Synthesis of glyceryl glycosides related to A-type prymnesin toxins

Article abstract of DOI:10.1016/j.carres.2018.04.008

A suite of glycosylated glycerol derivatives representing various fragments of the glycosylated ichthyotoxins called prymnesins were chemically synthesised. Glycerol was used to represent a small fragment of the prymnesin backbone, and was glycosylated at

Full text of DOI:10.1016/j.carres.2018.04.008

Accepted Manuscript
Synthesis of glyceryl glycosides related to A-type prymnesin toxins
Edward S. Hems, Sergey A. Nepogodiev, Martin Rejzek, Robert A. Field
PII:
S0008-6215(18)30153-8
10.1016/j.carres.2018.04.008
CAR 7553
DOI:
Reference:
To appear in: Carbohydrate Research
Received Date: 8 March 2018
Revised Date: 13 April 2018
Accepted Date: 14 April 2018
Please cite this article as: E.S. Hems, S.A. Nepogodiev, M. Rejzek, R.A. Field, Synthesis of glyceryl
glycosides related to A-type prymnesin toxins, Carbohydrate Research (2018), doi: 10.1016/
j.carres.2018.04.008.
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ACCEPTED MANUSCRIPT
Synthesis of glyceryl glycosides related to A-type prymnesin toxins
Edward S. Hems, Sergey A. Nepogodiev, Martin Rejzek, Robert A. Field*
Department of Biological Chemistry, John Innes Centre, Norwich Research Park, Norwich, NR4 7UH,
UK
Abstract
A suite of glycosylated glycerol derivatives representing various fragments of the glycosylated
ichthyotoxins called prymnesins were chemically synthesised. Glycerol was used to represent a
small fragment of the prymnesin backbone, and was glycosylated at the 2° position with the sugars
currently reported to be present on prymnesin toxins. Neighbouring group participation was utilised
to synthesise 1,2-trans-glycosides. SnCl2-promoted glycosylation with furanosyl fluorides gave 1,2-
cis furanosides with moderate stereocontrol, whilst TMSOTf promoted glycosylation with a furanosyl
imidate gave a 1,2-cis furanoside with good stereocontrol. The chemical synthesis of two larger
glyceryl diglycoside fragments of prymnesin-1, glycosylated with α-ʟ-arabinopyranose and α-ᴅ-
ribofuranose, is also described. As the stereochemistry of the prymnesin backbones at this region is
undefined, both the 2R- and 2S- glycerol isomers were synthesised. The separated diastereoisomers
were distinguished by comparing NOESY NMR with computational models.
Key words: Algae, Prymnesium parvum, toxins, prymnesins, glyceryl glycosides
1. Introduction
Prymnesins-1 and -2 are potent ichthyotoxins produced by Prymnesium parvum, a flagellated alga
which is widely distributed across brackish waters.[1] As a result, P. parvum has been attributed to
large scale fish kills globally, resulting in detrimental ecological and economic impacts to affected
areas.[2] There are currently 16 different prymnesins reported in literature.[3,4] According to the
structure of their ladder-frame polyether backbones primnesins can be divided into three main
types. The backbone of A-type prymnesins is composed of 91 carbon atoms, B-type of 85 and C-type
of 83 carbon atoms. The high diversity of these natural products arises from different patterns of
glycosylation combined with the various extent of backbone chlorination. Chemical structures of
three prymnesins, prymnesin-1 and prymnesin-2 derived from the A-type backbone (Figure 1) and
Prymnesin-B1, a B-type backbone derivative, have been elucidated, although the absolute
configuration of several stereocenters in these molecules remains unknown.[3–6]
* Corresponding author: rob.field@jic.ac.uk
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Figure 1 The reported structures of the A-type prymnesin ichthyotoxins.[3–6]
In order to gain insight into the structure and properties of prymnesin toxins, we aimed to
synthesise a library of glyceryl glycosides which can serve as model fragments of specific toxins
relating to prymnesin-1 and prymnesin-2 (Figure 2). A few glyceryl glycosides have been reported in
natural products literature and prepared by chemical synthesis. Thus, 1,3-dihydroxypropan-2-yl α-ᴅ-
glucopyranoside is an osmolyte used by cyanobacteria to combat the effects of salt-stress and
drought.[7] Glyceryl glycosides have also found use as moisturising ingredients in cosmetics.[8]
‘Floridoside’ (1,3-dihydroxypropan-2-yl α-ᴅ-galactopyranoside) has been isolated from the red alga
Mastocarpus stellatus[9] and found to be a potent activator of the classical complement pathway.[9]
Although relevant to B-type prymnesins, its’ chemical synthesis and characterisation has been
reported in the literature,[10,11] and so will not be repeated in this study. The synthesis of α-ʟ-
arabinopyranoside (2), which is also relevant to our study, was published in 1972 but with little
characterisation data.
Figure
2
Target glyceryl glycosides as fragments of prymnesin-1 (β-ᴅ-galactofuranoside 1, α-ʟ-
arabinofuranoside 2 and α-ᴅ-ribofuranoside 3)[3] and prymnesin-2 (either α-ʟ-xylofuranoside 4 or β-ᴅ-
arabinofuranoside 5).[3,4]
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One of the problems associated with detection of the prymnesin toxins is that current methods rely
on detection of the parent organism.[12,13] None of these methods can provide quantification of
toxin levels in water samples. Furthermore, the link between amount of P. parvum in water and
prymnesin concentration is not straight forward.[1,13,14] Thus, it has been noticed that in some
cases blooms of P. parvum do not necessarily lead to fish kills.[15] Work in our group revealed a
new mechanism of P. parvum toxicity, which results from mass cell lysis caused by the infection of
alga with a newly discovered double stranded DNA megavirus, PpDNAV.[16] Therefore sensitive and
quantitative techniques for direct detection for P. parvum toxins are highly desirable. As an example,
antibody recognition has been developed for polyketide algal toxins such as okadaic acid and
brevetoxins.[17,18] We decided to focus on a glycosylated polyol fragment of prymnesin-1 and
synthesise diglycosides 6 and 7 (Figure 3) incorporating α-ᴅ-ribofuranosyl and α-ʟ-arabinopyranosyl
residues as possible candidates for antigens for anti-carbohydrate antibodies production. These
fragments represent the region between C76-C78 on the prymnesin-1 backbone (Figure 1). The
design of 6 and 7 consists of glycerol as a polyol backbone mimic which is glycosylated at positions 1
and 2 and alkylated at position 3 with 3-aminopropyl linker. The linker is required for conjugation of
6 and 7 via cross-linking techniques to a protein carrier, with the aim to make carbohydrate antigens
for antibody production.[19,20] As the exact stereochemistry of the glycosylated backbone region of
prymnesin-1 is not yet known,[3] both the (2R)- and (2S)- glycerol diastereoisomers 6 and 7 were
required.
Figure 3 Target diastereomeric diglycosides 6 and 7.
2. Results and Discussion
2.1. Synthesis of 1,2-trans-linked glyceryl glycoside fragments of
prymnesin-1
Tin (IV) chloride-promoted glycosylation of 1,3-di-O-benzyl glycerol (9) with per-O-benzoyl-β-ᴅ-
galactofuranose (8)[21,22] produced exclusively 1,2-trans-glycoside 10 (Scheme 1).
After
hydrogenation to give 11, the benzoyl protecting groups were removed using a 5:2:1 mixture of
MeOH-H2O-Et3N leading to β-galactofuranoside 1. The configuration of the newly formed glycosidic
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linkage was confirmed by the appearance of H-1’ signal either as a singlet (δ 5.61 for 10) or a doublet
(δ 4.99 with J1,2 1.7 Hz for 1) in the 1H NMR spectra. These J1,2 values are known to be characteristic
for a β-galactofuranosidic linkage.[22]
Scheme 1 Synthesis of 1,3-dihydroxypropan-2-yl β-ᴅ-galactofuranoside (1 Reagents and conditions: (a) SnCl4,
CH2Cl2, 39%; (b) H2, 10% Pd/C, EtOAc, Et3N, 51%. (c) MeOH-H2O-Et3N (5:2:1), 90%.
The synthesis of 1,3-dihydroxypropan-2-yl α-ʟ-arabinopyranoside (2) has previously been
described,[23] but the product was only characterised by optical rotation. Silver carbonate-
promoted glycosylation of alcohol 9 with bromide 13 prepared from per-O-benzoyl-β-ʟ-
arabinopyranose (12)[24,25] gave arabinopyranoside 14 in 50 % yield (Scheme 2). The presence of
2
-O-benzoyl group in donor 13 ensured the formation of only the 1,2-trans α-anomer 14, and this
was confirmed by NMR spectra which showed signals for the anomeric proton (doublet at 5.06 ppm
with J1’,2’ 5.9 Hz) and anomeric carbon (100.4 ppm) consistent with α-configuration as followed from
comparison with spectra of methyl 2,3,4-tri-O-benzoyl-α- and β-arabinopyranosides.[26] The
glycerol benzyl ether protecting groups were removed by hydrogenation over a Pd/C catalyst,
followed by NaOMe catalysed removal of the benzoates to give 1,3-dihydroxypropan-2-yl α-ʟ-
1
arabinopyranoside (2) as a white powder. H NMR spectroscopy analysis of 2 showed a H-1’ signal at
4.35 ppm as a doublet with a larger J1’,2’ coupling value of 7.5 Hz, which confirmed the presence of
the α-anomer.
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Scheme 2 – Synthesis of α-ʟ-arabinopyranoside 2. Reagents and conditions: (a) HBr/AcOH, CH2Cl2,
95%; (b) 1,3-di-O-benzyl glycerol, Ag2CO3, Drierite™, toluene, 50%; (c) H2, 10% Pd/C, EtOAc, Et3N,
22%; (d) NaOMe, MeOH, 76%.
2.2. Synthesis of 1,2-cis linked glyceryl furanoside fragments
Synthesis of 1,2-cis furanosides can prove challenging as it can be difficult to control the
stereochemistry at the anomeric centre.[27,28] Unless indirect glycosylation approaches are
used,[29–31] application of non-participating protecting groups remains the main method of
minimising the formation of 1,2-trans glycofuranosylation products.
For syntheses of 1,2-cis furanosides 3-5, we resorted to Mukaiyama’s general methodology
employing SnCl2-promoted glycosylation with benzylated glycofuranosyl fluorides.[32,33] 2,3,5-Tri-
O-benzylated glycofuranosyl fluorides 16-18 were synthesised in high yield by fluorination of the
corresponding hemiacetals with DAST (for more details see Suppl. Information).[34,35] In reactions
with 1,3-di-O-benzyl-glycerol β-D-ribofuranosyl fluoride 16 the purified β-anomer was used. As this
resulted in a mixture of α- and β-linked products (19a & 19b), we decided that there was no benefit
in removing the 1,2-cis fluoride. Therefore, in the case of ʟ-xylofuranosyl and ʟ-arabinofuranosyl
glycosides 17 and 18, respectively, α/β-mixtures mixtures of the fluoride anomers were employed
(Scheme 3).
OBn
BnO
HO
BnO
BnO
F
O
a
O
b
d
O
O
O
OBn
OH
OH
OBn
OBn
19a
20a
+
O
O
BnO OBn
HO OH
BnO OBn
BnO OBn
16
19a
19b
3
OBn
OBn
OH
OH
O
BnO
O
OBn
O
O
HO
O
OBn
O
c
F
OBn
OBn
+
BnO
BnO
BnO
BnO
O
HO
HO
BnO
BnO
BnO
BnO
OH
OBn
4
17
20a
20b
OH
OH
OBn
OBn
O
HO
O
OBn
O
O
BnO
O
OBn
O
c
e
F
21a
+
OBn
OBn
O
OH
BnO
BnO
BnO
18
21a
21b
5
Scheme 3 – Synthesis of prymnesin-1 fragments 3-5. Reagents and conditions: (a) 1,3-di-O-benzyl glycerol, 1
equiv. SnCl2, 1 equiv. TrClO4, MS 4Å, Et2O; (b) H2, 10% Pd/C, MeOH-EtOH (5:1); (c) 1,3-di-O-benzyl glycerol, 1
equiv. SnCl2, 4Å MS,Et2O; (d) H2, 10% Pd/C, MeOH-PrOH (9:1); (e) H2 10% Pd/C, MeOH-EtOAc (9:1).
All glycosylations, which were promoted by one equivalent of SnCl2 and carried out in the presence
of 4Å MS in diethyl ether, resulted in the the formation of anomeric mixtures of furanosides 19-21 in
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which the 1,2-cis-glycosides dominated. It has been previously noted that in Mukaiyama’s synthesis
of ribofuranosides both yields and stereoselectivity could be improved with the addition of
triphenylmethyl perchlorate (TrClO4).[32] In our hands, addition of TrClO4 to the reaction involving
donor 16 had only a negligible effect on the reaction outcome, so this variation was not applied in
glycosylation with 17 and 18.
To establish the anomeric configuration of each component of mixtures of glycosylation products
19-21, individual glycosides were isolated and characterised by NMR spectroscopy. It is well known
that α- and-β-glycofuranosides can be distinguished on the basis of the differences in values of their
1
3JH-1,H-2 coupling constants in H NMR spectra, which are in the range of 0-2 Hz for of 1,2-trans-
furanosides and ~4 Hz larger for 1,2-cis-furanosides.[36–38] In addition, resonances of anomeric
carbons of 1,2-cis-furanosides in 13C NMR spectra appear 4-6 ppm upfield compared to
corresponding signals of 1,2-trans-furanosides.[37] Thus, the stereochemistry of the glycosidic
1
linkages in pairs of anomeric glycosides 19-21 was determined using these characteristic H and 13C
NMR signals, as summarised in Table 1. Finally, per-O-benzylated 1,2-cis-furanosides structurally
related to A-type prymnesin were deprotected by catalytic hydrogenolysis over Pd/C (Scheme 3).
NMR analyses of resulting α-ᴅ-ribofuranoside 3, α-L-xylofuranoside 4 and β-ᴅ-arabinofuranoside 5
confirmed their anomeric configuration (Table 1)
Table 1 Characteristic data for anomeric signals in 1H and 13C NMR spectra of pentofuranosides 3-4
and 23-25.
Compound
Configuration
δH-1 (ppm)
J1,2 (Hz)
4.3
4.4
4.7
4.3
0
δC-1 (ppm)
102.0
101.3
98.0
3
1,2-cis (α-
D
-ribo)
-xylo)
-arabino)
-ribo)
1,2-trans (β- -ribo)
1,2-cis (α- -xylo)
1,2-trans (β- -xylo)
1,2-cis (β- -arabino)
1,2-trans (α- -arabino)
5.11
4
1,2-cis (α-
L
5.09
5
1,2-cis (β-
D
5.05
19a
19b
20a
20b
21a
21b
27
1,2-cis (α-
D
5.38
101.5
104.9
99.9
D
5.30
L
5.35
4.3
1.9
4.4
<1
L
5.31
107.1
100.8
106.1
101.5
101.4
D
5.32
D
4.08
1,2-cis (α-
D
-ribo)
-ribo)
5.21
4.2
4.4
28
1,2-cis (α-
D
5.23
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2.3. Synthesis of diastereomeric diglycoside 6 and 7
We initially tried to synthesise the separate enantiomers of 23 by DMTST promoted glycosylation of
optically pure glycidol with ethyl 2,3,4-tri-O-benzoyl-1-thio-β-L-arabinopyranoside.[39]
Unfortunately the glycosylation was very low yielding and gave a complex mixture of products which
proved challenging to separate. As such we changed our approach and for the synthesis of 1,2-O-
glycosylated glycerol derivatives 6 and 7 we used allyl alcohol as a precursor. It was first
arabinosylated, then oxidised to a diastereomeric mixture of epoxides which were then
regioselectively ring-opened with 3-azidopropanol in the presence of Sc(OTf)3 (Scheme 4). This
generated an acceptor for a final ribosylation step and introducing a functionalised linker at the
same time (Scheme 4). Attempts to glycosylate allyl alcohol with per-O-benzoyl β-ʟ-arabinopyranose
(12) in the presence of BF3.OEt2 gave a 3:1 mixture of α and β anomers, which proved very
challenging to separate, in a combined 76% yield. To ensure that only the desired α-anomer was
synthesised, Koenigs-Knorr glycosylation conditions with perbenzoylated ʟ-arabinopyranosyl
bromide 13 were applied, leading to the target 1,2-trans-ʟ-arabinopyranoside 22 in 62% yield.
Scheme 4. Synthesis of diastereomeric diglycosides 6 and 7. Reagents and conditions: (a) AllOH, BF3-OEt2,
CH2Cl2, α/β 3:1; (b) HBr/AcOH, CH2Cl2, 85%; (c) AllOH, Ag2CO3, DCE, 4Å MS, 62%; (d) mCPBA, DCE, reflux, 56%;
(e) 3-azidopropanol, Sc(OTf)3, toluene, 40%; (f) TMSOTf, CH2Cl2, 4Å MS, -78 °C, 15% (27), 10% (28); (g) i) H2,
10% Pd/C, EtOAc-MeOH 1:1. ii) MeOH/H2O/Et3N (5:2:1), 59% (6), 41% (7).
7

