Colloid Polym Sci (2014) 292:219–228
DOI 10.1007/s00396-013-3070-2
ORIGINAL CONTRIBUTION
A new type of monodisperse porous, hydrophilic microspheres
with reactive chloroalkyl functionality: synthesis
and derivatization properties
Çiğdem Kip & Bilginur Maraş & Oğulcan Evirgen &
Ali Tuncel
Received: 20 July 2013 / Accepted: 6 September 2013 / Published online: 22 September 2013
# Springer-Verlag Berlin Heidelberg 2013
Abstract A seeded polymerization method based on a new
functional monomer, 3-chloro-2-hydroxypropyl methacrylate
(HPMA-Cl), was proposed for the synthesis of a new type of
monodisperse porous, hydrophilic microspheres with reactive
character. By applying the method, poly(3-chloro-2-
hydroxypropyl methacrylate-co-ethylene dimethacrylate)
(poly(HPMA-Cl-co-EDMA)) microspheres in the range of
4–7 μm, with specific surface areas between 2 and 146 m2/
g, were obtained. The microspheres are hydrophilic in nature
due to the hydroxyl groups and are easily derivatizable due to
the reactive chloropropyl moiety. Ligands in the form of small
molecules carrying hydrophobic alkyl or hydrophilic ion ex-
changer groups were covalently attached onto the micro-
spheres via simple and one-pot reactions via their
chloropropyl functionality. Using the same functionality, click
chemistry and surface-initiated atom transfer radical polymer-
ization were also applied for the generation of triazole ring and
zwitterionic molecular brushes on the microspheres, respec-
tively. Poly(HPMA-Cl-co-EDMA) microspheres seem to be a
promising hydrophilic reactive material particularly for the
synthesis of ion exchanger resins and chromatographic sta-
tionary phases.
Keywords Monodisperse porous beads . Glycidyl
methacrylate . 4-Vinylbenzyl chloride . Ion exchange resin .
Stationary phase
Ç. Kip : B. Maraş : O. Evirgen : A. Tuncel
Chemical Engineering Department, Hacettepe University, Ankara,
Turkey
A. Tuncel (*)
Division of Nanotechnology and Nanomedicine, Hacettepe
University, Ankara, Turkey
e-mail: atuncel@hacettepe.edu.tr
Introduction
Monodisperse macroporous microspheres prepared with reac-
tive monomers attracted significant attention for the synthesis
of ion exchangers and chromatographic stationary phases with
different functional groups. Monodisperse porous micro-
spheres synthesized by using 4-chloromethystyrene (CMS)
are one of the commonly investigated materials. The function-
al groups widely found on ion exchangers and chromato-
graphic stationary phases, like secondary amine, quaternary
ammonium, sulfonic acid, or carboxyl, can be easily generated
on the CMS-based microspheres mostly by single-stage and
simple derivatization reactions using their chloromethyl func-
tionality. A staged shape template suspension polymerization
protocol was developed for the preparation of monodisperse
macroporous poly(4-chloromethylstyrene-co-divinylbenzene)
(poly(CMS-co-DVB)) microspheres by Liang et al. [1].
Monodisperse cross-linked core–shell microspheres in the
form of poly(CMS-co-DVB) copolymer were then prepared
by precipitation polymerization [2]. Nearly monodisperse,
micrometer-sized beads with chloromethyl functionality were
synthesized on the basis of single-step swelling of uniform
polystyrene microbeads [3]. Monodisperse porous
poly(CMS-co-DVB) microspheres carrying different ion ex-
changer groups and chloromethyl functionalized porous
poly(styrene-co-divinylbenzene) microbeads were used as
separation medium in different chromatographic applications
[4–6]. On the other hand, porous microspheres suitable for ion
exchange or chromatographic applications were also prepared
by the copolymerization of divinylbenzene with various func-
tional monomers [7–10]. Monodisperse porous poly(glycidyl
methacrylate-co-ethylene glycol dimethacrylate) micro-
spheres synthesized with staged shape template polymeriza-
tion and poly(glycidyl methacrylate) (poly(GMA))-based po-
rous microspheres obtained with different cross-linking agents
220
have been also used as separation medium in different chro-
matographic applications [11–17].
