Synthesis of polylactide with thiol end groups

Article abstract of DOI:10.1135/cccc20031131

Four synthetic routes to poly(L-lactide) with thiol end groups based on ring-opening polymerization of L-lactide (LA) catalysed with tin(II) 2-ethylhexanoate (Sn(Oct)₂) are reported. The following alcohols were used as co-initiators of polymerization: 2-sulfanylethan-1-ol, 2-[(2,4-dinitrophenyl)sulfanyl]ethan-1-ol, 2-(tritylsulfanyl)ethan-1-ol and allyl alcohol. End groups introduced into polymers by co-initiators were transformed to thiol groups by a subsequent modification reaction. The efficiencies of the used synthetic methods were evaluated and discussed. The best results were obtained with co-initiator 2-(tritylsulfanyl)ethan-1-ol.

Full text of DOI:10.1135/cccc20031131

Synthesis of Polylactide
1131
SYNTHESIS OF POLYLACTIDE WITH THIOL END GROUPS
a2,b
Štěpán POPELKAa1,b,* and František RYPÁČEK
a
Institute of Macromolecular Chemistry, Academy of Sciences of the Czech Republic,
Heyrovského nám. 2, CZ 162 06 Prague 6, Czech Republic; e-mail: popelka@imc.cas.cz,
1
2
ryp@imc.cas.cz
b
Center for Cell Therapy and Tissue Repair, Charles University, 2nd Faculty of Medicine,
Institute of Neurosciences, V Úvalu 84, 150 18, Prague 5, Czech Republic
Received April 4, 2002
Accepted March 3, 2003
Four syn th etic routes to poly(L-lactide) with th iol en d groups based on rin g-open in g poly-
m erization of L-lactide (LA) catalysed with tin (II) 2-eth ylh exan oate (Sn (Oct)2) are reported.
Th e followin g alcoh ols were used as co-in itiators of polym erization : 2-sulfan yleth an -1-ol,
2-[(2,4-din itroph en yl)sulfan yl]eth an -1-ol, 2-(tritylsulfan yl)eth an -1-ol an d allyl alcoh ol. En d
groups in troduced in to polym ers by co-in itiators were tran sform ed to th iol groups by a sub-
sequen t m odification reaction . Th e efficien cies of th e used syn th etic m eth ods were evalu-
ated an d discussed. Th e best results were obtain ed with co-in itiator 2-(tritylsulfan yl)eth an -1-ol.
Keyw ord s: Polylactides; Th iol-fun ction alized polym ers; En d group m odification ; Th iols;
Rin g-open in g polym erization s; Adsorption on gold.
Polylactide (PLA) is widely used as a biom aterial. For application such as
tissue en gin eerin g, kn owledge of in teraction of PLA surfaces with com po-
n en ts of biological fluids is very im portan t. Th e surface plasm on reson an ce
(SPR) tech n ique1 is an efficien t m eth od for studies of protein adsorption
on surfaces exposed to protein solution s or com plex biological fluids, such
as blood plasm a or lym ph . Th e SPR tech n ique uses gold-plated optical
substrate, th e gold surface of wh ich h as to be first coated with a very th in
layer of th e polym er un der study. Usin g SPR, th e ch an ges at th e polym er
surface due to adsorption of m olecules from solution can be followed in
real tim e an d th e dyn am ics of adsorption process can be evaluated in de-
pen den ce on th e m aterial properties. To use SPR for evaluation of PLA-
based m aterials, a con tiguous th in layer of PLA well-adh eren t to th e gold
surface h as to be prepared.
To prepare a PLA layer stron gly attach ed to th e gold surface we decided to
use th iol-m on ofun ction alized PLA. Low-m olecular-weigh t th iols are well
kn own to form h igh ly regular an d stable self-assem bled m on olayers on
Collect. Czech. Chem. Commun. (Vol. 68) (2003)
doi:10.1135/cccc20031131