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1
The 1,2-trans configuration of 22 was confirmed by H NMR spectrum which showed the H-1 signal
at 4.82 ppm as a doublet with a J1’,2’ coupling value of 6.1 Hz. Epoxidation of the alkene 22 was
achieved using mCPBA[8] giving compound 23 as an inseparable 1:1 mixture of (R,S)-diastereomers,
as judged by integration of the anomeric proton signals at 4.92 and 4.82 ppm. Using a combination
1
of HSQCed and COSY 2D NMR spectra, it was possible to fully assign the H and 13C NMR spectra of
the individual isomers. Since both (R)- and (S)-isomers of the prymnesin-1 fragments were targeted,
the synthesis continued without diastereomer separation at this stage.
The epoxide 23 was ring-opened with 3-azidopropanol in the presence of Sc(OTf)3, leading to
installation of a linker with azido functional group, suitable for later reduction and conjugation of the
glycoside fragment to a carrier protein for antibody generation. The resulting alcohol 24 remained
an 1:1 mixture of (R,S)-diastereomers which were inseparable, but again it was possible to fully
1
assign their H and 13C NMR spectra using a combination of HSQCed and COSY 2D NMR. To effect
1,2-cis-ribofuranosylation of 24, we employed imidate 25 which has been used several times to good
effect for stereoselective ribosylations using TMSOTf as a promotor and either DCM or DCE as a
solvent.[40–42] The initial attempt to couple 24 to 25 was carried out in DCE at -30 °C and resulted
in predominant formation of the β-riboside (α/β 1.0:5.8, as judged by 1H NMR). The
stereoselectivity improved when the glycosylation was conducted in CH2Cl2 at -78 °C. Since acceptor
24 was a 1:1 (R,S)-mixture, the glycosylation resulted in four products. 1H NMR of the mixture of
ribosylation products showed a slight excess of the desired α-ribosides, with an α/β ratio of 1.4:1.
The target α-ribosides 27 and 28 were separated from the (R,S)-mixture of β-ribosides 26 using
preparative TLC. The stereochemistry of the newly formed 1,2-cis-ribofuranosyl linkages in 27 and
1
28 was confirmed by H and 13C NMR, which showed typical values for 1,2-cis-furanosides (Table 1).
To assign (R,S)-configurations in 27 and 28, we used a combination of NMR spectroscopy and
modelling approaches. The lowest energy conformations for both isomers were calculated using
MarvinSketch 15.1.19.0 Conformer Calculator Plugin (Drieding Force Field), and revealed a
noticeable difference in the orientation of the benzyl protecting groups with respect to the azido-
propyl linker (Figure 4). In the computational model for the (S)-isomer 28, the phenyl ring of the 2-
OBn group of the ribosyl residue is in close proximity to the azidopropyl group. In contrast, the
model of (R)- isomer 27 showed the aromatic benzyl protecting groups clustered together and
oriented away from the azidopropyl group. The conclusions arising from the computational models
were reinforced by 2D NOESY NMR spectra of compound 28, which show a clear cross-peak between
signals for the azidopropyl and the benzyl protecting groups. In contrast, no such cross-peaks can be
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identified in NOESY spectra of diastereoisomer 27. These observations allowed us to conclude that
the chiral centres of the aglycones in compounds 27 and 28 have (R)- and (S)-configurations,
respectively.
Finally, global deprotection of compounds 27 and 28 was achieved by first Pd/C-catalysed
hydrogenolysis of benzyl groups with simultaneous reduction of the azide to amine, followed by de-
O-benzoylation using a mixture of MeOH-H2O-Et3N (5:2:1). In this way, diastereomeric 2,3-di-O-
glycosylated derivatives of glycerol, 6 and 7, equipped with an aminopropyl linker were synthesised
in 59% and 41% yields, respectively.
2D NOESY of compound 27 shows no
Lowest energy conformation model for (2R)-isomer 27
long range cross-peak.
2D NOESY of compound 28 shows long
Lowest energy conformation model for (2S)-isomer 28
range cross-peak.
Figure 4 - 3D Model representations of lowest energy calculations (MarvinSketch 15.1.19.0 Conformer
Calculator Plugins) for diastereomers 27 and 28 and expanded 2D NOESY NMR spectra.
9