Till today, considerable research effort was focused on the
preparation of monodisperse porous poly(CMS-co-DVB) mi-
crospheres that are highly hydrophobic in nature. According
to the author’s knowledge, no report is found on the prepara-
tion of hydrophilic, polar forms of monodisperse porous mi-
crospheres with reactive chlorine functionality. For this rea-
son, we selected an acrylate-based monomer, 3-chloro-2-
hydroxypropyl methacrylate (HPMA-Cl), and studied the
preparation of monodisperse porous poly(3-chloro-2-
hydroxypropyl methacrylate-co-ethylene dimethacrylate)
(poly(HPMA-Cl-co-EDMA)) microspheres as a new hydro-
philic (i.e., polar) and reactive support for the preparation of
chromatographic separation or ion exchange media with dif-
ferent functionalities. In this study, a new seeded polymeriza-
tion protocol was proposed for the preparation of the micro-
spheres, and the effects of polymerization conditions on the
average bead size, morphology, and pore properties of the
resulting microspheres were investigated.
Experimental section
Materials
The monomers, HPMA-Cl, glycidyl methacrylate (GMA), and
[2-(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium
hydroxide (MESH), and the cross-linking agent, EDMA, were
obtained from Aldrich Chem. Co., St. Louis, MO, USA and
used without further purification. The diluent, ethylbenzene
(EB), and the solvents, ethanol (EtOH, high-performance liquid
chromatography [HPLC] grade) and tetrahydrofuran (THF,
HPLC grade), were supplied by Aldrich. The stabilizer,
polyvinylpyrrolidone (PVP K-30, M r =40,000) was supplied
from Sigma Chemical Co., St. Louis, MO, USA. Sodium lauryl
sulfate (SLS) and polyvinyl alcohol (PVA, 87–89 % hydrolyzed,
molecular weight=85,000–146,000) were also obtained from
Sigma. The initiator, 2,2′-azobisizobutyronitrile (AIBN), was
obtained from Merck A.G., Darmstadt, Germany and
recrystallized from methanol before use. Benzoyl peroxide
was supplied from Merck and dried in vacuo at 30 °C. The
reagents, octadecylamine (ODA), sodium bisulfite,
polyethylenimine (PEI, M w =25,000 Da), triethylamine (TEA),
copper iodide, tetrabutylammonium iodide (TBAI), bipyridine
(BPy), sodium azide, and propargyl alcohol, were supplied from
Aldrich and used without further purification.
Synthesis and characterization of poly(HPMA-Cl-co-EDMA)
microspheres
Poly(GMA) seed latex 2.1 μm in size was synthesized by
dispersion polymerization performed in a sealed glass
Colloid Polym Sci (2014) 292:219–228
polymerization reactor placed in a temperature-controlled
shaking water bath kept at 70 °C for 24 h [18]. The monodis-
perse porous poly(HPMA-Cl-co-EDMA) microspheres were
obtained by a modified form of the two-step swelling and
polymerization protocol. Typically, the diluent, EB (2.5 mL),
was emulsified in water (60 mL) including SLS (0.15 g) by
sonication for 7 min. The aqueous dispersion containing
poly(GMA) seed microspheres (4 mL, solid content=0.35 g)
was added and the emulsion was stirred at room temperature
for 24 h. Next, the monomer phase containing HPMA-Cl
(2.5 mL), EDMA (2.5 mL), and AIBN (0.12 g) was emulsi-
fied in water (60 mL) including SLS (0.15 g) by sonication for
3 min. The monomer emulsion was then mixed with the
aqueous dispersion of diluent-swollen seed microspheres.
The resulting emulsion was magnetically stirred for 24 h.
