Langmuir 1995,11, 4313-4322
4313
Surface Chemical Characterization Using X P S and TOF-SIMS of Latex Particles Prepared by the Emulsion Copolymerizationof Functional Monomers with Methyl Methacrylate and 4Winylpyridine M. C. Davies,*$tR. A. P. Lynn,?>$ J. Heam,$A. J. Pau1,l J. C. Vickerman," and J. F. Watts# Department of Pharmaceutical Sciences, University of Nottingham, Nottingham NG7 2RD, U.K., Department of Chemistry and Physics, Trent University, Nottingham NG11 8NS, U.K., Centre for Surface and Materials Analysis, Armstrong House, Oxford Road, Manchester M l 7ED, U.K.,Department of Chemistry, UMIST, P.O. Box 88, Manchester M60 lQD, U.K., and Department of Materials Science and Engineering, University of Surrey, Guildford GU2 5XH, U.K. Received December 16, 1994. I n Final Form: July 10, 1995@ The surface chemical analyses of a series of colloids based on poly(methy1methacrylate) and poly(4vinylpyridine) have been carried out using X-ray photoelectron spectroscopy (XPS)and time-of-flight The colloids were prepared by the vinyl polymerization secondary ion mass spectrometry (TOF-SIMS). of monomers bearing the reactive functional groups hydroxyl, amide, and epoxide. XPS and TOF-SIMS provided complementary information on the functional groups at the colloid particle surfaces and gave good correlationwith electrophoreticmobility and particle size data. This paper demonstrates the capability of these surface analysis techniques for studying the complex surface chemistries of copolymer colloid systems.
1. Introduction The presence of functional groups at the surface of polymer latex particles is a n important requirement for a range of biomedical and industrial applications.1-6 In many cases, the selective introduction of surface chemical groups can be achieved by the use of vinyl polymerization technique^,^ where one or more appropriately functionalized monomers are incorporated into the polymerization recipe. While the ability to modify latex properties in this way is extremely useful, the use of additional monomers complicates the polymerization kinetics and particle nucleation and formation mechanisms. Thus, the surface composition of the resultant polymer latex particle may differ significantly from that of the bulk and from the initial monomer composition. For many latex applications, a knowledge of the surface polymer composition vis-a-vis the bulk and/or monomer composition is important. Analytical determinations of monomer or functional group distributions have involved
* To whom correspondence should be addressed. University of Nottingham. Present address: SmithKline Beecham Pharmaceuticals, New Frontiers Science Park, Third Ave., Harlow, Essex CM19 5AW, U.K. 6 Trent University. Centre for Surface a n d Materials Analysis. +
the use of methods such as titration6-10 and nuclear magnetic resonance spectroscopy.l' More recently, surface analysis techniques such as X-ray photoelectron spectroscopy (XPS) and secondary ion mass spectrometry (SIMS) have been employed to evaluate the surface compositions of polymer lattices.12-16 In this study, X P S and time-of-flight SIMS (TOF-SIMS)17J8have been used, in conjunction with conventional methodologies, for the determination of the surface compositionsoffunctionalized polymer latex particles based on methyl methacrylate (MMA) and 4-vinylpyridine (VP), which comprise the following structures: CK=CH I
M U
VP
Two groups of polymer latex particles, based on MMA and VP, were prepared by incorporating the functional monomers 2-hydroxypropyl methacrylate (HPMA),gly~~
~~~
(8) Okubo, M.; Kanaida, K.; Tsunetaka, M. J.Appl. Polym. Sci. 1987,