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3. Conclusions
A library of glyceryl glycoside fragments inspired by the structures of prymnesin toxins was
prepared. Central to these syntheses were efforts to control glycosylation stereochemistry. The 1,2-
trans-linked fragment 1,3-dihydroxypropan-2-yl β-ᴅ-galactofuranoside (1) was synthesised using
SnCl4 promoted glycosylation with excellent stereocontrol, while 1,3-dihydroxypropan-2-yl α-ʟ-
arabinopyranoside (2) was synthesised using Koenigs-Knorr methodology, again with excellent
stereocontrol at the anomeric position.[43] 1,2-cis Glycofuranosyl-glycerol fragments 3, 4 and 5
were successfully synthesised from the corresponding glycofuranosyl fluoride donors, but with only
moderate stereoselectivity.
Elaborating with respect to the stripped-down glycosyl glycerol fragments, two glyceryl diglycoside
fragments inspired by prymnesin-1, 6 and 7, were also chemically synthesised. As the
stereochemistry of the prymnesin backbone in this region has not been established, both the 2R-
and 2S- diastereoisomers with respect to the glycerol backbone were synthesised. This was
achieved by glycosylating 3-(3-azidopropoxy)-2-R/S-hydroxypropyl 2,3,4-tri-O-benzoyl-α-ʟ-
arabinopyranoside
(24)
with
2’,3’,5’-tri-O-benzyl-β-ᴅ-ribofuranosyl
(N-phenyl)-2,2,2-
trifluoroacetimidate (25), which gave a mixture of α/β-ribosides in a ratio of 1.4:1. The α-ribosides
were separable both from the β-ribosides (26) and also from each other, to yield the diastereomeric
(2R)- and (2S)-3-(3-azidopropoxy)-2-[(2’,3’,5’-tri-O-benzyl-α-ᴅ-ribofuranosyl)oxy]propyl-2’’,3’’,4’’-tri-
O-benzoyl-α-ʟ-arabinopyranoside (27 and 28 respectively). The assignment of the stereochemistry
at the 2° position of the glycerol backbone was achieved by comparing nOe NMR spectra with
computational models of the lowest energy conformations of both isomers. Global deprotection of
27 and 28 yielded the desired (2R)-3-(3-aminopropoxy)-2-(α-ᴅ-ribofuranosyloxy)propyl α-ʟ-
arabinopyranoside
(6)
and
(2S)-3-(3-aminopropoxy)-2-(α-ᴅ-ribofuranosyloxy)propyl
α-ʟ-
arabinopyranoside (7), respectively, which are suitably functionalised for prospective bioconjugation
studies.
4. Experimental
4.1 General
Reagents and anhydrous solvents were supplied by Sigma Aldrich, and were used without further
purification. Analytical grade solvents were supplied by Fischer Scientific. Protected sugars which
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were not synthesised in the lab were supplied by Carbosynth. Glassware was oven-dried and purged
with nitrogen immediately before use, and reactions requiring inert atmosphere were run under N2.
Reactions were monitored by thin-layer chromatography (TLC) on aluminium-backed, pre-coated
silica gel plates (Silica Gel 60 F254, Merk) with the indicated eluents, and the TLC plates were
visualised under UV light (λ 254 nm) and charring by dipping in ethanol-sulfuric acid (95:5, v/v)
followed by heating. Preparative TLC was run on Analtech preparative uniplates (silica gel 1000
micron, 20 × 20 cm) and flash column chromatography (FCC) was performed on a Biotage Horizon
Isolera One using pre-packed SNAP ULTRA 25 µm silica gel cartridges.
NMR spectra were recorded using a Bruker Ultrashield Plus 400 MHz spectrometer at 298 K and
analysed using TopSpin software. Chemical shifts (δ) are reported in ppm with respect to internal
tetramethylsilane or the residual HOD signal in D2O. NMR assignments were made with the aid of
COSY and HSQCed experiments, with signals of glycosyl residues being distinguished from those of
aglycon by numbers with prime and double prime as shown in Figure 5. Note that compounds 23
and 24 are a mixture of R- and S-isomers; whilst we could not determine which isomer was R- and
which was S-, we could determine which molecule the signals related to via 2D NMR. Therefore, all
signals marked with an asterisk * are for one diastereoisomer; those without an asterisk * are the
other.
Figure 5 Numbering system used for assigning NMR spectra
Optical rotation values were measured using a Perkin Elmer® Model 341 Polarimeter at 20 °C at a
wavelength of 589 nm unless otherwise noted. Infrared spectra were recorded using a Perkin Elmer®
SpectrumBX and UV-vis spectra using a Varian 50 Bio spectrometer. Low resolution mass
spectrometry was employed for monitoring reactions using an Advion Expression CMS spectrometer
by direct injection or extraction from a TLC plate using an Advion Plate Express, with a 9:1 mixture of
methanol-aqueous formic acid (0.1%) used as a mobile phase. For high resolution mass
spectrometry (HRMS) 0.1% (w/v) samples were prepared by dissolving or diluting in methanol-
formic acid (1:1) and infused into a Synapt G2-Si mass spectrometer (Waters, Manchester, UK) at 5-
10 µl min-1 using a Harvard Apparatus syringe pump. The mass spectrometer was controlled by
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MassLynx 4.1 software (Waters). It was operated in high resolution and positive ion mode and
calibrated using sodium formate. The sample was analysed for 2 min with 1 s MS scan time over the
range of m/z 50-1200 with 3.5 kV capillary voltage, 40 V cone voltage, 120°C cone temperature. Leu-
enkephalin peptide (1 ng µL-1, Waters) was infused at 10 µl min-1 as a lock mass (m/z 556.2766) and
measured every 10 s. Spectra were generated in MassLynx 4.1 by combining a number of scans, and
peaks were centred using automatic peak detection with lock mass correction. For completeness,
synthetic procedures to produce all known compounds used in this study are included as electronic
Supplementary Information.
4.2. 1,3-Bis(benzyloxy)propan-2-yl 2,3,5,6-tetra-O-benzoyl-β-ᴅ-galactofuranoside (10)
A solution of per-O-benzoyl-β-ᴅ-galactofuranose (8)[44] (250 mg, 0.36 mmol) dissolved in CH2Cl2 (4
mL) was cooled in an ice bath. SnCl4 solution (1M in CH2Cl2, 400 µL, 0.4 mmol) was slowly added by
syringe. After stirring for 15 minutes at 0 °C, 1,3- di-O-benzyl-glycerol (9) (80 µL, 0.32 mmol) was
added by syringe and the mixture was allowed to warm up to room temperature and was stirred for
18 h after which TLC analysis (hexane-EtOAc 7:3) showed complete consumption of 8. The mixture
was diluted with CH2Cl2 (30 mL) and washed with sat. aqueous NaHCO3 (2 × 10 mL). The organic
layers were combined, dried over MgSO4, filtered and dried in vacuo to give a crude residue which
was purified by FCC to 10 (120 mg, 39%) as a colourless oil; Rf 0.61 (hexane/EtOAc 7:3); [α]D -1.2 (c
1.0, CHCl3); 1H NMR (400 MHz, CDCl3) 8.07-8.04 (m, 2H, Ar), 7.97-7.95 (m, 1H, Ar), 7.90-7.88 (m, 1H,
Ar), 7.57-7.20 (m, 25H, Ar), 6.03-5.99 (m, 1H, H-5’), 5.61 (s, 1H, H-1’), 5.59 (d, J3’,4’ = 5.4 Hz, 1H, H-3’),
5.55 (s, 1H, H-2’), 4.75 (dd, J3’,4’ = 5.4 Hz, J4’,5’ = 3.4 Hz, 1H, H-4’), 4.73 (dd, J5’,6’a = 7.7 Hz, 2J6’a,6’b = 12.0
2
2
Hz, 1H, H-6’a), 4.58-4.52 (m, 3H, H-6’b, PhCH2,), 4.51 (d, J = 11.9 Hz, PhCHH), 4.45 (d, J = 11.9 Hz,
PhCHH), 4.25-4.20 (m, 1H, H-2), 3.71-3.67 (m, 2H, H-1), 3,62 (d, J2.3 = 5.3 Hz, 2H, H-3); 13C NMR (100.6
MHz, CDCl3): 166.1, 165.8, 165.7, 165.4 (4 × C=O), 138.2, 138.1, 133.4, 133.3, 133.2, 133.0, 130.0,
129.9, 129.8, 129.7, 129.6, 129.2, 129.1, 128.4, 128.4, 128.3, 127.7, 127.6 (Ar), 105.0 (C1’), 82.1
(C2’), 81.4 (C4’), 77.8 (C3’), 74.6 (C2), 73.5, 73.5 (2 × PhCH2), 70.6 (C1), 70.3 (C5’), 70.2 (C3), 63.9
(C6’); HRMS (ESI+) m/z calc. for C51H46O12Na+ 873.2887 ([M+Na]+) found 873.2875 [M+Na]+.
4.3. 1,3-Dihydroxypropan-2-yl 2,3,5,6-tetra-O-benzoyl-β-ᴅ-galactofuranoside (11)
To a solution of 1,3-bis(benzyloxy)propan-2-yl 2,3,5,6-tetra-O-benzoyl-β-ᴅ-galactofuranoside (10)
(115 mg, 140 µmol) in EtOAc/EtOH (10:1) (20 mL) was added activated 10% palladium on charcoal
(10 mg). The system was flushed with N2 (×3) followed by H2 (×3) and stirred overnight at room
temperature. After the system had been flushed with N2 (×3) the catalyst was filtered off and the
solvent removed under reduced pressure to give 11 (60 mg, 64%) as a colourless oil; Rf 0.18 (hexane-
12

