1.5, HꢀC(6)); 2.15 (td, 2J ¼ 3J(8ax,9ax) ¼ 12.2,
3J(8ax,9eq) ¼ 2.7, HaxꢀC(8)); 2.02 (m, br. d-like, w1/2 ꢂ 18, HeqꢀC(10)); 1.84 (qt, 2J ¼ 3J(9ax,8ax) ¼
3J(9ax,10ax) ¼ 12.2, 3J(9ax,8eq) ¼ 3J(9ax,10eq) ¼ 3.8, HaxꢀC(9)); 1.74 (m, w1/2 ꢂ 30, 4J(10ax,P) ¼ 7.1,
HaxꢀC(10)); 1.47 (m, dq-like, w1/2 ꢂ 18, 2J ¼ 12.2, HeqꢀC(9)). 13C-NMR (100.6 MHz, (D6)acetone): 139.8
2
(C(1’)); 129.7 (C(3’), C(5’)); 129.2 (C(2’), C(6’)); 127.9 (C(4’)); 81.5 (d, J(1,P) ¼ 7.7, C(1)); 70.4 (dd,
2J(5,P) ¼ 7.3, 3J(5,F) < 1, C(5)); 58.9 (d, 3J(6,P) ¼ 5.3, C(6)); 57.5 (PhCH2); 52.5 (C(8)); 30.7 (d,
3J(10,P) ¼ 8.8, C(10)); 20.5 (C(9)). 31P-NMR (161.9 MHz, (D6)acetone): ꢀ 15.3 (dddt, 1J(P,F) ¼ 987,
3J(P,HeqꢀC(5)) ¼ 25.7, 4J(P,HaxꢀC(10)) ¼ 7.1, 4J(P,HaxꢀC(5)) ¼ 4J(P,6) ¼ 1.5). 19F-NMR (376.5 MHz,
1
(D6)acetone): ꢀ 83.6 (d, J(F,P) ¼ 987). EI-MS: 285 (20, Mþ), 194 (15, [M ꢀ PhCH2]þ), 186 (15), 172
(21), 160 (34), 147 (22), 91 (100, PhCHþ2 ), 65 (11).
(ꢀ)-(1R,3R,6R)-7-Benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-Oxide (¼(þ)-(2R,4aR,
8aR)-2-Fluorohexahydro-5-(phenylmethyl)-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-Oxide; (ꢀ)-
14a): [a]2D5 ¼ þ59.8 (c ¼ 0.36, acetone). All other data identical with those of (þ)-14a.
(þ)-(1S,3R,6S)-7-Benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-Oxide (¼(þ)-(2R,4aS,8aS)-
2-Fluorohexahydro-5-(phenylmethyl)-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-Oxide; (þ)-14b):
Colorless oil ((ꢁ)-14b: colorless plates, m.p. 93 – 94.58 [9]). Rf (hexane/Et2O 1:2) 0.26. [a]2D5 ¼ þ23.3
(c ¼ 0.28 acetone). IR (KBr): identical with that of (ꢁ)-14b [9]. 1H-NMR (400 MHz, (D6)acetone):
3
3
7.39 – 7.23 (m, PhCH2); 5.00 (A of ABX-P, 2J ¼ 11.8, J(5eq,P) ¼ 10.9, J(5eq,6) ¼ 7.0, HeqꢀC(5)); 4.95
3
3
4
2
3
(ddd, J(1,P) ¼ 12.8, J(1,6) ¼ 3.4, J(1,F) ¼ 2.7, HꢀC(1)); 4.62 (B of ABX-P, J ¼ 11.8, J(5ax,P) ¼ 14.0,
3J(5ax,6) ¼ 3.4, 4J(5ax,F) ¼ 2.3, HaxꢀC(5)); 4.04, 3.59 (AB, 2J ¼ 13.7, PhCH2); 3.21 (X of ABX-P,
3J(6,5eq) ¼ 6.8, 3J(6,5ax) ¼ 3J(6,1) ¼ 3.5, 4J(6,P) ¼ 1.8, HꢀC(6)); 2.72 (ddd, 2J ¼ 12.0, 3J(8eq,9ax) ¼ 7.4,
3J(8ax,9eq) ¼ 3.6, HaxꢀC(8)); 2.31 (ddd-like, 2J ¼ 12.0, 3J(8ax,9eq) ¼ 7.7, 3J(8eq,9ax) ¼ 3.3, HaxꢀC(8));