33, 1511. (9)Greene, B. W. J. Colloid Interface Sci. 1973,43(21,449. (10)Emelie, B.; Pichot, C.; Guillot, J. J . Dispersion Sci. Technol. II UMIST. 1983,4(3-4),393. # University of Surrey. (11)McDonald, C. J.J . Dispersion Sei. Technol. 1984,5(3-4),365. Abstract published inAduance ACSAbstruct, October 15,1995. (12)Pijpers, A. P.;Donners, W. A.B. J . Polym. Sci., Polym. Chem. (1)Rembaim, A.;Yen, S. P. S.;Molday, R. S.J . Macromol. Sci.Chem. Ed. 1986,23,453. 1979.A13 (5).603. (13)Okubo, M.; Okegami, K.;Yamamota,Y.Colloid. Polym. Sci. 1989, 267,193. (2jRembaum, A.; Yen, S. P. S.; Cheong, E.; Wallace, S.; Molday, R. (14)Okubo, M.; Yamamoto, Y.; Kamei, S. Colloid. Polym. Sci. 1989, S.; Gordon, I. L.; Dreyer, W. J. Macromolecules 1976,9, 328. 267,861. (3)Orsini, A. J.; Ingenito, A. C.; Needle, M. A.; Debari, V. A. Cell (15)Davies, M. C.; Lynn, R. A. P.; Davis, S. S.; Hearn, J.; Watts, J. Biophys. 1987,10,33. (4)Blackley, D. C. Science and Technology of Polymer Colloids. In F.;Vickerman, J. C.; Paul, A. J. Lungmuir 1993,9, 1637. Preparation and Reaction Engineering; Poehlein, G. W., Ottewill, R. (16) Davies, M. C.; Lynn, R. A. P.; Davis, S. S.; Hearn, J.;Watts, J. H.,Goodwin, J. W.,Eds.;NATO-ASISeriesE(No.67);NijhoEDordrecht, F.;Vickerman, J. C.; Johnson, D. Langmuir 1994,10, 1399. The Netherlands, 1983;Vol. 1, p 203. (17)Standing, K.G.; Chait, B. T.; Ens, W.; McIntosh, G.; Beavis, R. (5)Pichot, C. Makromol. Chem.,Macromol. Symp. 1990,35136,327. Nucl. Instrum. Methods 1982,33,198. (6)Upson, D. A.J . Polym. Sci., Polym. Symp. 1986,72,45. (18)Bletsos, I. V.; Hercules, D. M.; Greifendorf, D.; Benninghoven, (7) Arshady, R. J . Microencapsulation 1988,5(2), 101. A. Anal. Chem. 1986,57,2384. @
0743-7463/95/2411-4313$09.00/0 0 1995 American Chemical Society
Davies et al.
4314 Langmuir, Vol. 11, No. 11, 1995 cidyl acrylate (GA), acrylamide (AM), and 2-hydroxyethyl methacrylate (HEMA). These comprise the following structures: CH3 I CH2= C
HPMA
"2
1
2
3
4
100
60 40
50
50
25 25
~~
5
6
100
40 60
25 25
GA
I
I
~
methyl methacrylate 2-hydroxypropyl methacrylate glycidyl acrylate acrylamide hydroxyethyl methacrylate 4-vinylpyridine
I
C 0" \OC€I@iCH3 I OH
o" C\
Table 1. Monomer Compositions for the MMA- and VP-Based Colloids composn for given colloid (% (w/w))
C 0" \OCH2CQOH
u ~ e f u l . l ,These ~ ~ particle systems can be prepared in a single step up to 15pm in size and can be linked to proteins using cyanogen bromide activation. In addition, vinyl pyridines complex with transition metal ions to form colored latices and complex with electron-dense metals such as gold for electron microscopy application^.^^
2. Materials and Methods The objective with these copolymer compositions is to introduce functional groups a t the particle surface (i.e., hydroxyl, amide, epoxide) which are then available for reaction with biological molecules such as monoclonal a n t i b o d i e ~ The . ~ ~ synthesis ~~~ of these types of polymer particles was pioneered by Rembaum and co-workers1,2 for various biological and immunological applications. The presence on the surface of the desired functionality is usually determined empirically by the success or otherwise of the subsequent immobilization procedure. However, direct chemical evidence for the required functionality is desirable, particularly if attempts are made to vary the surface density of the reactive sites as a prelude to immobilization. The various methods of activating surface functionalities for biomolecule immobilization are well documented.21For example, hydroxyl groups can be activated with cyanogen bromide and reacted with amino groups on proteins.22 The carbodiimide reaction is used for carboxyl activation prior to reaction with amino groups.23 Amide groups can be converted to the hydrazide derivative and subsequently coupled to proteins via the acyl azide.24 Alternatively, both amide and amino groups can be coupled to amino groups on proteinslantibodies using dialdehydes such as g l ~ t a r a l d e h y d e .Incorporation ~~ of epoxide groups permits direct reaction with proteins.26 One of the problems encountered with some coupling reagents is biomolecule inactivation. With this in mind, some authors have devised methods of preparing particles from activated monomers such as 1-(methacry1oxy)benzotriazole and N-(a~rylo~y)~~~~inimide.~~-~~ Of the more unusual materials used for latex preparation, those based on the vinyl pyridines are particularly (19) Rolland, A.: Bourel. D.: Genetet. B.: Le Verge. R. Int. J . Pharm.
1987, 39,
173.
