d e n t a l m a t e r i a l s 2 2 ( 2 0 0 6 ) 845–851

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journal homepage: www.intl.elsevierhealth.com/journals/dema

Analysis of residual monomers in dendritic methacrylate

copolymers and composites by HPLC and headspace-GC/MS

Eeva K. Viljanen a,∗ , Sarka Langer b , Mikael Skrifvars c , Pekka K. Vallittu a

a University of Turku, Institute of Dentistry, Department of Prosthetic Dentistry and Biomaterials Science,
Lemminkäisenkatu 2, FI-20520 Turku, Finland
b SP Swedish National Testing and Research Institute, Chemistry and Materials Technology, Borås, Sweden
c University College of Borås, School of Engineering, Borås, Sweden

a r t i c l e i n f o a b s t r a c t

Article history: Objectives. The aim of this study was to analyze the residual monomer content of photopoly-
Received 31 May 2005 merized dendritic methacrylate copolymers and particulate filler composites. Headspace-
Received in revised form gas chromatography/mass spectrometry (HS-GC/MS) was compared with high performance
21 September 2005 liquid chromatography (HPLC).
Accepted 24 October 2005 Methods. The resin mixtures consisted of a dendritic methacrylate monomer, methyl
methacrylate and acetoacetoxyethyl methacrylate in varied proportions. In addition, one
of the composites contained 1,4-butanediol dimethacrylate. Camphorquinone and 2-(N,N-
Keywords: dimethylamino)ethyl methacrylate were used as the light-activated initiator system. The
Dendrimers content of residual methyl methacrylate and acetoacetoxyethyl methacrylate after 40 s pho-
Dental polymers topolymerization were analyzed with HPLC and HS-GC/MS.
Residual monomer Results. The content of residual methyl methacrylate decreased and residual acetoace-
High performance liquid toxyethyl methacrylate increased with increasing concentration of acetoacetoxyethyl
chromatography methacrylate in the resin mixture. The results with both methods had the same trend.
Headspace-gas Significance. The addition of acetoacetoxyethyl methacrylate enhanced the copolymerization
chromatography/mass spectrometry of methyl methacrylate, but did not decrease the total residual monomer content. The HS-
GC/MS method was found to be a feasible method in the analysis of low-boiling residuals
in dental polymers.
© 2005 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

1. Introduction of the sample. The pipetting required in the dilution is also a

According to ISO standards, the residual monomers in
most dental polymers are analyzed with a semi-quantitative
method of solubility. The qualitative and quantitative stan-
dard method for the analysis of residual monomers in
denture base polymers is gas chromatographic (GC) or
high performance liquid chromatographic (HPLC) analysis of
methyl methacrylate, extracted with an organic solvent [1].
The method requires laborious and time-consuming sample
preparation with a 3-day extraction and subsequent dilution
possible source of error in the analysis.
For low-volatile compounds, an ideal solvent is gas. In
headspace (HS) sampling, the extraction of analytes from a
solid or a liquid sample is achieved in a closed vial with an
inert gas that carries the vaporized analytes straight into a gas
chromatographic column. Quantitative headspace analysis is
based on an equilibrium between the analyte concentrations
in the sample and the headspace, the gaseous phase in the
vial. The sampling method is simple and convenient and does
not necessitate extensive manipulation of the sample prior to

∗
Corresponding author. Tel.: +358 2 333 51; fax: +358 2 333 8390.
E-mail address: eeva.viljanen@utu.fi (E.K. Viljanen).
0109-5641/$ – see front matter © 2005 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.
doi:10.1016/j.dental.2005.11.012
846 d e n t a l m a t e r i a l s 2 2 ( 2 0 0 6 ) 845–851