2.06 (m, w1/2 ꢂ 20, HeqꢀC(10)); 1.99 – 1.90 (m, HaxꢀC(10)); 1.62 – 1.52 (m, CH2(9)). 13C-NMR
(100.6 MHz, (D6)acetone): 139.8 (C(1’)); 129.4 (C(3’), C(5’)); 129.3 (C(2’), C(6’)); 128.0 (C(4’)); 81.7
2
3
2
3
(dd, J(1,P) ¼ 8.8, J(1,F) ¼ 1.2, C(1)); 67.7 (d, J(5,P) ¼ 6.8, C(5)); 58.5 (PhCH2); 57.0 (d, J(6,P) ¼ 9.0,
C(6)); 49.0 (C(8)); 28.8 (d, J(10,P) ¼ 2.7, C(10)); 22.3 (C(9)). 31P-NMR (161.9 MHz, (D6)acetone): ꢀ
3
15.6 (dddddd, 1J(P,F) ¼ 990, 3J(P,HaxꢀC(5)) ¼ 14.0, 3J(P,HꢀC(1)) ¼ 12.8, 3J(P,HeqꢀC(5)) ¼ 10.9,
4J(P,HaxꢀC(10)) ¼ 3.4, 4J(P,6) ¼ 1.8)23). 19F-NMR (376.5 MHz, (D6)acetone): ꢀ 74.5 (m, dq-like,
1J(F,P) ¼ 990, J(F,HꢀC(1)) ꢂ J(F,HaxꢀC(5)) ꢂ J(F,HeqꢀC(5)) ꢂ J(F,HeqꢀC(10)) ꢂ 2). EI-MS: 285 (25,
4
4
4
5
Mþ), 194 (13, [M ꢀ PhCH2]þ), 186 (15), 172 (22), 160 (41), 91 (100, PhCHþ2 ), 65 (11).
(ꢀ)-(1R,3S,6R)-7-Benzyl-3-fluoro-2,4-dioxa-7-aza-3-phosphadecalin 3-Oxide (¼(ꢀ)-(2S,4aR,8aR)-
2-Fluorohexahydro-5-(phenylmethyl)-4H-1,3,2-dioxaphosphorino[5,4-b]pyridine 2-Oxide; (ꢀ)-14b):
[a]2D5 ¼ ꢀ22.4 (c ¼ 0.27, acetone). All other data: identical with those of (þ)-14b.
5. X-Ray Crystal-Structure Determinations of (ꢀ)-13a and (þ)-14a16). 5.1. General. All measure-
ments were made with a Nonius-KappaCCD area-detector diffractometer [25], graphite-monochro-
mated MoKa radiation (l 0.71073 ꢆ), and an Oxford-Cryosystems-Cryostream-700 cooler. Data
reduction was performed with HKL DENZO and SCALEPACK [26]. The intensities were corrected
for Lorentz and polarization effects, and an absorption correction based on the multi-scan method [27]
was applied. The space groups were uniquely determined by the systematic absences. Equivalent
reflections, other than Friedel pairs, were merged. Neutral-atom scattering factors for non-H-atoms were
taken from [28] and the scattering factors for H-atoms were taken from [29]. Anomalous dispersion
effects were included in Fc [30]; the values for f’ and f’’ were those of [31]. The values of the mass

Helvetica Chimica Acta – Vol. 95 (2012)
733
attenuation coefficients were those of [32]. All calculations were performed with the SHELXL97
program [33], and the crystallographic diagrams were drawn with ORTEPII [34].
