New syntheses for 11-(mercaptoundecyl)triethylene glycol and mercaptododecyltriethyleneoxy biotin amide

Article abstract of DOI:10.1016/j.tetlet.2005.04.091

Novel syntheses for mercaptododecyltriethyleneoxy biotin amide and 11-(mercaptoundecyl)triethylene glycol are presented here. Such alkyl thiols are popular components in creating monolayers capable of specifically binding proteins. The development of a variety of functionalized alkyl thiol compounds has a great impact on biosensor substrate design. In our synthesis of mercaptododecyltriethyleneoxy biotin amide, we couple aminotriethyleneoxydodecane disulfide to the NHS-activated biotin; this technique is amenable to attaching a carboxylated molecule of interest in order to create the functionalized alkyl thiol of choice. The 11-(mercaptoundecyl) triethylene glycol synthesis presented here is an alternative method easily completed in three steps.

Full text of DOI:10.1016/j.tetlet.2005.04.091

Tetrahedron
Letters
Tetrahedron Letters 46 (2005) 4813–4816
New syntheses for 11-(mercaptoundecyl)triethylene glycol and
mercaptododecyltriethyleneoxy biotin amide
Christie A. Canaria,a Jeffrey O. Smith,b C. J. Yu,a Scott E. Fraserb and Rusty Lansfordb,*
aChemistry Department, California Institute of Technology, 1200 E. California Blvd., Pasadena, CA91125, USA
bBiology Department, California Institute of Technology, 1200 E. California Blvd., Pasadena, CA91125, USA
Received 3March 2005; accepted 26 April 2005
Available online 31 May 2005
Abstract—Novel syntheses for mercaptododecyltriethyleneoxy biotin amide and 11-(mercaptoundecyl)triethylene glycol are pre-
sented here. Such alkyl thiols are popular components in creating monolayers capable of specifically binding proteins. The devel-
opment of a variety of functionalized alkyl thiol compounds has a great impact on biosensor substrate design. In our synthesis
of mercaptododecyltriethyleneoxy biotin amide, we couple aminotriethyleneoxydodecane disulfide to the NHS-activated biotin; this
technique is amenable to attaching a carboxylated molecule of interest in order to create the functionalized alkyl thiol of choice. The
11-(mercaptoundecyl)triethylene glycol synthesis presented here is an alternative method easily completed in three steps.
Ó 2005 Elsevier Ltd. All rights reserved.
1. Introduction
tation. Such SAMs were poorer at presenting functional
groups to a substrate–liquid interface.
Self-assembled monolayers (SAMs) are a popular tool
for tailoring the reactive properties of a surface. These
functionalized surfaces are used in liquid mediated as-
says such as DNA chips, protein chips, and carbohy-
drate chips. To make the monolayers, molecules may
be physisorbed from solution or vapor or more tightly
attached by covalent bonds, as with Au substrates and
alkyl thiols. In this case, functionalized alkyl thiols ad-
sorb as thin films onto Au substrates. Based on the func-
tional group of the alkyl thiol, substrates may be made
to bind or repel proteins,1,2 carbohydrates, and pep-
tides,3–5 DNA,6,7 or other small molecules.8 SAMs
based on alkyl thiols and disulfides and gold surfaces
have been well studied and there are numerous literature
reports.9–12 For example, long all-trans alkyl chains
(n > 12) of adjacent alkylthiolates align via van der
Waals interactions to form a packing of molecules with
a preferred orientation: the thiolate group is in contact
with Au and the alkyl chains tilted at a 30° angle from
normal.13–15 However, alkyl chains shorter than 9–10
were found to form monolayers that lacked rigid orien-
In our ongoing research project of developing novel
SAM-based biosensors, we designed and synthesized a
new linker containing alkyl thiols and triethylene glycol
functionalized with biotin (compound 2, Fig. 1). In this
letter, we also report the modified procedures to synthe-
size the known SAM substrate 11-(mercaptoundecyl)tri-
ethylene glycol (1). Biotin is a small molecule with a
great avidity and specificity for proteins such as avidin
and streptavidin. The binding affinity of biotin to avidin
(Ka ꢀ 1015
M
À1) is so great that the bond is considered
irreversible. Biotin is also able to bind with high
avidity to alternative proteins such as streptavidin (Ka ꢀ
1013 MÀ1) and neutravidin (Ka ꢀ 1011
M
À1). Like-
wise, biotin derivatives are capable of binding with
avidin-like proteins with various binding affinities.
While the biotin–streptavidin interaction is strong with
high specificity, these proteins have been shown to bind
in a non-specific fashion when presented with a surface
comprising only straight-chain alkyl thiolates.16 In crea-
ting a reactive surface, it is important to promote only
high affinity binding events while repelling non-specific
ones.
Keywords: Self-assembled monolayer; SAM; Biotin; Alkyl thiol; Strep-
tavidin; Biofunctionalization.
In addition, attachment of polyethylene glycol (PEG)
units on surfaces has been shown to reduce the non-
specific binding of proteins and even cells to the
*
Corresponding author. Tel.: +1 626 395 2004; fax: +1 626 584
9527; e-mail: rusty@gg.caltech.edu
0040-4039/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2005.04.091