analysis. Another advantage of the two-phase system is that
there is no risk of contamination of the column by the non-
volatile sample compounds: they will not enter the chromato-
graph. This is a particular advantage of HS-GC over general
GC. The headspace sampling method has been in use since
the early days of gas chromatography. Analysis of alcohol in
aqueous solutions by the analysis of the vapor was already
indicated in a publication in 1939, the headspace sampling
method has been used since the 1950s and the gas chromato-
graphic analysis of headspace samples was first automated in
1967 [2]. The main drawbacks of HS-GC/MS are the sometimes
long equilibration time and the cost of the instruments.
Headspace sampling has been used in several studies of
residual monomers, for example, of residual MMA in polymer
latex [3] and residual styrene in polystyrene granules [4]. In
dental applications, the method has been utilized in the analy-
sis of substances released from adhesive pastes for prostheses
[5] and composite resin material [6]. Headspace sampling also
has other applications in dentistry and medicine, for example,
analysis of volatile sulfur compounds causing halitosis [7].
This study continues a series of studies on the suitabil-
ity of dendritic monomers as components in dental polymers
[8–10]. The dendritic resin systems previously studied con-
tained methyl methacrylate (MMA) as a low-viscosity diluent
monomer for the dendritic monomers. A diluent monomer
was needed because the viscosity of the dendritic monomer is
high for a manageable composition. MMA was selected as the
original comonomer because of its advantageous properties as
a solvent during the synthesis of the dendrimer molecules [11]
and because it copolymerizes with the dendritic monomer. In
the present study the resins were modified by the use of ace-
toacetoxyethyl methacrylate (AAEM) as a third co-monomer.
The aim was to enhance the copolymerization of methyl
methacrylate with a more reactive monomer and to lower the
viscosity with the purpose of allowing the polymerization to
extend further, resulting in fewer residual monomers. In the
photopolymerization of multifunctional monomers, conver-
sion is limited by the vitrification of the gelled cross-linked
system when the glass transition temperature (Tg ) exceeds the
reaction temperature [12–14]. In addition to the unfilled resins,
two experimental particulate filler composites containing the
dendritic monomer, MMA and AAEM along with an addi-
tional difunctional methacrylate, 1,4-butanediol dimethacry-
late (BDDMA), were investigated.
2. Materials and methods
2.1. Materials
The materials used in this study are presented in Table 1.
The ideal structural formula of the D12 dendrimer and the
formulas of methyl methacrylate (MMA), acetoacetoxyethyl
methacrylate (AAEM) and 1,4-butanediol dimethacrylate
(BDDMA) are presented in Fig. 1. The dendritic monomer D12
was synthesized according to the method by Rånby and Shi
[15]. It was synthesized by adding six moles of trimellitic
anhydride to a core of one mole of dipentaerythritol, and by
esterifying the formed carboxylic acid end groups with glycidyl
methacrylate, and then esterifying the hydroxyl groups of gly-
cidyl methacrylate with acetic anhydride. The product was
isolated by vacuum distillation. 20 wt.% of MMA was added
as a diluent monomer.
The basis of the resins was the prefabricated D12:MMA
(80:20, wt:wt) mixture, of which 0, 4, 8, 12 or 16 wt.%
was replaced with acetoacetoxyethyl methacrylate. Cam-
phorquinone (CQ) and 2-(N,N-dimethylamino)ethyl methacry-
late (DMAEMA) were used as the light-activated initiator
system, both in the quantity of 1.0 wt.%. To prepare the
resins, predetermined amounts of the D12:MMA mixture were
weighed on an analytical balance. The amounts of AAEM, CQ
and DMAEMA needed, were calculated and added to the mix-
tures. The resins were stored at (13 ± 1) ◦ C in light-protected
containers for 20 h to dissolve CQ.
The monomer composition in the particulate filler compos-
ites (PFC) was similar to that used in the unfilled resins. The
monomers in PFC1 consisted of 90 wt.% of the basic mixture
D12:MMA (80:20, wt:wt) and 10 wt.% of AAEM. PFC2 contained
70 wt.% of the basic mixture, 10 wt.% of AAEM and 20 wt.% of
BDDMA, which is a dimethacrylate used in dental polymers.
Both contained the initiator system (CQ and DMAEMA) and
an inhibitor (hydroquinone). The particulate filler contents in
PFC1 and PFC2 were 71.3 and 72.2 wt.%, respectively. The exact
filler compositions and initiator system concentrations of the