5.2. Determination of (ꢀ)-13a and (þ)-14a. The unit-cell constants and an orientation matrix for data
collection were obtained from a least-squares refinement of the setting angles of 66907 ((ꢀ)-13a) and
14905 ((þ)-14a) reflections in the range 48 < 2q < 568 and 48 < 2q < 608, resp. The mosaicity was 0.602(1)8
((ꢀ)-13a) and 0.432(1)8 ((þ)-14a). A total of 295 ((ꢀ)-13a) and 240 ((þ)-14a) frames were collected by
using w scans with k offsets, 22 and 38 s exposure time and a rotation angle of 1.58 and 2.08 per frame, and
a crystal-detector distance of 32.0 and 30.0 mm, resp. The data collection and refinement parameters are
given in Table 2. Aview of the molecules is shown in Fig. 2 ((ꢀ)-13a) and 3 ((þ)-14a). The structure were
solved by direct methods with SIR92 [35], which revealed the positions of all non-H-atoms. There were
three symmetry-independent molecules of the same enantiomer in the asymmetric unit of (ꢀ)-13a; the
atomic coordinates of the two molecules were tested carefully for a relationship from a higher symmetry
space group with the program PLATON [36], but none could be found; the molecules had very similar
conformations with only small differences in the orientation of the phenyl ring. The non-H-atoms were
refined anisotropically. All of the H-atoms were placed in geometrically calculated positions and refined
by using a riding model where each H-atom was assigned a fixed isotropic displacement parameter with a
value equal to 1.2 Ueq of its parent atom. The refinement of the structures was carried out on F2 by using
full-matrix least-squares procedures, which minimized the function Sw(F2o ꢀ Fc2 )2. The weighting scheme
was based on counting statistics and included a factor to downweight the intense reflections. Plots of
Sw(F2o ꢀ F2c )2 vs. Fc/Fc(max) and resolution showed no unusual trends. A correction for secondary
extinction was applied. Refinement of the absolute structure parameter [37] yielded a value of ꢀ 0.05(8)
((ꢀ)-13a) and 0.02(8) ((þ)-14a), which confidently confirmed that the refined models corresponded with
the true enantiomorphs.
6. Enzyme Kinetics. 6.1. General. For the detailed experimental procedure and the methods of data
analysis, see the preceding reports [1][2]. The following parameters were as in [1]: Apparatus and general
experimental conditions, phosphate buffer pH 7.00, AChE soln., ATC and DTNB solns., inhibitor solns.,
and the determination of Km. P(O)F(OiPr)2 (diisopropyl phosphorofluoridate) was used as the standard
reference.
6.2. Ellman-Assay [20]. In a polystyrene cell (4 ml, d ¼ 1 cm), phosphate buffer pH 7.00 (2 ml),
DTNB soln. (100 ml), and ATC soln. (20 ml) were mixed and thermostatted at 258 (ca. 5 min). Then,
inhibitor soln. (x ml, known [I], x ꢃ 25 ml) and MeCN ((25 – x) ml) were added. At t ¼ 0, the AChE soln.
(1 ml) was added and the mixture gently mixed for 10 s. After 20 s, the monitoring of the absorption at
412 ꢁ 2 nm (liberated bis-anion of 5-mercapto-2-nitrobenzoic acid) automatically started, and 600 data
points were collected for 10 min at various concentrations of the inhibitor. As in the Km determinations,
the total volume was 3.145 ml, the concentration of the substrate [S] ¼ 502 mm. Per inhibitor, at least five
measurements with different inhibitor concentrations were performed, the smallest one being ca. 1/5 –
1/10 of the largest one.
6.3. Data Analysis [21]. The integrated rate equation describing product generation (monitored by
the absorbance A at 412 ꢁ 2 nm) and the apparent rate constants (kobs) is given by Eqn. 1. It was fitted
(R > 0.999) to progress curves recorded at fixed [S] and variable [I] (primary plot, A ¼ f(t)) to obtain a
series of kobs values and their standard errors (SE). The inhibition parameters were obtained from the
secondary plots (kobs ¼ f([I])) that resulted from weighted (SEꢀ2) linear or nonlinear regression
according to Eqns. 2 or 3. The analysis of these plots enabled a differentiation between the inhibition
mechanisms: kobs depended linearly upon the inhibitor concentration for mechanism a and hyperbolically
for mechanism b (see Scheme 6). For mechanism a, the ka values were calculated according to Eqn. 2, its
slope kobs/[I] was obtained from the linear regression. The decisive plot for mechanism b was doubly
reciprocal (1/kobs ¼ f(1/[I]), Eqn. 3), and the KD- and kp-values were calculated by linear regression, the
slope being (KD/kp)(1 þ [S]/Km) and the intercept 1/kp. The overall inhibitory potency (ki) is expressed
by kp/KD (see Scheme 6).