The aqueous solution (10 mL) of PVA (0.8 g) was added into
the emulsion and polymerization in the swollen seed micro-
spheres was performed at 80 °C, with 120 cpm shaking rate
for 24 h. The monodisperse porous poly(HPMA-Cl-co-
EDMA) microspheres were obtained as the product. The
isolated microspheres were washed with EtOH several times
and extracted with THF. The microspheres were then exten-
sively washed with EtOH and distilled deionized (DDI) water
and finally dispersed in DDI water. In the polymerizations, the
effects of seed latex/monomer ratio and cross-linking agent/
monomer ratio on the average size and pore properties of
poly(HPMA-Cl-co-EDMA) microspheres were investigated.
The size distribution properties and the surface morpholo-
gy of poly(HPMA-Cl-co-EDMA) microspheres were deter-
mined by scanning electron microscopy (SEM; JEM 1200EX,
JEOL, Akishima, Tokyo, Japan). The pore properties (i.e.,
average pore size, pore size distribution, porosity, pore vol-
ume, and specific surface area) of poly(HPMA-Cl-co-EDMA)
microspheres were determined by inverse size exclusion chro-
matography (ISEC) according to the procedure described
elsewhere [19]. The polystyrene standards with different av-
erage molecular weights (M w =2,000,000, 550,000, 212,400,
114,200, 44,000, 18,700, 3,680, and 770) and toluene (M w =
92) were used in ISEC carried out with THF as the mobile
phase in stainless steel columns 50×7.8 mm i.d. with the
mobile phase flow rate of 1.0 mL/min at 22 °C. The specific
surface area of poly(HPMA-Cl-co-EDMA) microspheres was
determined by surface area and pore size analyzer
(Quantachrome, Nova 2200E, UK) using the Brunauer–
Emmett–Teller equation.
Derivatization of poly(HPMA-Cl-co-EDMA) microspheres
with different ligands
In all derivatization reactions, poly(HPMA-Cl-co-EDMA)
microspheres obtained with the formulation described previ-
ously were used. The following chemical routes were applied
for the derivatization of the microspheres:
Colloid Polym Sci (2014) 292:219–228
1. Octadecylation: Dried poly(HPMA-Cl-co-EDMA) mi-
crospheres (1.0) were reacted with ODA (4.0 g) in 1,4-
dioxane at 80 °C for 24 h at a stirring rate of 400 rpm. The
microspheres were then extensively washed with 1,4-
dioxane and EtOH by successive centrifugation–decanta-
tion. The ODA content of microspheres was determined
by nitrogen analysis.
2. Sulfonic acid attachment: Poly(HPMA-Cl-co-EDMA) mi-
crospheres (1.0) were reacted with NaHSO3 (1.5 g) in water
(20 mL) at 80 °C for 24 h at a stirring rate of 400 rpm. The
microspheres were extensively washed with DDI water by
successive centrifugation–decantation. The sulfonic acid
content of microspheres was determined by sulfur analysis.
3. Quaternization : Dried poly(HPMA-Cl-co-EDMA) mi-
crospheres (1.0) were reacted with TEA (2 ml) in
dimethylformamide (DMF, 10 mL) at 70 °C for 24 h at
a stirring rate of 400 rpm. The microspheres were then
extensively washed with DMF and EtOH by successive
centrifugation–decantation. The TEA content of the mi-
crospheres was determined by nitrogen analysis.
4. Amination with a macromolecular ligand : Dried
poly(HPMA-Cl-co-EDMA) microspheres (1.0) were
reacted with PEI (2.0 mL) in DMF (10 mL) at 70 °C for
24 h at a stirring rate of 400 rpm. The microspheres were
then extensively washed with DMF and EtOH by succes-
sive centrifugation–decantation. The PEI content of the
microspheres was determined by nitrogen analysis.
5. Click chemistry : The formation of triazole ring via click
chemistry on the poly(HPMA-Cl-co-EDMA) microspheres
was studied. For this purpose, dried poly(HPMA_Cl-co-
EDMA) microspheres (2 g) were reacted with NaN3 in
dimethyl sulfoxide (DMSO, 60 mL) containing TBAI
(20 mg) at 80 °C with 250 rpm stirring rate for 24 h. Then,
azide functionality was generated on the microspheres.
