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Chemical derivatization technique in ToF-SIMS for quantification analysis of surface amine groups
Applied Surface Science 252 (2006) 6632–6635 www.elsevier.com/locate/apsusc
Chemical derivatization technique in ToF-SIMS for quantification analysis of surface amine groups Tae Geol Lee a,*, Jinmo Kim a,b, Hyun Kyong Shon a, Donggeun Jung b, Dae Won Moon a a
Nano-Surface Group, Korea Research Institute of Standards and Science (KRISS), P.O. Box 102, Daejeon 305-600, Republic of Korea b Department of Physics, Brain Korea 21 Physics Research Division and Institute of Basic Science, Sungkyunkwan University, Suwon 440-746, Republic of Korea Received 12 September 2005; accepted 15 February 2006 Available online 2 May 2006
Abstract A chemical derivatization technique in ToF-SIMS along with principal component analysis (PCA) were used to perform a quantitative study of the surface amine density of the plasma-polymerized ethylenediamine (PPEDA) thin film. We used the scores on principal component (PC) 1 from a PCA of ToF-SIMS data for the PPEDA films and their chemical-derivatized surfaces for comparison with the surface amine densities. These surface amine densities were independently determined by UV–visible spectroscopy. Our work found a good linear relationship between the surface amine densities and the scores on PC 1 from a PCA of the ToF-SIMS data for the chemical-derivatized PPEDA surfaces, but not for the PPEDA thin films themselves. In addition to quantification, our PCA results provided insights into the surface chemical composition of each surface. # 2006 Elsevier B.V. All rights reserved. Keywords: Chemical derivatization; ToF-SIMS; PCA; PPEDA; UV–visible spectroscopy
1. Introduction Controlling the surface density and selectivity of specific functional groups that exist on the surface is crucial in biomaterial applications since these functional groups control the immobilization of proteins or cells [1]. To fully control the surface density of a specific functional group, it is important to first have a useful surface analysis tool to quantify it. Compared to X-ray photoelectron spectroscopy (XPS) [2], the ToF-SIMS technique has played only an ancillary role in the quantification analysis of specific surface chemical compositions. This is mainly due to the complex matrix effect and low molecular secondary ion efficiency from organic/bio materials [3]. We have recently developed a chemical derivatization technique in ToF-SIMS to quantify the surface amine density of a plasma-polymerized ethylenediamine (PPEDA) thin film deposited on a glass surface [4]. The surface amine density was controlled as a function of deposition plasma power and quantified using UV–visible absorption spectrometry. Chemi-
* Corresponding author. Tel.: +82 42 868 5129; fax: +82 42 868 5032. E-mail address: [email protected] (T.G. Lee). 0169-4332/$ – see front matter # 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.apsusc.2006.02.214
cal tags were hybridized with the surface amine groups and were detected in ToF-SIMS spectra as characteristic molecular secondary ions. We found good linear correlations between the surface densities of the amine groups and the normalized ToFSIMS intensities of several individual secondary ions. In this work, we performed a principal component analysis (PCA) [5] on ToF-SIMS data obtained from PPEDA and their chemical-derivatized surfaces. The scores on principal component (PC) 1 of each surface were compared with their surface amine densities to obtain a correlation curve for the quantitative analysis. 2. Experimental 2.1. Preparation of PPEDA film The details of PPEDA films deposited on glass slides are reported in elsewhere [6]. The following is a brief description. PPEDA film was deposited at the substrate at room temperature with a deposition pressure of 30 mTorr, deposition time of 2 min, and an Ar flow rate of 30 sccm. Ethylenediamine was used as the monomer and put in a stainless steel bubbler, which was heated to 50 8C for vaporization. Two plasmas were used to