1132
Popelka, Rypáček:
gold2 due to th e form ation of th iolate bon ds to th e m etal surface. Ph ysico-
ch em ical properties of th e m on olayer depen d on th e structure of th e rest of
th e th iol m olecule. Th e preparation of a th in polym er film on gold from
polystyren e with th iol en d groups h as been described in literature3. In th e
sam e way th iol-fun ction alized polylactides sh ould form eith er a con tiguous
coatin g or, at least, an an ch orin g layer on gold substrates, on to wh ich an
ultrath in film of PLA can be deposited by spin castin g.
Poly(lactic acid) as well as oth er aliph atic poly(h ydroxy acids) are m ost
com m on ly prepared by rin g-open in g polym erization of cyclic esters of
th ese acids. Tin (II) 2-eth ylh exan oate (Sn (Oct)2) as catalyst an d an alcoh ol
as co-in itiator are th e often used in itiatin g system of th ese polym erization s.
Pen czek’s group4,5 th orough ly in vestigated th e m ech an ism an d kin etics of
th e polym erization an d proven th e process to be “livin g”. Th e m olecular
weigh t of polym er products can be varied in a wide ran ge by adjustin g th e
in itial m on om er-to-ROH m olar ratio. Th e kin etic data an d MALDI-TOF
m ass spectra in dicate6 th at th e actual in itiator of th e polym erization is
tin (II) alkoxide (Oct)Sn OR form ed from Sn (Oct)2 an d th e co-in itiatin g alco-
h ol ROH in th e reaction : Sn (Oct)2 + ROH ↔ (Oct)Sn –OR + OctOH. Th e
propagation proceeds by a m on om er in sertion in to th e Sn –OR bon d of th e
tin (II)-alkoxide active cen tres reversibly form ed from polyester h ydroxy en d
groups an d Sn (Oct)2. Sin ce th e co-in itiatin g com poun d ROH is in corpo-
rated in to th e polym er as th e oth er ch ain en d group, th e polym erization
can be efficien tly used for th e preparation of fun ction alized polylactides
an d block copolym ers with polylactide blocks7,8
.
So far very few papers on th iol-fun ction alized aliph atic poly(h ydroxy
acid)s h ave been publish ed. Recen tly, th e preparation of poly(ε-capro-
lacton e) (i.e. poly(h exan o-6-lacton e)) with th iol groups h as been described
usin g an alcoh ol with a 2,4-din itroph en yl-protected th iol groups as
co-in itiator9. Th e th iol groups were recovered after cleavin g off th e
2,4-din itroph en yl groups by excess of a low-m olecular-weigh t th iol.
In th is paper we test an d com pare four syn th etic routes to th iol-
fun ction alized polylactide. Th ree of th em in troduce th iol groups th rough a
co-in itiator (fun ction alization by in itiatiator). As co-in itiator we tested
2-sulfan yleth an -1-ol with th e th iol groups protected by 2,4-din itroph en yl
an d trityl groups an d also with free th iol groups to ch eck th e n ecessity of
protection . Th e fourth route tested m akes use of a polym er precursor with
allyl en d groups wh ich are subsequen tly m odified by a sulfur reagen t (poly-
m er fun ction alization ). Th e suitability of each route for th e preparation of
well-defin ed polym er th iols is evaluated on th e basis of polym er param eters
in particular th e degree of fun ction alization .
Collect. Czech. Chem. Commun. (Vol. 68) (2003)