Azide-functionalized microspheres were washed with
DMSO three times by centrifugation–decantation. The mi-
crospheres (2 g) were then reacted with propargyl alcohol
(3 g) in acetonitrile (20 mL) containing CuI (40 mg) at
50 °C with 250 rpm stirring rate for 24 h. The microspheres
were extensively washed with acetonitrile and then DDI
water by centrifugation–decantation.
6. Surface-initiated atom transfer radical polymerization
(SI-ATRP ): In a typical SI-ATRP, a freshly prepared Cu-
BPy solution including CuBr (0.20 mmol) and BPy
(0.5 mmol) in water (2 mL) was mixed with the aqueous
solution of MESH (10 mmol, 4 mL) in a Schlenk tube. The
tube was sonicated and degassed with nitrogen for 10 min.
An aqueous dispersion (6 mL) of poly(HPMA-Cl-co-
EDMA) microspheres (1.0 g) was dispersed in the resulting
medium. The reactor was evacuated for 10 min. Polymer-
ization was performed at 80 °C for 6 h in a shaking water
bath. After polymerization, the microspheres were exten-
sively washed with EtOH and DDI water, dried in vacuo at
221
70 °C, and subjected to elemental analysis. The polyionic
ligand content of microspheres after heterogeneous ATRP
(in millimoles MESH per gram dry microspheres) was
calculated based on the elemental analysis of
poly(MESH)-g-poly(HPMA-Cl-co-EDMA) microspheres.
Results and discussion
Many applications using monodisperse porous polymer micro-
spheres like ion exchange processes or HPLC applications
involve the presence of various functional ligands on the micro-
spheres. So, the starting microspheres should have a reactive
group and the desired ion exchanger or chromatographic ligand
should be linked onto the microspheres using this reactive
moiety. Poly(CMS-co-DVB) and poly(GMA-co-EDMA) mi-
crospheres have been commonly utilized as starting material
for different chromatographic and ion exchange applications
[4–17]. In our case, monodisperse porous poly(HPMA-Cl-co-
EDMA) microspheres synthesized as an alternative reactive
starting material have 3-chloro-2-hydroxypropyl functionality
coming from the main monomer used in the synthesis (i.e.,
HPMA-Cl). The reactive character of the 3-chloro-2-
hydroxypropyl group allows the chemical derivatization of these
microspheres via simple and single-stage reactions. The molec-
ular structures of the main monomer and cross-linking agent are
given in Fig. 1. On the other hand, the HPMA-Cl unit also has
hydroxyl functionality, making the bead structure hydrophilic.
The common similar material with chloromethyl functionality
(i.e., monodisperse porous poly(CMS-co-DVB) microspheres)
containing aromatic rings in their structure is hydrophobic in
nature with respect to the material synthesized in our study. The
hydrophilic character allows the use of poly(HPMA-Cl-co-
EDMA) microspheres as a starting material for the synthesis
of stationary media for different chromatographic applications
like ion exchange, hydrophilic interaction, and reversed-phase
chromatography or various ion exchange processes.
The derivatization routes followed for obtaining monodis-
perse porous microspheres with different functional groups
Fig. 1 The molecular structures of a the main monomer, HPMA-Cl, and
b the cross-linking agent, EDMA
222
are shown in Fig. 2. The functional group contents and the
specific surface areas of the resulting microspheres are given
in Table 1. In reactions A, B, and C shown in Fig. 2, hydro-
phobic octadecyl functionality, strong cation exchanger sul-
fonic acid group, and strong anion exchanger quaternary
ammonium group were introduced into the microspheres via
one-pot and simple reactions. As seen in Table 1, the ligand
content was found as approximately 10 % (w/w) of the final
microspheres for these three ones. However, the octadecyl
group content of the resulting microspheres defined in terms
of “millimoles ligand per gram dry microspheres” was slightly
lower with respect to the other ligands (i.e., sulfonic acid and
quaternary ammonium). Note that ODA-attached poly(HPMA-
Cl-co-EDMA) microspheres contain hydrophobic alkyl chains
(C-18) on the hydrophilic porous core. Hence, this form can be
evaluated as a potential stationary phase in reversed-phase
chromatography [20]. On the other hand, sulfonic acid and
quaternary ammonium-functionalized poly(HPMA-Cl-co-
EDMA) microspheres are also potential hydrophilic stationary
phases for ion exchange chromatography and various ion ex-
change applications [21]. The specific surface area values
determined for the ligand-attached microspheres were slightly
lower with respect to the plain poly(HPMA-Cl-co-EDMA)
microspheres (Table 1). The suitability of poly(HPMA-Cl-co-
EDMA) microspheres for click chemistry was also tested by
the formation of the triazole ring (i.e., reaction D in Fig. 2). The
specific surface area and ligand content values of triazole-
carrying microspheres were similar to those found for the other
ligands (Table 1).