T.G. Lee et al. / Applied Surface Science 252 (2006) 6632–6635
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produce a high-quality amine surface. The inductively coupled plasma power varied from 3 to 70 W and was generated around the shower ring by a circular coil, which was connected to a 13.56 MHz radio frequency (RF) generator through a matching box. The 3 W of fixed substrate bias was generated from another RF generator and put into a substrate holder for plasma around the glass slide. The wall of the deposition chamber was grounded, and the base pressure of the deposition chamber was <10 6 Torr when pumped with a turbo molecular pump. 2.2. Chemical derivatization and UV–visible spectroscopy To determine the surface amine density of PPEDA film, UV– visible absorption spectrometry was used [4,7]. In a nitrogen atmosphere, a PPEDA-coated glass slide (1.5 cm 2.5 cm size) was allowed to react with excess 4-nitrobenzaldehyde (4NBA, 10 mg) anhydrous ethanol solution (25 mL) overnight at 50 8C. After the Schiff base reaction, the substrate was thoroughly washed and sonicated with absolute ethanol, methylene chloride, acetone and hexane for 3 min each sequentially and dried in a vacuum. At this stage, the chemically tagged substrate was submitted for ToF-SIMS analysis before hydrolysis. For hydrolysis, the imine-formed substrate was immersed in 1 mL volume of water overnight at 50 8C (Scheme 1). The aqueous solution of hydrolyzed 4-NBA (emax = 1.45 104 M 1 cm 1) was measured with a HP 8453 UV–visible spectrophotometer (HP, USA). All spectra were recorded after baseline correction and converted to surface amine densities in accordance with Beer’s law. 2.3. ToF-SIMS and PCA ToF-SIMS measurements were obtained with a TOF-SIMS V instrument (ION-TOF GmbH, Germany) using 25 keV Au+ primary ions (average current of 0.8 pA, pulse width of 16.8 ns, repetition rate of 5 kHz) at high-current bunched mode. The analysis area of 100 mm 100 mm was randomly rastered by primary ions and was charge compensated by low-energy electron flooding. The primary ion dose was kept below 1012 ions/cm2 to ensure static SIMS condition. Mass resolution was usually higher than 5000 at positive and negative modes. Mass calibration of positive and negative ion spectra was internally performed by using H+, H2+, CH3+, C2H3+ and C3H4+
Fig. 1. Positive ion ToF-SIMS spectra of PPEDA thin film deposited at 10 W of deposition plasma power (a) before and (b) after chemical tagging with 4-NBA.
peaks and H , C , CH , C2 and C2H peaks, respectively. A PCA of ToF-SIMS data was performed using PLS_Toolbox v. 3.5 (Eigenvector Research, Manson, WA) for MATLAB (MathWorks, Inc., Natick, MA). Raw data were normalized to the total secondary ion counts and mean-centered before the PCA process. 3. Results and discussion 3.1. ToF-SIMS analysis Six different PPEDA-coated thin films were left to react with 4-NBA and were measured using ToF-SIMS. Full spectra of PPEDA films obtained before and after the chemical tagging reaction are shown in Fig. 1a and b, respectively. We have included the full spectra for PPEDA made at 10 W plasma power as a typical case. Many peaks relating to aliphatic hydrocarbon (CxHy+) and aliphatic hydrocarbon containing nitrogen (CxHyNz+) are evident in a ToF-SIMS spectrum of the PPEDA film as shown in Fig. 1a. After a chemical tagging reaction with 4-NBA, several new peaks appear in the mass range from m/z 170 to 230, namely m/z 177, 191, 207 and 221. These new secondary ions originated from the chemical tag molecules 4-NBA and were tentatively assigned as C9H9N2O2+ (exact m/z 177.066, observed m/z 176.980), C10H11N2O2+ (exact m/z 191.082, observed m/z 190.996), C9H11N4O2+ (exact
Scheme 1.
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T.G. Lee et al. / Applied Surface Science 252 (2006) 6632–6635
Fig. 2. Plot of the surface amine densities of PPEDA film vs. scores on PC 1 from PCA of ToF-SIMS data for PPEDA and their chemical-derivatized surfaces (a and c) and their corresponding plots of m/z vs. loadings on PC 1 (b and d).