À
Á
vz
kobs
obsꢄt
1 ꢀ eꢀk
A ¼
(1)

734
Helvetica Chimica Acta – Vol. 95 (2012)
Table 2. Crystallographic Data of ( ꢀ )-13a and (þ)-14a
(ꢀ)-13a
(þ)-14a
Crystallized from
Empirical formula
Mr
Et2O
C13H17FNO3P
285.25
pentane/Et2O
C13H17FNO3P
285.25
Crystal color, habit
Crystal dimensions [mm]
Temperature [K]
colorless, prism
0.25 ꢆ 0.35 ꢆ 0.35
160(1)
colorless, prism
0.15 ꢆ 0.22 ꢆ 0.25
160(1)
Crystal system
Space group
Z
orthorhombic
P212121 (#19)
12
orthorhombic
P212121 (#19)
4
Reflections for cell determination
66907
14905
2q range for cell determination [8]
4 – 56
4 – 60
Unit cell parameters: a [ꢆ]
13.5669(2)
15.5403(2)
19.7914(3)
4172.7(1)
1800
9.7745(2)
10.0774(2)
13.5432(2)
1334.03(4)
600
1.420
0.221
f and w
60
b [ꢆ]
c [ꢆ]
V [ꢆ3]
F(000)
Dx [g cmꢀ3
]
1.362
0.212
m(MoKa) [mmꢀ1
Scan type
]
w
2q(max) [8]
56
Transmission factors (min; max)
Total reflections measured
Symmetry independent reflections
Rint
0.812; 0.951
62660
9854
0.854; 0.970
24752
3878
0.082
0.063
Reflections with I > 2s(I)
Reflections used in refinement
Parameters refined
8055
9854
515
3449
3878
173
Final R(F) (I > 2s(I) reflections)
0.0541
0.1346
0.0383
0.0958
wR(F2) (all data)
Weights
w ¼ [s2(Fo2) þ (0.073P)2
w ¼ [s2(Fo2) þ (0.0543P)2
þ 0.4328P]ꢀ1
,
þ 0.1646P]ꢀ1
,
where P ¼ (Fo2 þ 2Fc2)/3
where P ¼ (Fo2 þ 2Fc2)/3
Goodness of fit
1.079
1.068
Secondary extinction coefficient
Final Dmax/s
0.0107(9)
0.001
0.020(2)
0.001
D1 (max; min) [e ꢆꢀ3
s(d(CꢀC)) [ꢆ]
]
0.59; ꢀ 0.56
0.34; ꢀ 0.30
0.003 – 0.005
0.002 – 0.003
ꢀ
ꢁꢀ
ꢁ
kobs
½Iꢅ
½Sꢅ
ka ¼
1 þ
(2)
Km
ꢀ
ꢁꢀ
ꢁꢀ
ꢁ
1
kobs
KD
kp
½Sꢅ
1
½Iꢅ
1
kp
¼
1 þ
þ
(3)
Km
6.4. Results (Table 1). The secondary plot (kobs ¼ f([I])) exhibited a linear dependence for (ꢁ)-13a,
(þ)- and (ꢀ)-13a, (þ)- and (ꢀ)-14a, and P(O)F(OiPr)2. Hence, mechanism a was assigned. In the case of
(þ)- and (ꢀ)-13b, and (þ)- and (ꢀ)-14b, the secondary plot depended hyperbolically upon [I], and
mechanism b was assigned to these compounds.

Helvetica Chimica Acta – Vol. 95 (2012)
735
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Received December 15, 2011