Fig. 2 Some sample
derivatization routes for the
covalent attachment of ligands in
the form of single small
molecules onto the poly(HPMA-
Cl-co-EDMA) microspheres.
A Octadecylation with ODA,
B generation of strong cation
exchanger –SO3H groups,
C generation of strong anion
exchanger groups with TEA,
D triazole ring formation by click
chemistry (azide formation and
azide–alkine coupling)
Colloid Polym Sci (2014) 292:219–228
The chemical routes followed for the attachment of mac-
romolecular ligands onto the poly(HPMA-Cl-co-EDMA) mi-
crospheres are shown in Fig. 3. A polycationic ligand con-
taining a large number of ionizable groups (i.e., PEI) was
attached onto the microspheres via one-pot reaction between
amine and 3-chloro-2-hydroxypropyl groups (reaction A in
Fig. 3). The suitability of poly(HPMA-Cl-co-EDMA) micro-
spheres for the generation of molecular brushes via SI-ATRP
was tested (reaction B in Fig. 3). The presence of 3-chloro-2-
hydroxypropyl functionality on the poly(HPMA-Cl-co-
EDMA) microspheres allowed the direct use of these micro-
spheres to initiate the SI-ATRP process. Then, zwitterionic
poly(MESH) brushes were generated on the microspheres
using their 3-chloro-2-hydroxypropyl moiety as the initiator.
The order of magnitude of the macromolecular ligand con-
tents of the resulting microspheres was similar to those found
for the small molecules attached (Table 1). The specific sur-
face areas determined with the macromolecular ligands were
also lower with respect to the plain microspheres.
Note that monodisperse porous poly(HPMA-Cl-co-
EDMA) microspheres functionalized with triazole ring, PEI,
or zwitterionic molecular brushes represent a hydrophilic
porous core functionalized with polar, hydrophilic ligand that
can be potentially evaluated as a stationary phase in hydro-
philic interaction chromatography applications [22].
Similar derivatizations were also performed using mono-
disperse porous poly(CMS-co-DVB) microspheres synthe-
sized by different researchers [4–6]. However, poly(CMS-
co-DVB) microspheres were highly hydrophobic in nature.