Table 1 – Materials used in this study

Code Material Description Manufacturer

AAEM Monomer Acetoacetoxyethyl methacrylate Aldrich, Steinheim, Germany

BDDMA Monomer 1,4-Butanediol dimethacrylate Cray Valley, Villers St Paul, France
CQ Initiator Camphorquinone Fluka, Buchs, Switzerland
D12:MMA Experimental monomer mixture Dendritic methacrylate monomer VTT Chemical Technology, Espoo, Finland
and methyl methacrylate in
weight ratio 80:20
DMAEMA Activator 2-(N,N-dimethylamino)Ethyl Fluka, Buchs, Switzerland
methacrylate
PFC1 Experimental composite D12, MMA, AAEM, initiator, Contract manufacturer
activator, inhibitor, particulate
filler
PFC2 Experimental composite D12, MMA, AAEM, BDDMA, Contract manufacturer
initiator, activator, inhibitor,
particulate filler
 d e n t a l m a t e r i a l s 2 2 ( 2 0 0 6 ) 845–851 847

Fig. 1 – The structural formulas of (a) dendrimer D12, (b) methyl methacrylate, (c) acetoacetoxyethyl methacrylate and (d)
1,4-butanediol dimethacrylate.
848 d e n t a l m a t e r i a l s 2 2 ( 2 0 0 6 ) 845–851

Table 2 – Compositions of the uncured samples (wt.%)

D12 MMA AAEM BDDMA Initiator system Particulate filler

AAEM0 78.4 19.6 – – 2.0 –

AAEM4 75.3 18.8 3.9 – 2.0 –
AAEM8 72.1 18.0 7.8 – 2.0 –
AAEM12 69.0 17.2 11.8 – 2.0 –
AAEM16 65.9 16.5 15.7 – 2.0 –
PFC1 20.7 5.2 2.9 – × 71.3
PFC2 15.6 3.9 2.8 5.6 × 72.2

composites are proprietary information of the manufacturer. The HS-GC/MS system (Perkin Elmer, Boston, MA, USA)
The composition of the particulate filler composites was sim- consisted of a Turbo Matrix 40 Headspace Sampler, an
ilar to dental restorative materials. The compositions of the AutoSystem XL Gas Chromatograph and a Turbomass Mass
samples are shown in Table 2. Evaporation of low-boiling com- Spectrometer connected to TurboMass GC/MS software. The
ponents (mainly MMA) had most probably taken place during vials were thermostatted for 30 min at 110 ◦ C. A sample of the
the mixing of the fillers to the resin mixture. gaseous phase inside the vial was extracted for 0.04 min and
transferred to the gas chromatograph with the injection nee-
2.2. HPLC analysis dle and transfer line at 115 ◦ C. The sample components were
separated in a BPX-5 (5% phenyl polysilphenylene-siloxane)
The specimens were polymerized in a mold of 1.5 mm in depth capillary column (50 m, 0.32 mm i.d., 1.0 ␮m solid phase; SGE,
and 6.6 mm in diameter. They were irradiated for 40 s with Austin, TX, USA) with helium at 1.5 ml min−1 as carrier gas.
a visible light-curing unit (Optilux 501, SDS Kerr, Danbury, The GC oven was temperature programmed as follows: 50 ◦ C
CT, USA;
= 400–505 nm, I ≈ 650 mW cm−2 ). The residual MMA for 2 min, 4 ◦ C min−1 to 150 ◦ C, 8 ◦ C min−1 to 220 ◦ C, hold for
was extracted with tetrahydrofuran inhibited with 20 ppm of 2 min. The components were identified with a mass selective
hydroquinone. The polymer specimens were placed in the detector run in scan mode, simultaneously with selected ion
solution and stirred with a magnetic stirrer for (72 ± 2) h at monitoring. The ions used for detection of components were
(24 ± 1) ◦ C. An aliquot of the supernatant was diluted with
methanol inhibited with 20 ppm of hydroquinone. The sample
solution was filtered to remove precipitated polymers.
The residual unreacted MMA and AAEM contents in the
photopolymerized specimens were determined with high per-
formance liquid chromatography (HPLC). A modular HPLC
system (LC-2010; Shimadzu Corporation, Kyoto, Japan) was
used with the following components connected to Shimadzu’s
CLASS VP software: a system controller (SCL-10Avp), a liquid
chromatograph pump (LC-10Advp), a UV–vis detector (SPD-
10Avp), an on-line degasser (DGU-14A), and an auto injec-
tor (SIL-10Advp). The column used was a Phenomenex Luna
250 mm × 2.00 mm C18 column with a particle size of 5 ␮m
(with two 4 mm × 2.00 mm C18 guard columns) and the elu-
ent a 6:4 mixture of HPLC grade methanol and particle-free
water. The residual monomers were detected at 205 nm.
The quantities of released MMA and AAEM were calculated
on the basis of calibration standard curves by taking the area
under the HPLC peaks. The residual monomer contents were
calculated as a percentage of the mass of the resin in the
polymerized specimens. Data are presented as means of six
repetitions. The effect of AAEM concentration on the residual
MMA and AAEM content in the unfilled resins was analyzed
with the Kruskal–Wallis test (SPSS for Windows; SPSS Inc.,
Chicago, IL, USA).