ACCEPTED MANUSCRIPT
EtOAc 1:1); [α]D = -1.1 (c 1.0, CHCl3); 1H NMR (400 MHz, CDCl3): 8.09-8.06 (m, 2H, Ar), 8.02-7.97 (m,
4H, Ar), 7.93-7.90 (m, 2H, Ar), 7.59-7.51 (m 4H, Ar), 7.49-7.30 (m, 8H, Ar), 6.00-5.96 (m 1H, H-5’),
5.73 (dd, J2’,3’ = 2.1 Hz, J3’,4’ = 5.7 Hz, 1H, H-3’), 5.52 (s, 1H, H-1’), 5.49 (d, J2’,3’ = 2.1 Hz, 1H, H-2’), 4.85
2
(dd, J3’,4’ = 5.7 Hz, J4’,5’ = 3.8 Hz, 1H, H-4’), 4.79 (dd, J5’,6’a = 4.6 Hz, J6’a,6’b = 11.8 Hz, 1H, H-6’a), 4.72
2
(dd, J5’,6’b = 6.6 Hz, J6’a,6’b = 11.8 Hz, 1H, H-6’b), 3.94-3.89 (m, 1H, H-2), 3.78-3.72 (m 4H, H-2,3), 2.56
(bs, 1H, OH), 2.47 (bs, 1H, OH) ; 13C NMR (100.6 MHz, CDCl3): 166.2, 166.2, 165.7, 165.6 (4 × C=O),
133.7, 133.7, 133.4, 133.2, 130.0, 129.9, 129.8, 129.5, 129.3, 128.8, 128.6, 128.5, 128.5, 128.4 (Ar),
106.5 (C1’), 83.3 (C2’), 81.1 (C4’), 80.8 (C2), 77.3 (C3’), 70.3 (C5’), 63.1 (C1), 63.1 (C6’), 62.4 (C3);
HRMS (ESI+) m/z calc. for C37H34O12Na+ 693.1948 ([M+Na]+) found 693.1956 [M+Na]+.
4.4. 1,3-Dihydroxypropan-2-yl β-ᴅ-galactofuranoside (1)
1,3-Dihydroxypropan-2-yl 2,3,5,6-tetra-O-benzoyl-β-ᴅ-galactofuranoside (11) (55 mg, 82 µmol) was
dissolved into a solution of MeOH/H2O/NEt3 (5:2:1, 8 mL) and stirred vigorously for 18h at room
temperature. The solvent was removed in vacuo, the crude mixture was dissolved in MeOH (5 mL)
and passed through Dowex® 1X2-400 (OH-) ion exchange resin (1 g). The eluted solution was
concentrated and the residue was dried in vacuo to give 1 (7 mg, 51%) as a colourless oil; Rf 0.06
(toluene/MeOH, 8:2); [α]D -144 (c 0.5, MeOH); 1H NMR (400 MHz, CD3OD): 4.99 (d, J1’,2’ = 1.7 Hz, 1H,
H-1’), 3.93-3.92 (m, 2H, H-3’,4’), 3.90 (dd, J1’,2’ = 1.7 Hz, J2’,3’ = 4.0 Hz, 1H, H-2’), 3.65-3.58 (m, 2H, H-
5’,2), 3.56-3.48 (6H, H-1,3,6’a,6’b); 13C NMR (100.6 MHz, CD3OD): 107.8 (C1’), 83.4 (C3’), 81.6 (C2’),
78.8 (C2), 77.3 (C4’), 71.0 (C5’), 63.0 (C6’), 61.9 (C1), 61.1 (C3); HRMS (ESI+) calc. for C9H18O8Na+
277.0899 ([M+Na]+) found 277.0895 [M+Na]+.
4.5. 1,3-Bis(benzyloxy)propan-2-yl 2,3,4-tri-O-benzoyl-α-ʟ-arabinopyranoside (14)
2,3,4-Tri-O-benzoyl-β-ʟ-arabinopyranosyl bromide (13)[25] (2.1 g, 4.0 mmol) was dried
azeotropically with dry toluene (3 × 20 mL), dissolved into dry toluene (10 mL) under N2 and then
1,3-O-di-benzyl-glycerol (9) (0.9 mL, 3.8 mmol) and Ag2CO3 (1.8 g, 6.4 mmol) were added. The
mixture was stirred at 55 °C under N2 for 4 hours after which time TLC (hexane/EtOAc 3:1) showed
complete consumption of the glycosyl bromide. The reaction mixture was filtered and the filtrate
was concentrated under reduced pressure. Purification by FCC gave 14 (1.5 g, 50%) as a colourless
oil, Rf 0.48 (toluene/EtOAc 9:1); [α]D +94.4 (c 1.0, CHCl3); 1H NMR (400 MHz, CDCl3): 8.05-7.94 (m, 5H,
Ar), 7.58-7.17 (m, 20H, Ar), 5.71 (dd, J1’,2’ = 5.9 Hz, J2’,3’ = 8.3 Hz, 1H, H-2’), 5.67-5.65 (m, 1H, H-4’),
5.60 (dd, J2’,3’ = 8.3 Hz, J3’,4’ = 3.5 Hz, 1H, H-3’), 5.06 (d, J1’,2’ = 5.9 Hz, 1H, H-1’), 4.35 (s, 2H, PhCH2),
2
4.37-4.32 (m, 3H, H-5a’, PhCH2), 4.15-4.12 (m, 1H, H-2), 3.82 (dd, J4’,5’ = 2.3 Hz, J5a’,5b’ = 12.6 Hz, 1H,
H-5b’), 3.69 (dd, J1a,2 = 5.3 Hz, 2J1a,1b = 10.3 Hz, 1H, H-1a), 3.63-3.59 (m, 2H, H-1b,3a), 3.50 (dd, J2,3b
=
13

ACCEPTED MANUSCRIPT
6.4 Hz, J3a,3b = 10.3 Hz, 1H, H-3b); 13C NMR (100.6 MHz, CDCl3): 165.7, 165.6, 165.2 (3 × C=O), 138.2,
138.1, 133.4, 133.3, 129.9, 129.8, 129.5, 129.2, 128.5, 128.4, 128.3, 127.7, 127.6, 127.5, 127.5 (Ar),
100.4 (C1’), 77.6 (C2), 73.4, 73.4 (2 × PhCH2), 70.8 (C1), 70.4 (C3’), 70.4 (C3), 70.2 (C2’), 68.3 (C4’),
62.0 (C5’); HRMS (ESI+) m/z calc. for C43H40O10Na+ 739.2519 [M+Na]+ found 739.2513 [M+Na]+.
4.6. 1,3-Dihydroxypropan-2-yl 2,3,4-tri-O-benzoyl-α-ʟ-arabinopyranoside (15)
To a solution of 1,3-bis(benzyloxy)propan-2-yl 2,3,4-tri-O-benzoyl-α-ʟ-arabinopyranoside (14) (1.50
g, 2.1 mmol) in EtOAc/EtOH 10:1 (50 mL) was added palladium on activated charcoal (10% Pd basis)
(50 mg). The system was flushed with N2 (×3) followed by H2 (×3) and stirred overnight at room
temperature. After the system had been flushed with N2 (×3) the catalyst was filtered off and the
solvent removed under reduced pressure to give 15 (182 mg, 22%) as a white powder. Rf 0.21
1
(EtOAc/hexane 6:4); [α]D +261 (c 1.0, CHCl3); H NMR (400 MHz, CDCl3): 8.10-8.08 (m, 2H, Ar), 8.01-
7.99 (m, 2H, Ar), 7.89-7.87 (m, 2H, Ar), 7.62-7.27 (m, 9H, Ar), 5.80 (dd, J1’,2’ = 7.3 Hz, J2’,3’ = 9.7 Hz, 1H,
H-2’), 5.73-5.71 (m, 1H, H-4’), 5.62 (dd, J2’,3’ = 9.7 Hz, J3’,4’ = 3.4 Hz, 1H, H-3’), 4.90 (d, J1’,2’ = 7.3 Hz, 1H,
2
2
H-1’), 4.36 (dd, J4’,5’a = 2.8 Hz, J5’a,5’b = 13.2 Hz, 1H, H-5’a), 3.96 (dd, J4’,5’b = 1.5 Hz, J5’a,5’b = 13.2 Hz,
1H, H-5’b), 3.86-3.83 (m, 1H, H-2), 3.69-3.67 (m, 2H, H-1), 3.58-3.55 (m, 2H, H-3), 2.85 (bs, 1H,
OH),2.05 (bs, 1H, OH); 13C NMR (100.6 MHz, CDCl3): 165.7, 165.6, 165.6 (3 × C=O), 133.7, 133.6,
133.5, 129.9, 129.8, 129.7, 129.3, 129.0, 128.9, 128.6, 128.6, 128.4, (12 × Ar), 102.0 (C1’), 83.9 (C2),
71.0 (C3’), 70.4 (C2’), 68.7 (C4’), 64.0 (C5’), 62.7 (C1), 62.4 (C3); HRMS (ESI)+ calc. for C29H28O10Na+
559.1580 [M+Na]+ found 559.1577 [M+Na]+.
4.7. 1,3-Dihydroxypropan-2-yl α-ʟ-arabinopyranoside (2)
Sodium (20 mg) was added to dry methanol (18 mL) under N2. After the cessation of effervescence
1,3-dihydroxypropan-2-yl 2,3,4-tri-O-benzoyl-α-ʟ-arabinopyranoside (15) (180 mg, 0.3 mmol) was
added in dry MeOH (8 mL) and the mixture was stirred overnight at room temperature. The reaction
mixture was neutralised using Amberlite IR-120 (H+) resin, filtered and the solvent removed in vacuo.
The crude mixture was dissolved in H2O and passed through Dowex® 1X2-400 (OH- form) resin (1 g)
27
1
to give 2 (62 mg, 76%) as a colourless oil. [α]D + 6.8 (c 1.0, H2O) [Lit: [α]D + 5 (H2O)]; H NMR (400
MHz, D2O): 4.35 (d, J1’,2’ = 7.5 Hz, 1H, H-1’), 3.83-3.78 (m, 2H, H-4’,5’a), 3.77-3.73 (m, 1H, H-2), 3.65-
3.62 (m, 6H, H-1,2,5’b,3’), 3.45 (dd, J1’,2’ = 7.5 Hz, J2’,3’ = 9.4 Hz, 1H, H-2’); 13C NMR (100.6 MHz, D2O):
103.4 (C1’), 81.1 (C2), 72.9 (C3’), 71.3 (C2’), 68.4 (C4’), 66.2 (C5’), 61.4 (C1), 61.0 (C3); HRMS (ESI+)
m/z calc. for C8H16O7Na+ 247.0794 [M+Na]+ found 247.0798 [M+Na]+.
14