Colloid Polym Sci (2014) 292:219–228 223

Table 1 The specific surface area and ligand content values of functionalized poly(HPMA-Cl-co-EDMA) microspheres

Ligand type Heteroatom type Percent of Ligand contenta (mmol ligand/g SSA (m2/g)
heteroatom (w/w) dry microspheres)

None – – – 60.0
Octadecyl N 0.47 0.34 (90.4) 53.0
–SO3H S 3.78 1.18 (95.7) 58.8
Quaternary ammonium N 0.99 0.71 (71.7) 55.0
Triazole ring N 3.68 0.88 (73.6) 53.4
PEI N 3.31 – (101.2) 51.1
Poly(MESH) S 0.86 – (75.1) 52.5
a
Ligand content was also given in terms of “milligrams ligand per gram dry microspheres” in parenthesis

Fig. 3 The attachment of

macromolecular ligands onto the
plain poly(HPMA-Cl-co-EDMA)
microspheres. A The attachment
of a long-chain cationic
macromolecular ligand (i.e., PEI),
B the attachment of a long-chain
zwitterionic macromolecular
ligand by SI-ATRP
224 Colloid Polym Sci (2014) 292:219–228

Fig. 4 The SEM photographs of

poly(HPMA-Cl-co-EDMA)
microspheres derivatized with
different ligands. Ligand type: a
plain microspheres, b TEA, c
triazole ring, and d PEI

Fig. 5 SEM photographs of poly(HPMA-Cl-co-EDMA) microspheres 2.5 mL; SDS, 0.25 % (w/w) both in diluent and monomer swelling
obtained with different monomer/seed latex ratios. Monomer/seed latex stages; PVA, 0.8 g; diluent and monomer swelling, 22 °C, 24 h, and
ratio (in milliliters per gram): a 5/0.15, b 5/0.20, c 5/0.30, d 5/0.35, e 5/ 250 rpm stirring rate; polymerization, 80 °C, 24 h, and 120 cpm shaking
0.50, f 5/0.70. Conditions: HPMA-Cl, 2.5 mL; EDMA, 2.5 mL; EB, rate
Colloid Polym Sci (2014) 292:219–228 225

Owing to the their polyacrylate-based structure and hydroxyl
group content, poly(HPMA-Cl-co-EDMA) microspheres are
a new, alternative hydrophilic starting material for the synthe-
sis of chromatographic stationary phases, particularly for
reversed-phase, ion exchange, and hydrophilic interaction
chromatography modes.
The SEM photographs of functional microspheres obtained
from the plain poly(HPMA-Cl-co-EDMA) microspheres
using the chemical routes shown in Figs. 2 and 3 are exem-
plified in Fig. 4. As seen here, no significant change was
observed in the surface morphology by the attachment of
ligands either in the form of a small molecule (i.e., TEA or
triazole) or a macromolecule (i.e., PEI).
In the present study, plain poly(HPMA-Cl-co-EDMA) mi-
crospheres were synthesized with different size and pore
properties by changing the preparation conditions in the
staged shape template polymerization. First, the monomer/
seed latex ratio was changed in the range of 5.5–33.3 mL
monomer/g seed latex. The SEM photographs of
poly(HPMA-Cl-co-EDMA) microspheres obtained with dif-
ferent monomer/seed latex ratios are given in Fig. 5. The
experimental conditions and the size and pore properties of
the poly(HPMA-Cl-co-EDMA) microspheres are given in the
legend of Fig. 5 and Table 2, respectively. Monodisperse
poly(HPMA-Cl-co-EDMA) microspheres in the size range
of 3.8–6.8 μm were obtained by changing the monomer/
seed latex ratio. To increase the monomer/seed latex ratio,
the amount of seed latex was decreased by keeping the total
volume of the monomer phase constant. In this case, a large
amount of monomer should be utilized by each seed particle
during the swelling and polymerization processes. This leads
to an increase in average size as also observed in similar
studies performed with other monomers [23, 24]. No signifi-
cant change in the size distribution (i.e., CV values) was
observed in the studied range of the monomer/seed latex ratio.
Table 2 The size and pore properties of poly(HPMA-Cl-co-EDMA)
microspheres obtained with different monomer/seed latex ratios
The pore size distribution curves of poly(HPMA-Cl-co-
EDMA) microspheres obtained with different monomer/seed
latex ratios are given in Fig. 6. As seen here, most of the pores
were collected in the size range of 1–100 nm for the micro-
spheres obtained with different monomer/seed latex ratios.
The median pore size increased with decreasing monomer/
seed latex ratio (i.e., increasing amount of seed latex) (Table 2).
This finding was also supported by the presence of larger
pores clearly observed on the surface of the poly(HPMA-Cl-
co-EDMA) microspheres synthesized with lower monomer/
seed latex ratios (i.e., Fig. 5e, f). Note that a clear decrease was
also observed in the specific surface area with decreasing
monomer/seed latex ratio (i.e., increasing amount of seed
latex) (Table 2). The increase in the median pore size should
be the most important factor providing lower surface area. The
findings obtained for the effect of monomer/seed latex ratio on
the pore properties of polymer microspheres are consistent
with the results reported for the monodisperse porous micro-
spheres synthesized with different functional monomers [23,
24]. The viscosity of the porogen solution increased with
decreasing monomer/seed latex ratio due to its increasing
linear polymer content. According to the formation mecha-
nism of porous microspheres, excessive aggregation of larger
cross-linked nuclei forming with higher linear polymer con-
centration within the forming microspheres leads to the for-
mation of porous structure with larger pores and lower surface
area [25].