-
I
(20) Illum, L.; Jones, P. D. E. Methods Enzymol. 1985, 112, 67. (21)Dean. P. D. G., Johnson. W. S.. Middle. F. A,. Eds. Afinitv
Chromatography-A Practical Apiroach; IRL Press: Oxford,U.K.;'1986. (22) Cuatrecasas, P. J.Biol. Chem. 1970,245 (12),3059. (23)Goodfriend,T. L.; Levine, L.; Fasman, G. D. Science 1964,144, 1344. (24) Inman, J. K.; Dintzis, H. M. Biochemistry 1969, 8, 4074. (25)Molday, R. S.; Dreyer, W. J.; Rembaum, A.; Yen, S. P. S. J.Cell Biol. 1975, 64, 75. (26) Hosaka,S.;Murao, Y.; Masuho, S.;Miura, K. Immunol. Commun. 1983, 12 (5), 509. (27) Yoshida, M.; Asano, M.; Yokota, T.; Chosdu, R.; Kumakura, M. J . Polym. Sci., Polym. Lett. Ed. 1989,27, 437. (28) Yoshida,M.; Asano, M.;Yokota,T.; Kumakura,M. Polymer 1990, 31 (2). , ~ ,371. -, -(29) Morita, Y.; Yoshida, M.; Asano, M.; Kaetsu, I. Colloid Polym. Sci. 1987, 265, 916. ~~
2.1. Monomer and Initiator Purification. Methyl methacrylate monomer (MMA, Polysciences, Wanington, PA), supplied with polymerization inhibitor(s),was purified according to the method of Riddle.31 This involved washing 100 mL of MMA monomer 5 times with 20-mL aliquots of a 20% (w/v)/5% (w/v) NaCUNaOH solution, followed by distillation at 63 "C in a nitrogen atmosphere at a pressure of about 200 mmHg. 2-Hydroxypropyl methacrylate monomer (HPMA,97%, Aldrich, Gillingham, U.K.), also containing a polymerization inhibitor (1200 ppm hydroquinone monomethyl ether), was purified by distillation under nitrogen a t 57 "C and 0.5 mmHg pressure. Glycidyl acrylate monomer (GA, 95%, Aldrich) was supplied free ofpolymerization inhibitorbut was distilled at 62-65 "C and a pressure of 5 mmHg to remove monomeric impurities. Acrylamide monomer (AM, 97%, Sigma, Dorset, U.K.) was recrystallized from chloroform (mp 84-86 "C). 2-Hydroxyethyl methacrylate monomer (HEMA, Aldrich), containing 300 ppm hydroquinone monomethyl ether, was purified by distillation at 95 "C and 1 mmHg pressure in the presence of 0.5%(w/v) hydroquinone. 4-Vinylpyridine monomer (VP,Aldrich) was supplied containing 100 ppm hydroquinone, which was removed by distillation in the dark over KOH pellets at 62-65 "C and 15 mmHg pressure.30 All purified monomers were stored under nitrogen a t 4 "C and protected from light until required. Potassium persulfate initiator (Analar grade, BDH, Poole, U.K.) was recrystallized twice from double-distilled water and dried in a desiccator. This process was repeated after 2 weeks' storage. 2,2'-azobis(isobutyramidine) dihydrochloride (ABA, Polysciences) was used as received. 2.2. heparation of Latex Particles. In order to minimize contamination of the latex samples during preparation and subsequent manipulations, all glassware was cleaned before use with chromic acid and rinsed repeatedly with double-distilled water. A series of six colloids based on MMA and VP and incorporating various functional monomers was prepared in the absence of emulsifier. The monomer compositions employed are shown in Table 1, and the preparation procedures were based on those described e1sewhe1-e~~ with the modifications described below.33 The polymerization recipes for the six colloids are shown in Table 2. Colloids 1 and 2 were prepared in the usual manne132%33 by heating the required quantity of double-distilled water to 70 "C and then adding the monomer(s1 immediately followed by the potassium persulfate (initiator) solution. For colloids 3 and 4 , the acrylamide solution was added to the remaining water and heated to 70 "C. At this point, the comonomer was added followed by the initiator. The fmal persulfate and monomer concentrations (30) Schwartz, A.; Rembaum, A. Methods Enzymol. 1985,112, 175. (31)Riddle, E. H. Monomeric Acrylic Esters; Reinhold: New York, 1954; p 7. (32)Goodwin, J. W.; Hearn, J.;Ho, C. C.; Ottewill,R. H. Br. Polym.