Synthesis of Polylactide
1133
EXPERIMENTAL
Materials
1,4-Dioxan e for polym erization s was dried by refluxin g over sodium . 2-Sulfan yleth an -1-ol
(Fluka AG) (co-in itiator 1) was distilled an d dried over m olecular sieve. Allyl alcoh ol (co-
in itiator 2) was distilled from CaH2. L-Lactide was syn th esized10 from L-lactic acid an d re-
crystallized from a m ixture of dry toluen e an d eth yl acetate (1:1 v/v) prior to use. 2,2′-Azo-
bisisobutyron itrile (AIBN) (Aldrich ) was recrystallized from m eth an ol. N-(2,4-Din itroan ilin o)-
m aleim ide (DNAMI) (Sigm a) was used as received. Tin (II) 2-eth ylh exan oate (Fluka AG) was
purified by vacuum distillation . Trifluoroacetic acid (TFA) an d trieth ylam in e (TEA) were dis-
tilled prior to use. All oth er reagen ts were purch ased from Aldrich an d used as received.
Measurem en ts
NMR spectra were recorded on a Bruker Avan ce DPX-300 spectrom eter in CDCl3 with
tetram eth ylsilan e as an in tern al stan dard. Calculation of th e n um ber-average m olecular
weigh t is based on th e ratio of in tegrated sign als of m eth in e proton s of th e COCH(CH3)OH
en d group (4.29 ppm ) an d of th e COCH(CH3)O m ain -ch ain un its (5.09 ppm ) in th e 1H NMR
spectra. Size exclusion ch rom atograph y (SEC) was perform ed on a Waters HPLC-SEC m odu-
lar system usin g PLgel 103 Å, 10 µm (7.5 × 600 m m ) colum n , eluen t THF an d Waters 410 RI
an d Waters 484 UV detectors. Th e colum n s were calibrated with polystyren e stan dards an d
th e m olecular-weigh t/elution volum e depen den ce was recalculated for PLLA by usin g
Mark–Houwin k coefficien ts for polystyren e an d PLLA 11
.
Determ in ation of Th iol Groups
Two parallel m eth ods were used: (i) By NMR spectroscopy, th e con ten t of free th iol en d
groups was determ in ed from ratio of in ten sities of m eth ylen e (2.68 dt, 2 H, HSCH2CH2O or
4.17 t, 2 H, HSCH2CH2O) an d m eth in e en d groups (4.29 q, 1 H, COCH(CH3)OH) in
1H NMR spectrum . (ii) By th e reaction of th iol groups with N-(2,4-din itroan ilin o)m ale-
im ide12. A polym er sam ple an d th ree equivalen ts of DNAMI were dissolved in DMF an d al-
lowed to react at room tem perature for 5 h (Sch em e 1). To separate th e labeled polym er
fraction from an excess of un reacted DNAMI an d to quan tify it, SEC with UV detection (λ =
323 n m ) was used. Th e m olar am oun t of th e tran sform ed th iol groups was calculated from
th e peak area of th e labeled polym er. By com parison of th e m olar am oun t of th e in jected
polym er an d th e m easured m olar am oun t of th iol groups, th e fraction of th iol-fun ction -
alized PLA was calculated.
R
O
O
S
O2N
O2N
HN
N
+
HS
R
O2N
O2N
HN
N
O
O
SCHEME 1
Collect. Czech. Chem. Commun. (Vol. 68) (2003)