The variation of back pressure with the flow rate of the
mobile phase for the stainless steel columns packed with
poly(HPMA-Cl-co-EDMA) microspheres obtained with dif-
ferent monomer/seed latex ratios is given in Fig. 7. As seen
here, the back pressure exhibited a linear change with the flow
rate for the columns packed with all microspheres. The back
pressure at constant flow rate clearly increased with decreas-
ing monomer/seed latex ratio. As known, the back pressure is
inversely proportional to the column permeability depending
on both intra-particular and inter-particular porosities. The

M/SL (mL/g) Dp (μm) CV (%) d m (nm) ε (%) SSA (m2/g)

5.5 (5.0/0.90) 3.8 4.7 nd nd 13.5

7.1 (5.0/0.70) 4.3 4.0 108 29.4 21.1
10.0 (5.0/0.50) 4.3 4.9 4.82a 30.2 34.7
14.2 (5.0/0.35) 4.6 3.8 nd 33.8 47.8
16.7 (5.0/0.30) 5.5 3.3 14 45.9 60.0
20.0 (5.0/0.25) 5.1 4.2 nd 34.9 62.3
25.0 (5.0/0.20) 5.7 4.6 6 39.7 67.4
33.3 (5.0/0.15) 6.8 5.5 6 30.1 70.7

M/SL monomer/seed latex ratio, Dp mean bead size, CV coefficient of

variation for size distribution, d m median pore size, ε bead porosity, SSA
specific surface area Fig. 6 The pore size distribution curves of poly(HPMA-Cl-co-EDMA)
a
Bidisperse pore size distribution with two maxima microspheres obtained with different monomer/seed latex ratios
226
Fig. 7 The variation of column back pressure with the flow rate of the
mobile phase for the columns packed with poly(HPMA-Cl-co-EDMA)
microspheres obtained with different monomer/seed latex ratios. Column,
50×7.8 mm; mobile phase, distilled water
decrease in the average size of the microspheres involves a
decrease in inter-particular porosity and then an increase in
back pressure. On the other hand, lower bead porosity values
were obtained with the lower monomer/seed latex ratios (Ta-
ble 2). Owing to decreasing inter-particular and intra-
particular porosities, back pressure increases with decreasing
monomer/seed latex ratio (Fig. 7).
A set of polymerization runs was performed by changing the
HPMA-Cl/EDMA volume ratio in the monomer phase. The
pore size distribution curves of poly(HPMA-Cl-co-EDMA)
microspheres obtained with different HPMA-Cl/EDMA ratios
are given in Fig. 8. The size and pore properties of
poly(HPMA-Cl-co-EDMA) microspheres synthesized with
different HPMA-Cl/EDMA volume ratios are given in Table 3.
As seen in Table 3, no significant change was observed in the
average bead size with HPMA-Cl/EDMA ratio. The micro-
spheres approximately 5–6 μm in size and with narrow size
distribution were obtained with all HPMA-Cl/EDMA ratios.