2.3. HS-GC/MS analysis

The specimens had dimensions identical to the HPLC speci-
mens. They were irradiated for 40 s with a visible light-curing
unit (Elipar Highlight, Espe, Seefeld, Germany;
= 400–500 nm,
I ≈ 600 mW cm−2 ). The specimens were sealed in headspace
vials containing butyl acetate as an internal standard.
Fig. 2 – The residual monomer content in dendrimer
D12/methyl methacrylate/acetoacetoxyethyl methacrylate
copolymers and composites analyzed with (a) HPLC (N = 6)
and (b) HS-GC/MS (N = 2). The error bars represent standard
deviation.
 d e n t a l m a t e r i a l s 2 2 ( 2 0 0 6 ) 845–851
Fig. 3 – Typical (a) HPLC and (b) HS-GC/MS chromatograms (AAEM12).
m/z = 41 for MMA, m/z = 69 for AAEM, and m/z = 43 for butyl
acetate.
Six consecutive extractions were taken from each vial (mul-
tiple headspace extraction, MHE). The total residual monomer
contents in the specimens were calculated using linear regres-
sion analysis of the peak areas obtained in the six extrac-
tions calibrated against the internal standard. The method
was adapted from Kolb and Ettre [2]. Data are presented as
means of two repetitions.
3. Results
On the basis of the HPLC analysis the addition of AAEM was
found to promote the photopolymerization of MMA (p = 0.014).
The amount of residual MMA decreased from 2.7 to 1.6%
of specimen weight as the initial concentration of AAEM
increased from 0 to 16 wt.%, but at the same time the amount
of residual AAEM increased from 0.3 to 1.7% (Fig. 2a). The effect
on residual AAEM was significant (p = 0.023). The total resid-
ual monomer content remained almost constant at 2.7–3.3%.
A typical HPLC chromatogram is presented in Fig. 3a.
The residual amounts of MMA in the PFC1 and PFC2 speci-
mens were 0.10 and 0.08% of resin weight and the amounts
of AAEM 0.65 and 0.77%, respectively (Fig. 2a). These val-
ues are comparable to the amounts of leaching residuals
measured previously in commercial composites [16,17]. The
small amount of residual MMA in PFC specimens compared
to unfilled resins was probably mainly due to earlier evapora-
tion of MMA at the manufacturing stage.
849
The results obtained with the HS-GC/MS method had a sim-
ilar trend as the HPLC results (Fig. 2b). The amount of MMA
detected in the PFC specimens was too small to be quanti-
fied with the MHE method. Minor amounts of CQ, DMAEMA
and toluene (from the manufacture of the dendritic monomer)
were also detected in the specimens with HS-GC/MS. The
amount of CQ ranged from less than 0.01–0.05% of the weight
of the whole specimen, and the amount of toluene from less
than 0.01–0.11%. The amounts were the lowest in the PFC spec-
imens. The amount of DMAEMA could not be quantified due
to nonlinear evaporation from the specimens. BDDMA was
not analyzed because it was present in only one experimental
composite. A typical HS-GC/MS chromatogram is presented in
Fig. 3b.
4. Discussion
AAEM is used in thermoset coatings to give lower solution
viscosity and to give lower glass transition temperature for
the polymer [18]. In this study, it was found to enhance the
polymerization of MMA which was seen as a decrease in
the residual MMA content, also when compared to the orig-
inal MMA content. Moreover, AAEM enhanced the polymer-
ization of the dendritic monomer, observed as an increase
in the degree of conversion of the copolymer from 52 to
60% as the original AAEM concentration changed from 0 to
16 wt.% [19]. This was most probably a result of the sys-
tem being less viscous and having more flexibility and thus
being able to polymerize further before vitrification. The same