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C. A. Canaria et al. / Tetrahedron Letters 46 (2005) 4813–4816
Figure 1. (1) Polyethylene glycol alkane thiol and (2) biotinylated alkane thiol.
substrates.17–20 In protein binding assays, the biotinyl-
ated SAMs with the best avidity for streptavidin are those
that present both biotin and PEG groups to the sub-
strate–liquid interface. The mechanism for protein resis-
tance by PEG is not fully understood, but a study by
Grunze et al. suggests that it is related to the interfacial
adsorption of water molecules with the PEG groups.21
The uncharged hydrophilic PEG oligo groups adopt
random conformations in aqueous solution and coordi-
nate very well with water. When a protein approaches
the PEG-covered surface, there is a loss in the available
volume and conformations for each oligomer, creating a
repulsive force. Since PEG is hydrophilic, the coordi-
nated water cannot be displaced by proteins as they ap-
proach the surface, and so the proteins are repelled
away.22 However, in the presence of a surface receptor
with high binding affinity to the solvated protein, the
thermodynamic stability of the protein–receptor pair
overcomes the repulsive effects of PEG and results in
protein adsorption. PEG groups are an important and
integral component of a functional and specific-binding
SAM; they are mostly inert and therefore may be incor-
porated with a number of functional groups besides bio-
tin. For example, substrates modified with PEG groups
are incorporated with peptides, quinones, and carbohy-
drates. Many of these substrates are under development
to create bio-sensing devices.
in decent yield.23,24 The results in our lab, however, were
inconsistent. Instead, we developed the synthesis shown
in Figure 2 to produce compound 1. In the first step, tri-
ethylene glycol is deprotonated in anhydrous DMF by
NaH, then reacted to large excess with 1,11-dibromo-
undecane to yield 11-(bromoundecyl)triethylene glycol
(41% yield). The introduction of thioacetate was
achieved by reacting 11-(bromoundecyl)triethylene gly-
col to sodium thioacetate, prepared by a reaction of so-
dium methoxide and thioacetic acid, giving the desired
product, 11-(thioacetylundecyl)triethylene glycol, in
excellent yield (93%). The deprotonation of thiol ester
was straightforward, via ammonia hydrolysis in metha-
nol as a solvent (54% yield).
A biotinylated alkylthiol reagent in the literature is de-
picted as compound 324,25 (Fig. 3). The body of the com-
pound consists of three sections: biotin, PEG, and alkyl
chain. In compound 3, an amide bond connects the
dodecyl chain and the ethylene glycol groups. Com-
pound 2 presented here has an ether bond connecting
the alkyl chain to the ethylene glycol group, more closely
mimicking the structure of 1. Another difference
between the syntheses of 2 and 3 is the direction in which
the compounds are made. As outlined in the procedures
by Nelson et al., the molecule is constructed by
the attachment of the PEG group to the biotin,
followed by the attachment of the alkyl chain to the
PEG group.
2. Synthesis
The synthesis of compound 12 is described in Figure 4.
The selective tosylation of triethylene glycol in two
steps using NaH and p-toluenesulfonyl chloride in
Literature reports the use of radical chemistry initiated
by photolysis to achieve the synthesis of compound 1
Figure 2. (a) (i) NaH/DMF, (ii) 1,11-dibromoundecane; (b) (i) NaH/CH3OH, (ii) CH3COSH; (c) NH4OH/CH3OH, 40 °C.