1134
Popelka, Rypáček:
2-[(2,4-Din itroph en yl)sulfan yl]eth an -1-ol (3)
Adoptin g th e procedure described in literature9, a solution of 2-sulfan yleth an -1-ol (30 m m ol,
2.1 m l) in 30 m l of CHCl3 was slowly added in to a m ixture of 1-fluoro-2,4-din itroben zen e
(5.6 g, 30 m m ol) an d TEA (7 m l). Th e brown ish reaction m ixture was stirred at room tem -
perature overn igh t. Th e reaction m ixture was diluted with CHCl3 (200 m l), extracted with 1 M
HCl (1 × 20 m l) an d water (2 × 20 m l). Th e ch loroform ph ase was separated, dried over an -
h ydrous MgSO4 an d filtered. Th e yellow product was twice recrystallized from CHCl3. Yield
4.5 g (61%); m .p.: 99.5–101 °C. For C8H8N2O5S (244.2) calculated: 39.34% C, 3.30% H,
11.47% N, 13.13% S; foun d: 39.40% C, 3.20% H, 11.30% N, 13.06% S. 1H NMR (CDCl3):
8.95 s, 1 H, J(3,5) = 2.5 (H-3), H-arom .; 8.28 dd, 1 H, J(3,5) = 2.5, J(5,6) = 9.0 (H-5), H-arom .;
7.62 d, 1 H, J(5,6) = 9.0 (H-6), H-arom .; 3.97 t, 2 H, J = 5.9, SCH2CH2OH; 3.24 t, 2 H, J =
5.9, SCH2CH2OH.
2-(Tritylsulfan yl)eth an -1-ol (4)
A procedure described in literature was followed13. Trityl ch loride (4 g, 14 m m ol) was dis-
solved in 60 m l of petroleum eth er an d 1 m l (14 m m ol) of 2-sulfan yleth an -1-ol was added.
Th e m ixture was refluxed for 15 m in . Wh ite crystals th at precipitated in reaction m ixture
were collected by filtration an d th e product was crystallized from eth an ol. Fin al
recrystallization was from a m ixture of toluen e an d petroleum eth er (1:1 v/v). Yield 3.2 g
(71%); m .p.: 115–116 °C. For C21H20OS (320.5) calculated: 78.71% C, 6.29% H, 10.01% S;
foun d: 78.6% C, 6.3% H, 9.9% S. 1H NMR (CDCl3): 7.14–7.38 m , 15 H, H-arom .; 3.31 t, 2 H,
J = 6.0, SCH2CH2OH; 2.41 t, 2 H, J = 6.0, SCH2CH2OH.
Polym ers
Polym ers were prepared by rin g-open in g polym erization of L-lactide in dioxan e in itiated
with tin (II) 2-eth ylh exan oate an d co-in itiated with on e of th e four differen t co-in itiators
possessin g OH groups (Sch em e 2). Th e m on om er-to-Sn (Oct)2 m olar ratio was 50:1 except th e
case of polym erization with co-in itiator 1 wh en th e 15:1 ratio was used. Th e m on om er-to-
co-in itiator ratio was varied as sh own in Table I. Th e polym erization was carried out un der
n itrogen atm osph ere in sealed am poules at 60 °C for 40 h . Th e reaction m ixture was fin ally
poured in to m eth an ol an d th e precipitated polym er was isolated by filtration or cen tri-
fugation .
Modification of En d Groups
Isolated polym er precursors 1a–4a were subjected to th e followin g treatm en t, wh ich con -
verted th e in itiator en d groups in to th iol groups.
To regen erate th iol en d groups from precursor 1a, an excess of low-m olecular-weigh t th iol
was used. An am oun t of 1.5 g of polym er 1a was dissolved in dich lorom eth an e (16 m l) an d
4 m l of 2-sulfan yleth an -1-ol was dropwise added to th e solution . Upon stirrin g for 30 m in ,
dark-red precipitation of low-m olecular-weigh t tin (II) th iolate was form ed. It was rem oved
by filtration an d th e polym er solution was precipitated in to m eth an ol.
Allyl en d groups of th e polym er 2a were m odified by radical addition of triph en yl-
silan eth iol to th e double bon d usin g th e procedure described for low-m olecular-weigh t
alken es14. Polym er 2a, correspon din g to 146 µm ol of ch ain en ds, triph en ylsilan eth iol (121 m g,
Collect. Czech. Chem. Commun. (Vol. 68) (2003)

Synthesis of Polylactide
1135
CH3
O
HS
OH
O
H
H
(Oct)SnS
O
O
HSCH2CH2OH
n
1
1a
CH3
O
O
O
1. Ph3SiSH/AIBN
2.TFA
OH
CH3
O
O
n
O
2
3
CH3
CH3
2a
O
O
H
HS
Sn(II) 2-ethylhexanoate
O
n
CH3
O
5
S
O2N
O
H
OH
HSCH2CH2OH
TEA
O2N
S
O
O
NO2
n
NO2
3a
TFA
CH3
O
Ph
Ph
Ph
Ph
OH
Ph
S
Ph
O
H
S
n
4
4a
SCHEME 2
414 µm ol) an d AIBN (17 m g, 104 µm ol) were dissolved in 1.3 m l of ben zen e, h eated to 95 °C
an d stirred for 5 h . Th e reaction m ixture con tain in g th e polym er-th iol adduct was cooled
an d treated with 5 equivalen ts of TFA an d stirred for 0.5 h to rem ove th e triph en ylsilyl pro-
tectin g groups. Th e reaction m ixture was precipitated in to m eth an ol.
2,4-Din itroph en yl protectin g groups of polym er 3a were cleaved with an excess of
2-sulfan yleth an -1-ol. An am oun t of 1.5 g of polym er 3a was dissolved in 15 m l of m ixture
CHCl3/2-sulfan yleth an -1-ol (2:1 v/v) con tain in g TEA in 1% (w/w) con cen tration . Th e solu-
tion was stirred at room tem perature for 15 h an d th en precipitated in to m eth an ol.
Th e trityl protectin g groups of polym er 4a were rem oved by trifluoroacetic acid. An
am oun t of 1.5 g of polym er 4a was dissolved in 10 m l of TFA. Yellow colour of th e
triph en ylm eth yl cation im m ediately appeared. After 1 h of stirrin g at room tem perature, th e
solution was precipitated in to m eth an ol.
RESULTS AND DISCUSSION
Th e th iol en d groups con ten t is a key param eter for com parison of th iol-
fun ction alized polym ers prepared by differen t routes. Th e determ in ation
based on th e in ten sity of peaks in 1H NMR spectrum (2.68 dt, 2 H,
HSCH2CH2O an d 4.17 t, 2 H, HSCH2CH2O) is n ot quite reliable due to a
partial overlap of th ese sign als with th ose of th e oth er en d groups of PLA
ch ain (4.29 q, 1 H, COCH(CH3)OH an d 2.62 s, 1 H, COCH(CH3)OH).
Th erefore, we h ave developed an altern ative an alytical m eth od derived
Collect. Czech. Chem. Commun. (Vol. 68) (2003)