850 dental materials
has been observed in the dilution of Bis-GMA with TEGDMA
[20].
However, about one tenth of the original amount of AAEM
remained unpolymerized in all resin mixtures. This resulted
in the total residual monomer content remaining high, which
may have an adverse effect on the mechanical properties
and biocompatibility of the polymer. The composition used
in the study has been assessed for cytotoxicity and neither
the unpolymerized resin nor the copolymer exhibited signs
of cytotoxicity [21]. However, AAEM monomer as such, is a
moderate skin and eye irritant and has potential for allergic
skin reactions, like many other methacrylate monomers [22],
references
which needs to be taken into consideration.
The amount of residual MMA detected in the PFC speci-
mens with HS-GC/MS could not be quantified with the MHE
method. The detection limit of HS-GC/MS appears to be higher
than that of HPLC and should be verified before use for routine
analysis. In the analysis of AAEM with HPLC some uncertainty
may arise from complications in the integration of the AAEM
chromatographic peak, due to the relatively low intensity of
the signal, especially at low AAEM concentrations, and limits
in the peak separation that can be achieved. It is also possible
that some unknown component elutes at the same time with
AAEM.
There are also other possible sources of difference in
the HPLC and HS-GC/MS measurements, if extensive paral-
lel series were measured and statistically compared. In HPLC,
an external standard was used [1] and the reliability of the
results was dependent on the reproducibility of pipetting in
the sample preparation. In HS-GC/MS, an internal standard
was used, which was a more reliable and practical standard-
ization method and minimized the number of measurements
[2].
The equilibration of the headspace specimens requires bal-
ancing between disadvantages. If the temperature is too high,
the polymer material may disintegrate. If the temperature is
too low, the diffusion of analytes from the sample may be
incomplete due to the long diffusion path from the polymer
matrix. In this study, an equilibrium between the solid and
gaseous phases in the headspace vial was reached in 30 min
at 110 ◦ C for MMA and AAEM, which was indicated by linear
MHE graphs, i.e. the partition coefficients were independent
of the analyte concentration. This thermostatting time was
considered reasonable. Therefore, no reduction of the size of
the solid specimen to facilitate faster equilibration was nec-
essary, thus avoiding an additional sample preparation step,
e.g. freeze-grinding. The temperature of 110 ◦ C has been used
previously for MMA and found suitable [3].
5. Conclusion
The addition of acetoacetoxyethyl methacrylate enhanced the
copolymerization of methyl methacrylate and the dendritic
monomer, but did not decrease the total residual monomer
content. The results obtained with the HS-GC/MS method had
a similar trend to the results from the standard HPLC method
in the analysis of low-boiling residuals in dental polymers. HS-
GC/MS has the advantages of being less laborious and also its
ability to identify other volatile compounds.
22 ( 2 0 0 6 ) 845–851
Acknowledgements
This study was supported by the Magnus Ehrnrooth Founda-
tion and Instrumentarium Science Foundation, Finland. The
research is part of the Bio- and Nano-polymers Research Cen-
tre of Excellence named by the Academy of Finland. Support
by the Finnish National Technology Agency (TEKES) is also
acknowledged. Ms. Minttu Vigren is acknowledged for prepar-