J. 1973, 5, 347. (33)Lynn, R. A. P. Ph.D Thesis, University of Nottingham, U.K., 1991.
Langmuir, Vol. 11, No. 11, 1995 4315
Latex Particles by Emulsion Copolymerization
Table 2. PolymerizationRecipes for Colloids Based on MMA and VP (Total Monomer Concentration 3% (w/v) for MMA-Based Systems and 1.5% (w/v) for VP-Based Systems) vol added for ~ v e colloid n (mL) 1 2 3 4 5 6 3.21 1.93 0.80 0.80 methyl methacrylatea 1.13 2-hydroxypropyl methacrylate" 0.34 glycidyl acrylatea 25.0 25.0 acrylamide (1.5%(w/v)in HzO) 0.37 0.29 hydroxyethyl methacrylatea 0.77 0.46 4-vinylpyridinea 3.0 3.0 1.5 1.5 potassium persulfate soln (0.1% (w/v)) 2,2'-azobis(isobutyramidine)-2HCl soln (0.5%(w/v)) 8.0 8.0 water (double distilled) 93.8 93.9 22.3 22.4 41.2 41.3 100.0 100.0 50.0 50.0 50.0 50.0 total vol (mL) ~
a
Based on the following densities (g/mL): MMA, 0.936; HPMA, 1.066; Ga, 1.099; HEMA, 1.034; 4-W, 0.975.
tion data were analyzed by a Malvern applications program (7025 in colloids 1-4 were 1.11 x mol dm-3 and 3% (w/v), Spect. I VI, Malvern) to yield particle size and polydispersity respectively. The polymerizations were continued for 8 h at 70 data. The polydispersity, Q, is derived from Koppel's method of "C. cummulants34 and is used to express the polydispersity numeriColloids 5 and 6 were also prepared using the methodology cally. Using this approach, monodisperse lattices have a value detailed e l s e ~ h e r e . In ~ ~preliminaq ,~~ experiments, it was found of Q = 0.03, but correlative data from electron microscopy suggest that the use of an anionic initiator (persulfate) in the presence that values of Q less than 0.1 reflect particle sizes that are of a basic monomer (VP) produced unstable systems. This narrowly distributed. A total of 20 measurements were recorded mol dm-3 ABA problem was remedied by the use of 1.48 x for each sample and mean size and polydispersity values as initiator, which produces cationic polymer end groups.33 The calculated. total monomer concentration was also reduced to 1.5% (wlv). The reactions were terminated after 6 h at 70 "C. The determination of electrophoretic mobility (EPM) and t Prior to cleaning, aggregates were removed from all colloids potential (ZP) was performed using laser Doppler anemomby filtration through Whatman No. 1 filter paper (Whatman, etry.34-36 Measurements were carried out using a Malvern U.K.). The percentage yield of polymer in latex form was Zetasizer 11. EPM measurements were made over the pH range estimated by taking a known volume of the cooled filtered latex 3.0-8.3 using phosphate-citrate buffers of constant ionic and evaporating to dryness. For colloids 3 and 4, excess strength (0.01 M). Samples were prepared by the addition of acrylamide was removed by exhaustive dialysis prior to drying. 200-300 pL of latex to 5 mL of the appropriate buffer solution. The weight of the polymer was determined, and, in each case, This level of dilution provided a suitable scattered light intensity the yield was greater than 80%,indicating satisfactory monomerwithout compromising buffering capacity. Five measurements to-polymer conversion. were made for each latex sample a t each pH. 2.3. CleaningandStorage of Latex Particles. The colloids 2.4.2. X-ray Photoelectron Spectroscopy. X P S spectra based on MMA were cleaned by repeated centrifugation and were obtained using aVG ESCALAB Mk I1 electron spectrometer replacement of the supernatant with double-distilled water. (VGScientificLtd., East Grinstead, Sussex, U.K.) employing Mg Briefly, 1-2-mL samples were centrifuged a t 13 000 rpm for 10 Ka X-rays (hv = 1253.6 eV) at an electron take-off angle of 45". min using a Micro Centaur (MSE, Fisons, Loughborough, U.K.). The base pressure of the spectrometer was typically mbar. The supernatant was carefully removed using a Pasteur pipet The X-ray gun was operated a t 10 kV and 20 mA, corresponding and replaced with water. The particles were resuspended by to a power of 200 W. A wide scan (0-1000 eV) was recorded for shaking, and the procedure was then repeated. The process was each sample (single scan) followed by the C Is, 0 Is, N Is, and monitored by performing a UV scan on each supernatant. The S 2p regions where appropriate (5 scans). The analyzer was colloid was judged to be clean when the UV absorbance of the operated in fixed analyzer transmission (FAT)mode with a pass supernatant was negligible over a wide range of wavelengths a t energy of 50 eV (wide scan) and 20 eV (C Is,0 Is, N Is, and S the maximum sensitivity of the spectrometer (Kontron Uvikon 2p regions). 