1136
Popelka, Rypáček:
from th e m eth od com m on ly used in protein ch em istry for quan tification of
cystein e residues15. It is based on th e SH groups labelin g with a ch rom o-
ph ore an d its subsequen t spectroph otom etric determ in ation . Both m eth ods
gave SH-con ten ts, wh ich are in a reason able agreem en t, th us con firm in g
th at th ey represen t th e actual th iol con ten t. Th ey are given togeth er with
th e oth er param eters of th e prepared polym ers in Table I.
Th e polym erization co-in itiated with 2-sulfan yleth an -1-ol (1) could be
th e sim plest way to obtain polym er th iol with out th e n ecessity of prepara-
tion of a com m ercially un available protected co-in itiator. However, th e for-
m ation of tin (II) th iolate as a product of reaction of Sn (Oct)2 with th e th iol
groups an d, con sequen tly, an in fluen ce on th e course of polym erization is
to be expected. To com pen sate th e loss of catalytic activity of Sn (Oct)2, th e
am oun t of th e catalyst in th e reaction m ixture was in creased up to
equim olar to th e co-in itiator. Tin (II) th iolate apeared as a brown ish turbid-
ity in th e reaction m ixture in th e course of polym erization . Th e in fluen ce
of side reaction between co-in itiator an d catalyst was m arkedly reflected in
a low degree of fun ction alization an d in lower m olecular weigh t of th e
polym ers (polylactides 1-1 an d 1-2) th an expected from th e com position of
th e startin g reaction m ixture. To get a h igh er degree of th iol fun ction -
alization , eith er protection of th e th iol group in 2-sulfan yleth an -1-ol or an -
oth er ch em ical strategy h as to be used.
Accordin g to literature9, 2-[(2,4-din itroph en y)sulfan yl]eth an -1-ol (3) was
used as a protected form of 2-sulfan yleth an -1-ol in th e syn th esis of th iol-
fun ction alized poly(ε-caprolacton e), givin g h igh yields of fun ction alization .
Alth ough our polym erization con dition s were com parable to th ose de-
scribed in literature, we did n ot succeed in obtain in g h igh degree of fun c-
tion alization , as sh own in Table I (polylactides 3-1 an d 3-2). Sim ilar
in dication s of a side reaction between th e catalyst an d co-in itiator (proba-
bly its NO2 groups) were observed to th ose in th e case of th e polym eriza-
tion with co-in itiator 1: low fun ction alization , lower th an th eoretical
weigh t an d a broad MWD. Moreover, th e con dition s used for deprotection
caused partial degradation of PLA, th us decreasin g th e con ten t of th iol en d
groups.
Trityl was tested as an oth er th iol protectin g group for 2-sulfan yleth an -
1-ol. Th e products of polym erization in itiated with 4 (polylactide 4-1 to
4-6) were quan titatively fun ction alized as follows from th e ratio of peak
in tegrals of PLA en d groups: quartet 4.29 ppm in OCH(CH3)OH, triplets
3.80 an d 2.38 ppm in trityl-SCH2CH2O-PLA (Fig. 1). Ch aracteristics of th e
precursor polym ers 4a in dicate th at th e polym erization was con trolled. Af-
ter deprotection th e con ten t of th iol en d groups rem ain ed h igh . Th e
Collect. Czech. Chem. Commun. (Vol. 68) (2003)

Synthesis of Polylactide
1137
Collect. Czech. Chem. Commun. (Vol. 68) (2003)