C. A. Canaria et al. / Tetrahedron Letters 46 (2005) 4813–4816
4815
Figure 3. Biotinylated alkane thiol described by Knoll and Nelson et al.
Figure 4. (a) N-hydroxysuccinimide/DCC/DMF; (b) (i) NaH/DMF, (ii) p-toluenesulfonyl chloride; (c) NaN3/DMF/H2O, 58 °C; (d) (i) NaH/DMF,
(ii) 1,12-dibromododecane; (e) (i) thiourea/EtOH/H2O, 88 °C (ii) NaOH; (f) (i) Ph3P/THF/H2O, (ii) ditertbutyl dicarbonate/THF/TEA; (g) TFA/
CH2Cl2/triisopropyl silane; (h) 7/DMF/TEA; (i) DTT/THF/MeOH/TEA.

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C. A. Canaria et al. / Tetrahedron Letters 46 (2005) 4813–4816
anhydrous DMF led to the formation of 5 (73% yield).
The azide 6 was prepared in almost quantitative yield
(99%) by stirring 5 in large excess of NaN3 in DMF/
H2O. Reaction of this compound to excess amount of
1,12 dibromododecane resulted in the attachment of
the azotriethylene glycol to the alkyl chain to yield 12-
(azotriethyleneoxy)-1-bromododecane 7 in low yield
(24% yield). The resulting bromoalkane 7 was refluxed
with thiourea in ethanol, followed by hydrolysis by
NaOH solution, affording the diazodo disulfide 8
(46% yield). The reduction of the azo group by triphen-
ylphosphine in THF provided the corresponding
amine; after protection by tert-butoxycarbonate, 9
was obtained in excellent yield (82% yield). The removal
of the tert-butoxycarbonate group by trifluoroacetic
acid (TFA) in CH2Cl2 gave ammoniumtriethyleneoxy-
dodecane disulfide 10 (80% yield). The introduction of
biotin to the linker through a reaction of the corre-
sponding amine to the activated biotin derivative 4
was straightforward. The benefit of this reaction is its
application for the attachment of a carboxylated mole-
cule of choice.26 The final compound 12 can be pre-
pared by a simple reduction of the disulfide 11 by
dithiothreitol (DTT) in basic medium.
References and notes
1. Spinke, J.; Liley, M.; Guder, H.-J.; Angermaier, L.; Knoll,
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In summary, we have designed and synthesized two bio-
functionalized alkyl thiols. The synthesis of 11-(mer-
captoundecyl)triethylene glycol 1 in three steps has
provided a feasible alternative method. Furthermore,
mercaptododecyltriethyleneoxy biotin amide 12 is novel
and viable for the formation of SAMs. The intermediate
aminotriethyleneoxydodecanethiol 11 will be very useful
for attachment of any activated carboxyl groups in
order to make novel receptors for biosensor develop-
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will be published separately.
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Acknowledgements
This research was funded under DARPA grant
MLR.00018-3AFOSR.000019. The authors thank B.
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BioNEMS group for support in substrate fabrication.
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Supplementary data
Materials, detailed synthesis, and product characteriza-
tion can be found in the Supplementary data.
Supplementary data associated with this article can
be found, in the online version, at doi:10.1016/j.tetlet.
2005.04.091.
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gate Techniques; Academic: San Diego, 1996, pp 169–181.