ACCEPTED MANUSCRIPT
4.8. 1,3-Bis(benzyloxy)propan-2-yl 2,3,5-tri-O-benzyl-α-ᴅ-ribofuranoside (19a) and 1,3-
bis(benzyloxy)propan-2-yl 2,3,5-tri-O-benzyl-β-ᴅ-ribofuranoside (19b)
2,3,5-Tri-O-benzyl-β-ᴅ-ribofuranosyl fluoride (16)[35] (210 mg, 0.5 mmol), SnCl2 (95 mg, 0.5 mmol)
and triphenylmethyl perchlorate (170 mg, 0.5 mmol) were dissolved into dry Et2O (5 mL) containing
4Å MS (1.0g) under N2. The reaction vessel was wrapped with aluminium foil to exclude light, cooled
to -15 °C. 1,3-Di-O-benzyl glycerol (9) (100 µL, 0.40 mmol) was added in a single portion and the
reaction mixture was stirred at -15 °C for 6 hours before kept at 4 °C overnight. The reaction mixture
was diluted with Et2O (50 mL), filtered, and the filtrate washed with saturated aqueous NH4Cl
solution (3 × 10 mL). The organic layer was separated, dried over MgSO4 and filtered before the
solvent was removed under reduced pressure. The anomers were separated by FCC to give 19a and
19b as colourless oils.
1
The α-anomer 19a: Rf 0.10 (hexane/Et2O 7:3); [α]D +52.5 (c 1.0, CHCl3); H NMR (400 MHz, CDCl3):
7.32-7.18 (m, 25H, Ar), 5.38 (d, J1’,2’ = 4.3 Hz, 1H, H-1’), 4.73 -4.38 (m, 10H, 5 × PhCH2), 4.26-4.24 (m,
1H, H-4’), 4.23-4.17 (m, 1H, H-2), 3.82 (dd, J2’,3’ = 7.1 Hz, J3’,4’ = 3.9 Hz, 1H, H-3’), 3.75 (dd, J1’,2’ = 4.3
2
Hz, J2’,3’ = 7.1 Hz, 1H, H-2’), 3.74-3.59 (m, 4H, H-1,3), 3.41 (dd, J4’,5’a = 3.9 Hz, J5’a,5’b = 10.6 Hz, 1H, H-
2
5’a), 3.34 (dd, J4’,5’b = 4.2 Hz, J5’a,5’b = 10.6 Hz, 1H, H-5’b); 13C NMR (100.6 MHz, CDCl3): 138.6, 138.6,
138.4, 138.1, 138.0 (5 × Ar), 128.3, 128.3, 128.2, 128.0, 127.9, 127.6, 127.6, 127.6, 127.5, 127.5,
127.4 (Ar), 101.5 (C1’), 81.5 (C4’), 77.2 (C2’), 75.6 (C2), 75.5 (C3’), 73.4, 73.4, 73.3 72.2, 72.0 (5 ×
CH2Ph), 71.7 (C1), 70.7 (C3), 69.9 (C5’); HRMS (ESI+) m/z calc. for C43H46O7Na+ 697.3141 [M+Na]+
found 697.3129 [M+Na]+.
1
The β-anomer 19b: Rf 0.20 (hexane/Et2O 7:3); [α]D + 42.4 (c 1.0, CHCl3); H NMR (400 MHz, CDCl3):
7.31-7.22 (m, 25H, Ar), 5.30 (s, 1H, H-1’), 4.65-4.39 (m, 10H, PhCH2), 4.36-4.30 (m, 1H, H-4’), 4.05-
4.01 (m, 1H, H-2), 4.03 (dd, J2’,3’ = 4.8 Hz, J3’,4’ = 7.4 Hz, 1H, H-3’), 3.92 (d, J2’,3’ = 4.8 Hz, 1H, H-2’), 3.65-
3.42 (m, 6H, H-1,3,5’a,5’b); 13C NMR (100.6 MHz, CDCl3): 138.4, 138.3, 138.3, 138.0, 138.0, 128.4,
128.4, 128.3, 128.0, 127.8, 127.7, 127.6, 127.6, 127.5, 127.5 (Ar), 104.9 (C1’), 80.4 (C4’), 79.7 (C2’),
78.5 (C2), 74.8 (C3’), 73.4, 73.2, 73.0, 72.4, 72.1 (5 × PhCH2), 71.5 (C5’), 70.2 (C1), 70.2 (C3).
4.9. 1,3-Dihydroxypropan-2-yl α-ᴅ-ribofuranoside (3)
1,3-Bis(benzyloxy)propan-2-yl 2,3,5-tri-O-benzyl-α-ᴅ-ribofuranoside (19a) (80 mg, 120 µmol) was
dissolved in MeOH-EtOH (5:1, 12 mL) and palladium on activated charcoal (10% Pd basis) (50 mg)
was added. The system was flushed with N2 (×3) followed by H2 (×3) and stirred overnight at room
temperature. After the system had been flushed with N2 (×3) the catalyst was filtered through
Celite® and the filter was washed with AcOH (50 mL). The organic washes were combined and the
1
solvent was removed under reduced pressure to give 3 (20 mg, 75%); [α]D +68.2 (c 1.0, MeOH); H
15

ACCEPTED MANUSCRIPT
NMR (400 MHz, CD3OD): 5.11 (d, J1’,2’ = 4.3 Hz, 1H, H-1’) 4.01-3.89 (m, 3H, H-4’,2’,3’), 3.72-3.47 (m,
7H, H-2,5’a,5’b,1,3); 13C NMR (100.6 MHz, CD3OD): 102.0 (C1’), 85.9 (C4’), 79.0 (C2), 71.9 (C2’), 70.1
(C3’), 61.9 (C5’), 61.5 (C1), 60.8 (C3); HRMS (ESI+) m/z calc. for C8H16O7Na+ 247.0794 [M+Na]+ found
247.0798 [M+Na]+.
4.10. 1,3-Bis(benzyloxy)propan-2-yl 2,3,5-tri-O-benzyl-α-ʟ-xylofuranoside (20a) and 1,3-
bis(benzyloxy)propan-2-yl 2,3,5-tri-O-benzyl-β-ʟ-xylofuranoside (20b) 2,3,5-Tri-O-benzyl-α/β-ʟ-
xylofuranosyl fluoride (17) (200 mg, 0.47 mmol) and 1,3-di-O-benzyl glycerol (10) (100 µL, 0.4 mmol)
were dissolved into Et2O (5 mL) containing 4Å MS (1.0 g). The suspension was cooled in an ice bath
and SnCl2 (90 mg, 0.47 mmol) was added in a single portion to initiate the reaction. The reaction
mixture was then kept overnight at 4 °C, filtered and the filtrate was concentrated in vacuo to give a
1
crude product, which contained a mixture of α/β -glycosides in 1.0:0.8 ratio as judged by H NMR.
Purification by FCC afforded 20a and 20b as colourless oils.
The α-anomer 20a: colourless oil; yield 59 mg, (19%); Rf 0.48 (hexane/Et2O 7:3); [α]D -39.6 (c 1.0,
DCM); 1H NMR (400 MHz, CDCl3): 7.31-7.22 (m, 25H, Ar), 5.35 (d, J1’,2’ = 4.3 Hz, 1H, H-1’), 4.71 (d, 2J =
2
2
11.9 Hz, 1H, PhCHH), 4.63 (d, J = 11.9 Hz, 1H, PhCHH), 4.57 (d, J = 12.1 Hz, 1H, PhCHH), 4.53-4.45
(m, 7H,3.5 × PhCH2), 4.43-4.41 (m, 1H, H-4’), 4.31 (dd, J2’,3’ = 5.9 Hz, J3’,4’ = 7.0 Hz, 1H, H-3’), 4.20-4.15
(m, 1H, H-2), 3.97 (dd, J1’,2’ = 4.3 Hz, J2’,3’ = 5.9 Hz, 1H, H-2’), 3.74-3.54 (m, 6H, H-5’a,1,3,5’b); 13C NMR
(100.6 MHz, CDCl3): 138.4, 138.4, 138.3, 138.3, 137.9, 128.4, 128.4, 128.3, 128.3, 128.0, 127.7, 127.7,
127.6, 127.6, 127.5, 127.5 (Ar), 99.9 (C1’), 83.8 (C2;), 81.6 (C3’), 76.0 (C4’), 75.4 (C2), 73.4, 73.4, 73.3,
72.4, 71.8 (4 × PhCH2), 71.5 (C1), 70.4 (C3), 69.4 (C5’); HRMS (ESI+) m/z calc. for C43H46O7Na+
697.3141 ([M+Na]+) found 697.3129 [M+Na]+.
The β-anomer 20b: colourless oil; yield 32 mg(12%); Rf 0.54 (hexane/Et2O 7:3); [α]D +5.6 (c 1.0,
1
DCM); H NMR (400 MHz, CDCl3): 7.32-7.23 (m, 25H, Ar), 5.31 (d, J1’,2’ = 1.9 Hz, 1H, H-1’), 4.58-4.40
(m, 11H, 5 × PhCH2, H-4’), 4.11-4.06 (m, 3H, H-2’,3’,2), 3.76 (dd, J4’,5a’ = 4.9 Hz, 2J5’a,5’b = 10.3 Hz, 1H, H-
2
5’a), 3.71 (dd, J4’,5’b = 3.7 Hz, J5’a,5’b = 10.3 Hz, 1H, H-5’b), 3.71-3.55 (m, 4H, H-1,3); 13C NMR (100.6
MHz, CDCl3): 138.5, 138.4, 138.3, 138.0, 137.7, 128.4, 128.3, 128.3, 128.3, 127.8, 127.7, 127.7, 127.6,
127.6, 127.6, 127.5, 127.5, 127.5 (Ar), 107.0 (C1’), 86.8 (C2’), 82.0 (C3’), 79.9 (C4’), 75.7 (C2), 73.4,
73.3, 73.3, 72.0, 71.8 (5 × PhCH2), 70.7 (C1), 70.4 (C3), 69.8 (C5’);
4.11. 1,3-Dihydroxypropan-2-yl α-ʟ-xylofuranoside (4)
1,3-Bis(benzyloxy)propan-2-yl 2,3,5-tri-O-benzyl-α-ʟ-xylofuranoside (20a) (100 mg, 145 µmol) was
dissolved in a mixture of MeOH/n-PrOH (9:1) (20 mL) and palladium on activated charcoal (10% Pd
16