1138
Popelka, Rypáček:
deprotected polym er, h owever, still con tain s a sm all fraction of th e pro-
tected species detected as triplets 3.80 an d 2.38 ppm (Fig. 2). If th e low-
m olecular-weigh t (Mn = 2500) polym er precursor was deprotected, origin al-
ly un im odal MWD ch an ged to bim odal. Molecular weigh t of th e n ew poly-
m er fraction in th e polym er was twice h igh er th an th e origin al on e. Th is
in dicates partial oxidation of th e polym er th iol to polym er disulfide, wh ich
is a typical reaction of low-m olecular-weigh t th iols but was also observed
with polym er th iols16. Th e triplet 2.86 ppm in 1H NMR spectrum was as-
sign ed to CH2 group in th e substructure OCH2CH2S–SCH2CH2O. Th e oxida-
tion of polym er th iol of h igh er m olecular weigh t th an 2500 was n ot
1
observed. Th e triplet at 3.05 ppm in th e H NMR spectrum (Fig. 2) of de-
protected polym er 4a in dicates th e presen ce of a substructure SCH2CH2O
different from the HSCH2CH2O, trityl-SCH2CH2O and OCH2CH2S–SCH2CH2O
groups. Th is triplet was also observed in 1H NMR spectra of polym ers pre-
pared by deprotection of 1a an d 3a. We assum e th at th is substructure (see
Sch em e 3) m ay origin ate from th e reesterification of polym er ester with free
th iol. Th e estim ated value (CS Ch em Draw Ultra 5.0) of ch em ical sh ift is
3.15 ppm for OCOSCH2CH2O, wh ich correspods to th e experim en tal value.
e
b
d′
a′′
f
d
c
e′
a′
a
a
a′ + a′′ +CHCl
e + e′
d
3
c
b
d′
f
7.0
6.0
5.0
4.0
3.0
2.0
ppm
FIG. 1
1H NMR spectrum of poly(L-lactide) with 2-(tritylsulfan yl)eth oxy en d groups (4-1, before
deprotection )
Collect. Czech. Chem. Commun. (Vol. 68) (2003)

Synthesis of Polylactide
1139
Th e con ten t of th is substructure in th e product n ever exceeded 10% of all
SCH2 structures.
CH3
O
CH3
O
CH3
O
CH3
O
O
H
S
O
H
O
O
O
m
n
SCHEME 3
Fun ction alization of a well-defin ed polym er precursor with a sulfur re-
agen t is an altern ative to fun ction alization by in itiator. Triph en ylsilan e-
th iol an d allyl-term in ated PLA were ch osen for th e fourth syn th etic route,
sin ce triph en ylsilan eth iol sh ould n ot react with m on om er un its in radical
addition of SH on th e term in al allyl double bon d. Alth ough , allyl en d
groups were in troduced with h igh efficien cy in to th e PLA by polym eriza-
tion co-in itiated with allyl alcoh ol (2), th e subsequen t radical addition of
th iol did n ot proceed as efficien tly as described for low-m olecular-weigh t
1
an alogues16. Th e H NMR spectra did n ot in dicate an y sign ifican t m odifica-
tion of th e allyl groups un der th e described con dition s.
a
a′
e
b
d′
a′′
f
d
e′
c
g
h
a′ + a′′ +CHCl
d
h
e + e′
3
f + g
d′
a
c
b
7.0
6.0
5.0
4.0
3.0
2.0
ppm
FIG. 2
1H NMR spectrum of poly(L-lactide) with 2-sulfan yleth oxy en d groups (4-1, after deprotection )
Collect. Czech. Chem. Commun. (Vol. 68) (2003)

1140
Popelka, Rypáček:
CONCLUSIONS
2-(Tritylsulfan yl)eth an -1-ol (4) was foun d to be th e best in itiator out of
th ose tested for preparation of th iol-m on ofun ction alized polylactide. Poly-
m ers prepared in th is way sh ow th iol con ten ts suitable for future adsorp-
tion studies on gold substrates. Molecular param eters of th e fun ction alized
polym ers can be reason ably con trolled by th e reaction con dition s.
The financial support of the Grant Agency of the Czech Republic (grant No. 203/99/0576) is
gratefully acknowledged.
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