860, Kontron, St. Albans, Herts., U.K.). In practice, this required Data analysis was performed on aVGS 5000 data system based a total of about five repeats. on a DEC PDP 11/73 computer. The methodology employed for This technique could not be used for the lattices based on VP peak fitting of the C Is and 0 1s envelopes has been described since centrifugation resulted in particle aggregation. These in detail el~ewhere.~'Typically, 1.5- 1.6-eV line widths and colloids were, therefore, cleaned by exhaustive dialysis against GaussianLorentzian ratios of 30% were employed for the double-distilled water using thoroughly cleaned Spectropor components of the C 1senvelope. Atomic percentage values and membrane (molecular weight cutoff, 12 000-14 000; Spectrum elemental ratios were calculated from the peak areas using Medical Industries, Los Angeles). The dialyzate was changed sensitivity factors and background subtraction. Spectra were every 24 h for a period of 14 days, by which time its conductivity corrected for sample charging by referencingphotoelectronpeaks was equal to that of water. The removal of residual monomer to C-CIC-H at 285 eV.3a and soluble oligomer was monitored in the manner described 2.4.3. Time-of-FlightSecondaryIon Mass Spectrometry. above, by removing samples at various time intervals, centrifugTOF-SIMS spectra were obtained using a VG M23S instrument39 ing, and performing a W scan on the supernatant. comprising a Poschenrieder TOF analyzer, a pulsed liquid metal Purified latex samples were stored in chromic acid-cleaned ion source (Ga+,30 keV) and a pulsed electron flood source for glass tubes a t 4 "C until required. charge c o m p e n ~ a t i o n .The ~ ~ secondary ions were accelerated to 2.4. Characterizationof Colloids. 2.4.1. Size and Elec5 keV for the TOF analysis by applying a bias to the sample. trophoretic Mobility Determination. Particle size measurements were carried out by photon correlation s p e c t r o ~ c o p y . ~ ~ J ~ (36)Earnshaw, J. C., Steer, M. W., Eds. The Applications o f h s e r Briefly, the system comprised a helium neon laser (Siemens, Light Scattering to the Study ofBiological Motion; Plenum: New York, Germany), a Malvern K7025 64-channel multibit correlator 1983. (Malvern Instruments Ltd., U.K.) and a Commodore PET 2001(37)Sherwood,P. M.A. In Practical Surface Analysis by Auger and 32N microcomputer (CommodoreBusiness Machines). CorrelaX-ray Photoelectron Spectroscopy; Briggs, D., Seah, M. P., Eds.; Wiley: Chichester, U.K., 1985;p 445. (38)Andrade, J.D.In Surface and Interfacial Aspects ofBiomedica1 (34)Cummins, H. Z., Pike, E. R., Eds. Photon Correlation and Light Polymers; Andrade, J. D., Ed.; Surface Chemistry and Physics;Plenum, Beating Spectroscopy; NATO-AS1 Series B (No. 3);Plenum: New York, New York, 1985; Vol. 1,p 105. 1974. (39)Eccles,A. J.;Vickerman, J. C. J. Vac. Sci. Technol. l989,A7 (2), (35)Cummins,H.Z.,Pike,E.R., Eds.Photon CorrelationSpectroscopy 234. and Velocimetry;NATO-AS1 Series B (No. 23);Plenum:New York, 1977.
4316 Langmuir, Vol. 11, No. 11, 1995
Davies et al.
Table 3. Particle Size (dz), Polydispersity Index (Q), Electrophoretic Mobility (EPM), and 5' Potential (ZP) Obtained for the Colloids EPMb (ccm 6-1 cm V-1) ZP (mV) monomer composn
dz" Q -
MMA MMA/HPMA MWGNAM MMAIHEWAM VP VPiHEMA
269 296 216 275 357 321
0.048 0.068 0.054 0.040 0.049 0.047
pH3.2
pH 7.1 pH 3.2 pH 7.1
-0.91 -0.93 -0.10 -0.20 2.64 2.80
-1.94 -2.08 -0.37 -1.05 1.52 -1.71
-11.9 -12.2 -1.3 -2.6 34.6 36.7
-25.5 -27.3 -4.9 -13.8 19.9 -22.5
a Mean of20 measurements, coefficientofvariation