ACCEPTED MANUSCRIPT
basis) (50 mg) was added. The system was flushed with N2 (×3) followed by H2 (×3) and stirred
overnight at room temperature. After the system had been flushed with N2 (×3) the catalyst was
filtered through Celite® and the filter was washed with MeOH (20 mL). The organic washes were
combined and the solvent was removed under reduced pressure to give 4 (16 mg, 45%) as a
1
colourless oil; Rf 0.28 (DCM/MeOH 85:15); [α]D -139 (c 1.0 MeOH); H NMR (400 MHz, CD3OD): 5.09
(d, J1’2’ = 4.4 Hz, 1H, H-1’), 4.16-4.08 (m, 2H, H-3’,4’), 3.92 (dd, J1’,2’ = 4.4 Hz, J2’,3’ = 4.4 Hz, 1H, H-2’),
3.65-3.51 (m, 7H, H-2,1,3,5’); 13C NMR (100.6 MHz, CD3OD): 101.3 (C1’), 80.0 (C2), 78.7 (C4’), 78.1
+
(C2’), 75.7 (C3’), 61.7 (C5’), 61.2 (C1), 61.0 (C3); (HRMS ESI+) m/z calc for C8H16NaO7 247.0794
([M+Na]+) found 247.0785 [M+Na]+.
4.12. 1,3-Bis(benzyloxy)propan-2-yl 2,3,5-tri-O-benzyl-β-ᴅ-arabinofuranoside (21a) and 1,3-
bis(benzyloxy)propan-2-yl 2,3,5-tri-O-benzyl-α-ᴅ-arabinofuranoside (21b)
2,3,5-Tri-O-benzyl-α,β-ᴅ-arabinofuranosyl fluoride (18) (950 mg, 2.3 mmol) and 1,3-di-O-benzyl
glycerol (9) (540 µL, 2.2 mmol) were dissolved into a suspension of 4Å MS (1.0 g) in Et2O (7 mL). The
suspension was cooled in an ice bath and SnCl2 (440 mg, 2.3 mmol) was added in a single portion to
initiate the reaction. The reaction mixture was then kept in a refrigerator overnight at 4 °C, the solid
materials were filtered off and the solvent removed in vacuo to give a mixture of anomers which had
very close Rf. Three rounds of purification by FCC gave the β-anomer 21a (560 mg), α-anomer 21b
(47 mg) and some remaining unseparated mixture of both.
Compound 21a: colourless oil; Rf 0.46 (hexane/Et2O 6:4); [α]D -38.7 (c 1.0, CHCl3); 1H NMR (400 MHz,
CDCl3): 7.32-7.24 (m, 25H, Ar), 5.32 (d, J1’,2’ = 4.4 Hz, 1H, H-1’), 4.71-4.36 (m, 10H, 5 × PhCH2), 4.13-
4.06 (m, 3H, H-3’,4’,2), 4.03 (dd, J1’,2’ = 4.4 Hz, J2’,3’ = 7.0 Hz, 1H, H-2’), 3.71 (dd, J4’,5’ = 3.4 Hz, 2J5a’,5b’
=
10.3 Hz, 1H, H-5a’), 3.64-3.47 (m, 5H, H-5b’,1,3); 13C NMR (100.6 MHz, CDCl3): 138.4, 138.3, 138.3,
138.2, 137.9 (5 × Ar), 128.4, 128.4, 128.4, 128.3, 128.0, 127.8, 127.7, 127.7, 127.7, 127.6, 127.6,
127.6, (Ar), 100.8 (C1’), 83.7 (C2’), 83.1 (C3’), 80.1 (C4’), 75.6 (C2), 73.5, 73.3, 73.1 (3 × PhCH2), 72.6
(C1), 72.3, 71.8 (2 × PhCH2), 71.3 (C5’), 70.3 (C3); HRMS (ESI+) m/z calc. for C43H46O7 697.3136
[M+Na]+ found 697.3132 [M+Na]+.
1
Compound 21b: colourless oil; Rf 0.46 (hexane/Et2O 6:4); [α]D + 26.8 (c 1.0, CHCl3); H NMR (400
MHz, CDCl3): 7.35-7.22 (m, 25H, Ar), 5.35 (bs, 1H, H-1’), 4.57-4.41 (m, 10H, 5 × PhCH2), 4.23 (dm, J3’,4’
= 7.3 Hz, 1H, H-4’), 4.15-4.09 (m, 1H, H-2), 4.08 (d, J2’,3’ = 3.4 Hz, 1H, H-2’), 3.92 (dd, J2’,3’ = 3.4 Hz, J3’,4’
= 7.3 Hz, 1H, H-3’), 3.70 (dd, J1,2 = 3.8 Hz, 2J1a,1b = 10.2 Hz, 1H, H-1a), 3.65-3.54 (m, 5H, H-1b,3,5’); 13
C
NMR (100.6 MHz, CDCl3): 138.4, 138.3, 138.2, 138.0, 137.7 (5 × Ar), 128.4, 128.3, 128.0, 127.8, 127.7,
127.7, 127.6, 127.6, 127.5 (Ar), 106.1 (C1’), 88.5 (C2’), 83.7 (C3’), 80.3 (C4’), 75.0 (C2), 73.4 73.3,
17

ACCEPTED MANUSCRIPT
73.3, 72.1, 71.8 (5 × PhCH2), 70.8 (C1), 70.5 (C3), 69.6 (C5’); HRMS (ESI+) m/z calc. for C43H46O7
697.3136 [M+Na]+ found 697.3132 [M+Na]+.
4.13. 1,3-Dihydroxypropan-2-yl β-ᴅ-arabinofuranoside (5)
1,3-Bis(benzyloxy)propan-2-yl 2,3,5-tri-O-benzyl-β-ᴅ-arabinofuranoside (21a) (560 mg, 0.8 mmol)
was dissolved in a mixture of MeOH/EtOAc (9:1) (50 mL) and palladium on activated charcoal (10%
Pd basis) (50 mg) was added. The system was flushed with N2 (×3) followed by H2 (×3) and stirred for
48 hours at room temperature. After the system had been flushed with N2 (×3) the catalyst was
filtered through Celite® and the filter was washed with MeOH (20 mL). The organic washes were
combined and the solvent was removed under reduced pressure to give 5 (80 mg, 45%) as a
colourless oil; Rf 0.45 (DCM/MeOH 85:15); [α]D -78.1 (c 1.0, MeOH); 1H NMR (400 MHz, CD3OD): 5.05
(d, J1’,2’ = 4.7 Hz, 1H, H-1’), 4.18 (dd, J2’,3’ = 8.0 Hz, J3’,4’ = 8.0 Hz, 1H, H-3’), 4.00 (dd, J1’,2’ = 4.7 Hz, J2’,3’
=
8.0 Hz, 1H, H-2’), 3.78-3.73 (m,3H, H-1,4’), 3.72-3.68 (m, 1H, H-2), 3.66-3.61 (m, 4H, H-5’,3); 13C NMR
(100.6 MHz, CD3OD): 101.3 (C1’), 82.5 (C2), 80.8 (C4’), 77.6 (C2’), 73.5 (C3’), 61.7 (C5’), 61.4 (C1,C3);
HRMS (ESI+) m/z calc. for C8H16O7Na+ 247.0794 [M+Na]+ found 247.0788 [M+Na]+.
4.14. Prop-2-en-1-yl 2,3,5-tri-O-benzoyl-α-ʟ-arabinopyranoside (22)
A mixture of 2,3,4-tri-O-benzoyl-β-ʟ-arabinopyranosyl bromide (13)[25] (1.8 g,3.5 mmol), allyl
alcohol (290 µL, 4.2 mmol) and 4Å MS (2.0 g) in DCE (30 mL) was stirred at room temperature for 30
minutes. Silver carbonate (1.2 g, 4.2 mmol) was added and stirring continued in the dark at room
temperature overnight. The reaction mixture was filtered through Celite® and the volatile
components were evaporated in vacuo to give a crude syrup which was purified by FCC to give α-
glycoside 22 (1.1 g, 62%) as a colourless oil; Rf 0.64 (hexane/EtOAc 7:3); [α]D +106° (c = 1.0, CHCl3); 1H
NMR (400 MHz, CDCl3): 8.05 – 8.01 (m, 4H, Ar), 7.95-7.93 (m, 2H, Ar), 7.59-7.31 (m, 9H, Ar), 5.86 (m,
1H, H-2), 5.74 (dd, J1’,2’ = 6.1 Hz, J2’,3’ = 8.7 Hz, 1H, H-2’), 5.71-5.68 (m, 1H, H-4’), 5.62 (dd, J2’,3’ = 8.7
Hz, J3’,4’ = 3.4 Hz, 1H, H-3’), 5.29 (dq, J2,3a = 17.3 Hz, 2J3a,3b = 1.7 Hz, 1H, H-3a), 5.17 (dq, J2,3b = 10.5 Hz,
2J3a,3b = 1.7 Hz, 1H, H-3b), 4.82 (d, J1’,2’ = 6.1 Hz, 1H, H-1’), 4.38 (ddt, J1a,1b = 13.1 Hz, J1a,2 = 5.0 Hz, 4J1a,3
= 1.6 Hz, 1H, H-1a), 4.33 (dd, J4,5a’ = 4.4 Hz, 2J5a’,5b’ = 12.7 Hz, 1H, H-5a’), 4.16 (ddt, J1a,1b = 13.1 Hz, J1b,2
= 6.2 Hz, 4J1b,3 = 1.3 Hz, 1H, H-1b), 3.90 (dd, J4,5b’ = 2.3 Hz, 2J5a’,5b’ = 12.7 Hz, 1H, H-5b’); 13C NMR (100.6
MHz, CDCl3): 165.7, 165.6, 165.3 (3 × C=O), 133.5 (C2), 133.4, 133.3, 133.3, 129.9, 129.9, 129.8,
129.8, 129.4, 129.4, 129.1, 128.5, 128.5, 128.4, 117.8 (C3), 99.4 (C1’), 70.5 (C3’), 70.0 (C2’), 69.7 (C1),
68.3 (C4’), 62.3 (C5’); HRMS (ESI+) m/z calc. for C29H26O8Na+ 525.1525 [M+Na]+ found 525.1522
[M+Na]+.
18