ing the resins.
[1] ISO 1567. Dentistry – Denture base materials, 1999.
[2] Kolb B, Ettre LS. Static headspace-gas chromatography:
theory and practice. New York: Wiley; 1997.
[3] Chai XS, Hou QX, Schork FJ. Determination of residual
monomer in polymer latex by full evaporation headspace
gas chromatography. J Chromatogr A 2004;1040:163–7.
[4] Garrigos MC, Marin ML, Canto A, Sanchez A.
Determination of residual styrene monomer in
polystyrene granules by gas chromatography-mass
spectrometry. J Chromatogr A 2004;1061:211–6.
[5] Gramiccioni L, Milana MR, Gesumundo C, Maggio A,
Denaro M. Experimental evaluation on adhesive pastes for
dental prostheses: I. Polyvinyl acetate-based formulations.
Rassegna Chim 1992;44:117–21.
[6] Örtengren U, Langer S, Göransson A, Lundgren T. Influence
of pH and time on organic substance release from a
model dental composite: a fluorescence
spectrophotometry and gas chromatography/mass
spectrometry analysis. Eur J Oral Sci 2004;112:530–7.
[7] Young A, Jonski G, Rolla G, Waler SM. Effects of metal
salts on the oral production of volatile sulfur-containing
compounds (VSC). J Clin Periodontol 2001;28:776–81.
[8] Viljanen EK, Skrifvars M, Vallittu PK. Degree of conversion
of a copolymer of an experimental monomer and methyl
methacrylate for dental applications. J Appl Polym Sci
2004;93:1908–12.
[9] Viljanen EK, Lassila LVJ, Skrifvars M, Vallittu PK. Degree of
conversion and flexural properties of a dendrimer/methyl
methacrylate copolymer: design of experiments and
statistical screening. Dent Mater 2005;21:172–7.
[10] Viljanen EK, Skrifvars M, Vallittu PK. Dendrimer/methyl
methacrylate copolymers: residual methyl methacrylate
and degree of conversion. J Bioma Sci Polym Ed
2005;16:1219–31.
[11] Vallittu PK, Lassila LVJ, Skrifvars M, Viljanen EK, Yli-Urpo
A. Dental and medical polymer composites and
compositions. International patent WO 02062901,
2002.
[12] Hale A, Macosko CW, Bair HE. Glass transition temperature
as a function of conversion in thermosetting polymers.
Macromolecules 1991;24:2610–21.
[13] Wise CW, Cook WD, Goodwin AA. Chemico-diffusion
kinetics of model epoxy-amine resins. Polymer
1997;38:3251–61.
[14] Scott TF, Cook WD, Forsythe JS. Photo-DSC cure kinetics of
vinyl ester resins. I. Influence of temperature. Polymer
2002;43:5839–45.
[15] Rånby B, Shi W. Radiation curable resins comprising
hyperbranched polyesters. US patent 5834118, 1998.
[16] Bagis YH, Rueggeberg FA. The effect of post-cure heating
on residual, unreacted monomer in a commercial resin
composite. Dent Mater 2000;16:244–7.
 dental materials
[17] Spahl W, Budzikiewicz H, Geurtsen W. Determination of
leachable components from four commercial dental
composites by gas and liquid chromatography/mass
spectrometry. J Dent 1998;26:137–45.
[18] Eastman. Utility of acetoacetoxyethyl methacrylate (AAEM)
in thermoset coatings. Eastman Chemical Company
Publication N-322D, 2003.
[19] Viljanen EK, Skrifvars M, Vallittu PK. Dendritic copolymers
and particulate filler composites for dental applications:
degree of conversion and thermal properties. Dent Mater,
submitted for publication.
22 ( 2 0 0 6 ) 845–851 851
[20] Morgan DR, Kalachandra S, Shobha HK, Gunduz N,
Stejskal EO. Analysis of a dimethacrylate copolymer
(Bis-GMA and TEGDMA) network by DSC and C-13 solution
and solid-state NMR spectroscopy. Biomaterials
2000;21:1897–903.
[21] Vallittu PK. Unpublished research results.
[22] Rustemeyer T, Frosch PJ. Occupational skin diseases in
dental laboratory technicians. (I) Clinical picture and
causative factors. Contact Dermatitis 1996;34:125–33.