Note that the total volume of the monomer phase was kept
constant in this set. Hence, no appreciable change was observed
Fig. 8 The pore size distribution curves of poly(HPMA-Cl-co-EDMA)
microspheres obtained with different HPMA-Cl/EDMA ratios
Colloid Polym Sci (2014) 292:219–228
Table 3 The size and pore properties of poly(HPMA-Cl-co-EDMA)
microspheres obtained with different HPMA-Cl/EDMA ratios
HPMA-Cl/EDMA Dp (μm) CV (%) d m (nm) ε (%) SSA (m2/g)
(mL/mL)
0.75/4.25 5.14 4.58 9 49.7 146.3
1.50/3.50 4.80 4.92 9 40.5 65.2
2.50/2.50 5.50 3.30 14 45.9 60.0
3.50/1.50 5.40 4.51 106 29.4 14.7
4.25/0.75 4.95 4.32 nd nd 2.14
Conditions: seed latex, 0.3 g; HPMA-Cl+EDMA, 5 mL; EB, 2.5 mL;
SDS, 0.25 % w/w both in diluent and monomer swelling stages; PVA,
0.8 g; diluent and monomer swelling, 22 °C, 24 h, and 250 rpm stirring
rate; polymerization, 80 °C, 24 h, and 120 cpm shaking rate
HPMA-Cl/EDMA volume ratio of HPMA-Cl and EDMA, Dp mean bead
size, CV coefficient of variation for size distribution, d m median pore
size, ε porosity, SSA specific surface area
in the particle size distribution. As seen in Table 3, the specific
surface area markedly increased with decreasing HPMA-Cl/
EDMA ratio (i.e., increasing cross-linking agent (EDMA) con-
centration). Poly(HPMA-Cl-co-EDMA) microspheres with a
specific surface area higher than 100 m2/g were obtained with
the highest cross-linking agent concentration (i.e., with the
HPMA-Cl/EDMA ratio of 0.75/4.25 mL/mL). As seen in
Fig. 8, the volume fraction of larger pores was reasonably
higher with the lowest cross-linking agent concentration (i.e.,
with the HPMA-Cl/EDMA ratio of 3.5/1.5 mL/mL). In this
case, the median pore size was larger, and the specific surface
area was lower with respect to the microspheres obtained with
the higher cross-linking agent concentrations.
The findings for the effect of monomer/cross-linking agent
ratio on the pore properties of polymer microspheres can be
explained by the pore formation mechanism proposed for the
monodisperse microspheres by Cheng et al. [25]. With in-
creasing concentration of cross-linking agent in the monomer
Fig. 9 The variation of column back pressure with the flow rate of the
mobile phase for the columns packed with poly(HPMA-Cl-co-EDMA)
microspheres obtained with different monomer phase compositions. Col-
umn, 50×7.8 mm; mobile phase, distilled water
Colloid Polym Sci (2014) 292:219–228
phase, a higher number of smaller nuclei should be produced
by the phase separation within the forming microspheres. A
structure with smaller pores dominantly contributing to the
specific surface area forms by the aggregation and fixation of
smaller nuclei within the forming microspheres.
The variation of back pressure with the flow rate in the
column packed with the poly(HPMA-Cl-co-EDMA) micro-
spheres obtained with different HPMA-Cl/EDMA ratios is
given in Fig. 9. As seen here, reasonably higher back pres-
sures were observed particularly with the microspheres
obtained with the low cross-linking agent concentrations
(i.e., with the HPMA-Cl/EDMA ratios of 3.50/1.50 and
4.25/0.75 mL/mL). Note that the ISEC study could not be
performed with the sample obtained with the HPMA-Cl/
EDMA ratio of 4.25/1.50 mL/mL since the column pressure
was so high. However, extremely high back pressures ob-
served with the microspheres synthesized with the HPMA-
Cl/EDMA ratios of 3.5/1.50 and 4.25/0.75 mL/mL should be
explained by the low porosity of these materials as also
supported by very low specific surface area values (Table 3).
Conclusion
Monodisperse porous, hydrophilic polyacrylate microspheres
with reactive character were synthesized by starting from a
new main monomer, HPMA-Cl. The size and pore properties
of microspheres were effectively controlled by changing the
polymerization conditions. The presence of 3-chloro-2-
hydroxypropyl functionality on the microspheres allowed
the synthesis of supports functionalized with different ligands
via one-pot and simple derivatization protocols. The ligands in
the form of single small molecules or in the form of macro-
molecules were covalently attached on the microspheres. It is
believed that monodisperse porous poly(HPMA-Cl-co-
EDMA) microspheres are a promising starting material for
the synthesis of stationary phases for ion exchange and chro-
matography applications.
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