ACCEPTED MANUSCRIPT
4.15. (R,S)-Oxiranyl)methyl 2,3,5-tri-O-benzoyl-α-ʟ-arabinopyranoside (23)
Prop-2-en-1-yl 2,3,5-tri-O-benzoyl-α-ʟ-arabinopyranoside (22) (1.1 g, 2.2 mmol) and mCPBA (450 mg,
2.6 mmol) were dissolved into DCE (20 mL) and heated to 80 °C overnight. The solvent was removed
in vacuo, the crude product was dissolved into EtOAc (10 mL) and washed with sat. NaHCO3 solution
(3 × 3 mL). The organic layer was separated and dried over MgSO4, then filtered and the solvent
evaporated in vacuo to give 23 (640 mg, 45%) as a white powder (1:1 R,S-mixture, as judged by the
integration of the H-1’ signals at 4.92 and 4.82 ppm). Rf 0.5 (hexane/EtOAc 7:3); 1H NMR (400 MHz,
CDCl3): 8.08-7.94 (m, 12H, Ar), 7.60-7.32 (m, 18H, Ar), 5.73 (dd, J1’,2’ = 6.0 Hz, J2’,3’ = 8.4 Hz, 2H, H-
2’,2’*), 5.70-5.69 (m, 2H, H-4’,4’*), 5.62 (dd, J2’3’ = 8.4 Hz, J3’,4’ = 3.4 Hz, 2H, H-3’,3’*), 4.92 (d, J1’a,2’
6.0 Hz, 1H, H-1’), 4.82 (d, J1’b,2’ = 6.0 Hz, 1H, H-1’*), 4.37-4.31 (m, 2H, H-5a’,5a’*), 4.09 (dd, J1a,1b
=
=
12.0 Hz, J1a,2 = 3.0 Hz, 1H, H-1a), 3.92-3.83 (m, 4H, H-5b’,5b’*,1a*, 1b*), 3.19-3.12 (m, 2H, H-2, 2*),
2.76-2.73 (m, 2H, H-3a,3a*), 2.62 (dd, 2J3b*,3a* = 5.1 Hz, J3b*,2* = 2.6 Hz, 1H, H-3b*), 2.58 (dd, 2J3b*,3a*
=
5.1 Hz, J3a*,2* = 2.6 Hz, 1H, H-3a*); 13C NMR (100.6 MHz, CDCl3): 165.7, 165.6, 165.3, (3 × C=O), 133.4,
133.4, 133.3, 130.2, 129.9, 129.8, 129.4, 129.3, 129.1, 128.5, 128.4, 128.3, 100.5 (C1’), 100.2 (C1’*),
70.4 (C3’), 70.2 (C3’*), 69.9, (C2’), 69.9 (C2’*), 69.9 (C1), 68.9 (C1*), 68.2 (C4), 68.1 (C4*), 50.7 (C2),
50.4 (C2*), 44.3 (C3), 44.1 (C3*); HRMS (ESI+) m/z calc. for C29H26O9Na+ 541.1469 [M+Na]+ found
541.1467 [M+Na]+. Two sets of signals belonging to spin systems of two individual diastereomers
are distinguished by marking one of the sets with asterisk.
4.16. 3-(3-Azidopropoxy)-2-(R,S)-hydroxypropyl 2,3,4-tri-O-benzoyl-α-ʟ-arabinopyranoside (24)
(R,S)-Oxiranyl)methyl 2,3,5-tri-O-benzoyl-α-ʟ-arabinopyranoside (23) (1.0 g, 1.9 mmol), 3-azido
propanol (230 µL, 2.5 mmol) and Sc(OTf)3 (140 mg, 0.3 mmol) were dissolved into toluene (100 mL)
and stirred vigorously at room temperature overnight. The reaction mixture was then washed with
sat. NaHCO3 solution (3 × 30 mL) and the organic layer dried over MgSO4, filtered and the solvent
evaporated in vacuo. The crude product was then purified by FCC to give 24 (480 mg, 40%) as a
1
colourless oil (1:1 mixture of diastereoisomers as judged by the integration of the H signals at 4.81
1
and 4.79 ppm). Rf 0.27 (hexane/EtOAc 7:3); H NMR (400 MHz, CDCl3): 8.08-8.00 (m, 8H, Ar), 7.93-
7.90 (m, 4H, Ar), 7.61-7.30 (m, 18H, Ar), 5.74 (dd, J1’,2’ = 6.5 Hz, J2’,3’ = 8.9 Hz, 2H, H-2’, H-2’*), 5.71-
5.67 (m, 2H, H-4’, H-4’*), 5.61 (dd, J2’,3’ = 8.9 Hz, J3’,4’ = 3.5 Hz, 2H, H-3’,3’*), 4.81 (d, J1’,2’ = 6.5 Hz, 1H,
H-1’), 4.79 (d, J1’*,2’* = 6.5 Hz, 1H, H-1’*), 4.34 (dd, J5’a,5’b = 12.9 Hz, J4’,5’a = 3.7 Hz, 2H, H-5’a,5’a*), 3.98-
2
3.86 (m, 6H, H-5’b,5’b*,2,2*,3a,3a*), 3.75 (dd, J3a,3b = 10.2 Hz, J2,3a = 4.4 Hz, 1H, H-3b), 3.67 (dd,
2J3a*,3b* = 9.8 Hz, J2,3a* = 3.5 Hz, 1H, H-3b*), 3.45-3.28 (m, 10H, H-4,1a,1a*,1b,1b*,6,6*), 2.35 (s, 1H,
OH), 2.17 (s, 1H, OH*), 1.78-1.73 (m, 2H, H-5,5*); 13C NMR (100.6 MHz, CDCl3): 165.7, 165.6, 165.3 (3
× C=O), 133.5, 133.4, 129.9, 129.9, 129.8, 129.4, 129.3, 129.2, 129.0, 128.5, 128.5, 128.4, 128.2,
19

ACCEPTED MANUSCRIPT
101.5 (C1), 101.4 (C1*), 71.6 (C4), 71.6 (C4*), 71.3 (C3), 71.1 (C3*), 70.6 (C3’3’*), 69.3 (C2), 69.3
(C2*), 68.4 (C4’,4’*), 68.0 (C1,1*), 63.0 (C5’,5’*), 48.4 (C6,6*), 29.0 (C5), 28.9 (C5*); HRMS (ESI+) m/z
calc. for C32H33N3O10Na+ 642.2058 [M+Na]+ found 642.2051 [M+Na]+. Two sets of signals belonging
to spin systems of two individual diastereomers are distinguished by marking one of the sets with
asterisk.
4.17.
(2R)-3-(3-Azidopropoxy)-2-(2,3,5-tri-O-benzyl-α-ᴅ-ribofuranosyloxy)propyl
2,3,4-tri-O-
benzoyl-α-ʟ-arabinopyranoside (27) and (2S)-3-(3-azidopropoxy)-2-(2,3,5-tri-O-benzyl-α-ᴅ-
ribofuranosyloxy)propyl- 2,3,4-tri-O-benzoyl-α-ʟ-arabinopyranoside (28)
A suspension of 3-(3-azidopropoxy)-2-hydroxypropyl 2,3,4-tri-O-benzoyl-α-ʟ-arabinopyranoside (24)
(200 mg, 0.3 mmol), 2,3,5-tri-O-benzyl-1-(N-phenyl)-2,2,2-trifluoroacetimidoyl)-β-ᴅ-ribofuranose
(25)[40] (270 mg, 0.45 mmol) and 4Å MS (1.0 g) in dry dichloromethane (10 mL was stirred under a
nitrogen atmosphere for 30 minutes. The mixture was then cooled to -78 °C and TMSOTf (11 µL, 60
µmmol) was added by syringe. After 2 hours of stirring at -78 °C, the reaction was quenched with
Et3N (20 µL) and the solvents were removed in vacuo. Excess of the acceptor was removed by FCC
and the diastereomeric products were then separated by preparative TLC (hexane/EtOAc 6:4) to give
27 (45 mg, 15%) and 28 (30 mg, 10%) as colourless oils.
Compound 27: Rf 0.45 (hexane/EtOAc 6:4); [α]D + 95.9 (c 1.0, CHCl3); 1H NMR (400 MHz, CDCl3): 8.01-
8.00 (m, 4H, Bz), 7.87 (dd, J = 8.2 Hz, J = 1.0 Hz, 2H, Bz), 7.56-7.22 (m, 24sH, Ar), 5.74 (dd, J1’,2’ = 7.0
Hz, J2’,3’ = 9.4 Hz, 1H, H-2’), 5.60-5.58 (m, 1H, H-4’), 5.52 (dd, J2’,3’ = 9.4 Hz, J3’,4’ = 3.6 Hz, 1H, H-3’),
2
5.21 (d, J1’’,2’’ = 4.2 Hz, 1H, H-1’’), 4.88 (d, J1’,2’ = 7.0 Hz, 1H, H-1’), 4.71 (d, J = 12.1 Hz, 1H, CHHPh),
4.69 (d, 2J = 12.1 Hz, 1H, CHHPh), 4.56 (d, 2J = 11.7 Hz, 1H, CHHPh), 4.53 (d, 2J = 11.7 Hz, 1H, CHHPh),
4.50 (d, 2J = 11.9 Hz, 1H, CHHPh), 4.44 (d, 2J = 11.9 Hz, 1H, CHHPh), 4.24 (dd, J3’’,4’’ = 8.3 Hz, J4’’,5’’ = 4.0
2
Hz, 1H, H-4’’), 4.18 (dd, J4’,5a’ = 3.3 Hz, J5a’,5b’ = 13.1 Hz, 1H, H-5a’), 4.02-3.95 (m, 2H, H-3a,2), 3.85-
2
3.79 (m, 2H, H-3b,3’’), 3.72-3.69 (m, 2H, H-5b’,2’’), 3.50 (dd, J4’’,5’’a = 5.0 Hz, J5a’’,5b’’ = 10.3 Hz, 1H, H-
5a’’), 3.45-3.39 (m, 2H, H-5b’’,1), 3.34-3.27 (m, 2H, H-4), 3.22 (t, J5,6 = 7.0 Hz, 2H, H-6), 1.67-1.61 (m,
2H, H-5); 13C NMR (100.6 MHz, CDCl3): 165.7, 165.6, 165.3 (3 × C=O), 133.3, 133.3, 129.9, 129.8,
129.4,129.4, 129.1, 128.5, 128.4, 128.4, 128.3, 128.1, 128.0, 127.7, 127.7, 127.6, 101.5 (C1’’), 101.1
(C1’), 81.6 (C4’’), 77.7 (C2’’), 75.7 (C2), 75.5 (C3’’), 73.5 (CH2Ph), 72.3 (CH2Ph), 72.3 (CH2Ph), 71.0
(C3’), 70.6 (C5’’), 70.2 (C2’), 70.0 (C1), 69.2 (C3), 68.9 (C4’), 67.9 (C4), 63.3 (C5’), 48.4 (C6), 29.0 (C5);
HRMS (ESI+) m/z calc. for C58H59N3O14Na+1044.3889 [M+Na]+ found 1044.3885 [M+Na]+.
Compound 28: Rf 0.53 (hexane-EtOAc 6:4); [α]D +120 (c 1.0, CHCl3); 1H NMR (400 MHz, CDCl3): 8.04-
8.00 (m, 4H, Bz), 7.93 (dd, J = 8.2 Hz, J = 1.1 Hz, 2H, Bz), 7.57-7.19 (m, 24H, Ar), 5.67 (dd, J1’,2’ = 6.0 Hz,
20

ACCEPTED MANUSCRIPT
J2’,3’ = 7.9 Hz, 1H, H-2’), 5.56 (m, 2H, H-4’,3’), 5.23 (d, J1’’,2’’ = 4.4 Hz, 1H, H-1’’), 4.91 (d, J1’,2’ = 6.0 Hz,
1H, H-1’), 4.69 (d, 2JCHHPh, CHHPh = 10.0 Hz, 1H, CHHPh), 4.66 (d, 2JCHHPh, CHHPh = 10.0 Hz, 1H, CHHPh), 4.56
(d, 2JCHHPh, CHHPh = 15.2 Hz, 1H, CHHPh), 4.53 (d, 2JCHHPh, CHHPh = 15.2 Hz, 1H, CHHPh), 4.46 (d, 2JCHHPh, CHHPh
= 12.2 Hz, 1H, CHHPh), 4.40 (d, 2JCHHPh, CHHPh = 12.2 Hz, 1H, CHHPh), 4.27 (dd, J3’’,4’’ = 7.4 Hz, J4’’,5’’ = 3.7
2
Hz, 1H, H-4’’), 4.20 (dd, J4’,5a’ = 3.8 Hz, J5a’,5b’ = 12.8 Hz, 1H, H-5a’), 4.03-4.00 (m, 1H, H-2), 3.92 (dd,
J2,3a = 6.3, J3a,3b = 10.9, 1H, H-3a), 3.86-3.81 (m, 2H, H-3’’,3b), 3.77 (dd, J1’’,2’’ = 4.4 Hz, J2’’,3’’ = 6.5 Hz,
2
1H, H-2’’), 3.72 (dd, J4’,5b’ = 1.8 Hz, J5a’,5b’ = 12.8 Hz, 1H, H-5b’), 3.51 (d, J1,2 = 5.4 Hz, 2H, H-1), 3.36
(dd, J4’’,5’’ = 3.7 Hz, 2J5a’’,5b’’ = 3.7 Hz, 2H, H-5a’’,5b’’), 3.34-3.26 (m, 2H, H-4), 3.27 (t, J5,6 = 6.8 Hz, 2H, H-
6), 1.69-1.62 (m, 2H, H-5); 13C NMR (100.6 MHz, CDCl3): 165.7, 165.6, 165.3 (3 × C=O), 138.4, 138.1,
138.0, 133.3, 133.3, 129.9, 129.9, 129.5, 129.2, 128.5, 128.4, 128.4, 128.4, 128.4, 128.3, 128.1,
127.8, 127.7, 127.7, 127.6, 127.6, 127.6, 101.4 (C1’’), 100.5 (C1’), 81.8 (C4’’), 75.7 (C2), 75.6 (C3’’),
74.4 (CH2Ph), 72.4 (CH2Ph), 72.2 (CH2Ph), 71.5 (C1), 70.5 (C3’), 70.1 (C2’), 70.0 (C5’’), 67.0 (C3), 68.5
(C4’), 67.9 (C4), 62.2 (C5’), 48.4 (C6), 29.1 (C5); HRMS (ESI+) m/z calc. for C58H59N3O14Na+ 1044.3889
[M+Na]+ found 1044.3890 [M+Na]+.
4.18. General procedure for deprotection of compounds 27 and 28.
(2R)- or (2S)-3-(3-Azidopropoxy)-2-(2,3,5-tri-O-benzyl-α-ᴅ-ribofuranosyloxypropyl 2,3,4-tri-O-
benzoyl-α-ʟ-arabinopyranoside (27) or (28) respectively was dissolved in a mixture of MeOH (10 mL)
and EtOAc (5 mL) and hydrogenated over 10% Pd/C (5 mg) under a H2 at 50 °C for 24 hours. The
resulting mixture was filtered through Celite®, the filter was washed with additional MeOH (25 mL)
and the solvent was removed in vacuo. The residue was dissolved in a mixture of MeOH/H2O/Et3N
(5:2:1 mL) and stirred at room temperature overnight. The mixture was concentrated in vacuo and
the residue was passed through a short column of Dowex® 1X2-400 resin (OH- form) in water. The
sample was lyophilised to give target compounds 6 and 7.
(2R)-3-(3-Aminopropoxy)-2-(α-ᴅ-ribofuranosyloxy)propyl α-ʟ-arabinopyranoside (6) was prepared
from 27 (45 mg, 44 µmol) as a white powder (10.7 mg, 59%); [α]D +40.4 (c = 1.0, H2O); 1H NMR (400
MHz, D2O): 5.24 (d, J1’’,2’’ = 4.4 Hz, 1H, H-1’’), 4.29 (d, J1’,2’ = 7.7 Hz, 1H, H-1’); 4.07 (dd, J3’’,4’’ = 6.6 Hz,
J4’’,5a’’ = 3.7 Hz, 1H, H-4’’), 4.04-4.00 (m, 2H, H-4’,2’’), 3.95 (dd, J2’’,3’’ = 3.5 Hz , J3’’,4’’ = 6.6 Hz, 1H, H-3’’),
3.90-3.83 (m, 4H, H-3a, 2, 5a’), 3.75 (dd, J2,3b = 4.4 Hz, J3a,3b = 11.4 Hz, 1H, H-3b), 3.68 (dd, J4’’,5a’’ = 3.7
Hz, J5a’’,5b’’ = 12.4 Hz), 1H, H-5a’’), 3.63-3.46 (m, 8H, H-1,5b’’,3’,5’,4,2’), 2.60 (t, J5,6 = 6.9 Hz, 1H, H-6),
1.70-1.61 (m, 1H, H-5); 13C NMR (100.6 MHz, D2O): 103.4 (C1’), 101.9 (C1’’), 84.7 (C4’’) 75.7 (C4’),
72.3 (C3’), 71.3 (C2’’), 70.7 (C2’), 70.2 (C1), 69.6 (C3’’), 69.3 (C4), 69.1 (C3), 68.2 (C2), 66.2 (C5’), 61.5
(C5’’), 37.7 (C6), 31.4 (C5); HRMS (ESI+) m/z calc. for C16H31NO11Na+ 414.1970 [M+Na]+ found
414.1968 [M+Na]+
.
21

ACCEPTED MANUSCRIPT
(2S)-3-(3-Aminopropoxy)-2-(α-ᴅ-ribofuranosyloxypropyl) α-ʟ-arabinopyranoside (7) was prepared by
deprotection of (28) (30 mg, 29 µmol) as a white powder; yield 7.4 mg (41 %); [α]D +35.5 (c 1.0,
H2O); 1H NMR (400 MHz, D2O): 5.24 (t, J1’’,2’’ = 3.9 Hz, 1H, H-1’’), 4.29 (d, J1’,2’ = 7.6 Hz, 1H, H-1’), 4.11
(dd, J3’’,4’’ = 7.6 Hz, J4’’,5’’ = 3.7 Hz, 1H, H-4’’), 4.05-3.92 (m, 4H, H-4’,2’’,3’’,3a), 3.87-3.82 (m, 2H, H-
5’,2), 3.74 (dd, J2,3a = 4.9 Hz, J3a,3b = 10.5 Hz, 1H, H-3b), 3.70-3.47 (m, 8H, H-1,5a’’,5’’b,3’,5b’,4,2’),
2.83 (t, J5,6 = 7.2 Hz, 1H, H-6), 1.80-1.73 (m, 1H, H-5); 13C NMR (100.6 MHz, D2O): 103.5 (C1’), 102.0
(C1’’), 84.7 (C4’’), 75.7 (C4’), 72.3 (C3’), 71.4 (C2’’), 70.7 (C2’), 69.7 (C1), 69.6 (C3’’), 69.6 (C3), 69.1
(C4), 68.2 (C2), 66.3 (C5’), 61.4 (C5’’), 37.7 (C6), 29.0 (C5); HRMS (ESI+) m/z calc. for C16H31NO11Na+
414.1970 [M+Na]+ found 414.1969 [M+Na]+.
5. Acknowledgements
These studies were supported in the UK by BBSRC Institute Strategic Programme on Understanding
and Exploiting Metabolism [BB/J004561/1] and the John Innes Foundation. We thank Gerhard
Saalbach for his assistance with recording HRMS spectra. MarvinSketch was used for displaying
lowest energy calculation 3D models, and Calculator Plugins were used for structure property
prediction and calculation. Marvin 15.1.19.0, 2015, ChemAxon (http://www.chemaxon.com).
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Highlights:
•
•
•
Synthesis of glyceryl glycosides inspired by carbohydrates on A-type prymnesins.
Di-glycosylated fragments of prymnesin-1 synthesised for bioconjugation studies.
NOESY NMR compared with computational models to assign fragment stereochemistry.