m/z 207.088, observed m/z 207.036) and C10H13N4O2+ (exact m/z 221.104, observed m/z 221.109) [4]. 3.2. PCA for quantitative analysis For comprehensive ToF-SIMS data interpretation, a PCA was performed on each set of raw data obtained from the PPEDA and the 4-NBA tagged surfaces. Two PCs, PC 1 (accounting for 78.26/82.79% of the variance) and PC 2 (accounting for 15.19/ 13.24% of the variance), were sufficient to explain the PPEDA/4NBA tagged PPEDA data set. Fig. 2a and b show the scores plot of PC 1 from PCA of the ToF-SIMS data for PPEDA as a function of the surface amine densities and the corresponding loadings plot of PC 1 as a function of m/z, respectively. For comparison, the scores and loading plots of PC 1 for 4-NBA tagged PPEDA are shown in Fig. 2c and d, respectively. Interestingly, there was a good linear correlation between the scores on PC 1 for 4-NBA tagged PPEDA and the surface amine densities. This is in comparison to the poor linear correlation between the scores on PC 1 for the PPEDA and their surface amine densities. Similar behaviors were observed when individual characteristic secondary ions signals from PPEDA and their chemical-derivatized surfaces were used for quantitative analysis [4]. The failure of the quantitative analysis with pure PPEDA may be due to the complex plasma-polymerization mechanism as a function of plasma deposition power, which results in non-linear correlations between the ToF-SIMS intensities and the surface amine densities for each individual peaks. In addition to the quantitative study, our PCA results gave us insights into the surface chemical compositions of each surface. For example, the negative scores on PC 1 for PPEDA film (i.e.,
4.4 amine groups/nm2 at 5 W, 3.6 amine groups/nm2 at 10 W, 3.2 amine groups/nm2 at 30 W, 2.9 amine groups/nm2 at 50 W and 2.7 amine groups/nm2 at 70 W) corresponded well to the peaks of m/z > 109. The positive scores on PC 1 for PPEDA film (i.e., 5.4 amine groups/nm2 at 3 W) corresponded well to the peaks of m/z < 67, as shown in Fig. 2a and b. This suggests that molecular cross-linking on the PPEDA film occurred due to the increased deposition plasma power [8]. To the contrary, the formation of small species containing intact amine groups occurred more frequently on the PPEDA surface at a lower deposition plasma power. Regarding the PCA results from the chemical-derivatized surfaces, the positive scores on PC 1 for the 4-NBA tagged PPEDA film (i.e., 5.4 amine groups/nm2 at 3 W and 4.4 amine groups/nm2 at 5 W) corresponded well to the peaks of m/ z > 177, as shown in Fig. 2c and d, suggesting that the more surface amine groups are on PPEDA surface, the more chemical tagging molecules are hybridized on the PPEDA surface. 4. Conclusions We have shown that a chemical derivatization technique in ToF-SIMS along with PCA is a useful method of studying the surface chemical composition of PPEDA thin films in a quantitative and systematic manner. There was a good linear relationship between the surface amine densities and scores on PC 1 from PCA of ToF-SIMS data for the chemical-derivatized PPEDA surface, compared to the poor linear relationship between the surface amine densities and scores on PC 1 for the PPEDA itself. We also found that PCA gave us insights into molecular cross-linking on plasma-polymerized thin films.
T.G. Lee et al. / Applied Surface Science 252 (2006) 6632–6635
Acknowledgement This work was supported by the R&D Program of Fusion Strategies for Advanced Technologies of MOCIE. References [1] D.G. Castner, B.D. Ratner, Surf. Sci. 500 (2002) 28. [2] D.S. Everhart, C.N. Reilley, Anal. Chem. 53 (1981) 665.
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[3] X.V. Eynde, in: J.C. Vickerman, D. Briggs (Eds.), ToF-SIMS: Surface Analysis by Mass Spectrometry, IM Publications and SurfaceSpectra Limited, Manchester, 2001, p. 543. [4] J. Kim, H.K. Shon, D. Jung, D.W. Moon, S.Y. Han, T.G. Lee, Anal. Chem. 77 (2005) 4137. [5] M.S. Wagner, D.J. Graham, B.D. Ratner, D.G. Castner, Surf. Sci. 570 (2004) 78. [6] J. Kim, H. Park, D. Jung, S. Kim, Anal. Biochem. 313 (2003) 41. [7] J.H. Moon, J.W. Shin, S.Y. Kim, J.W. Park, Langmuir 12 (1996) 4621. [8] N. Inagaki, Plasma Surface Modification and Plasma Polymerization, Technomic Publishing Co. Inc., Lancaster, USA, 1996, p. 153.
Chemical derivatization technique in ToF-SIMS for quantification analysis of surface amine groups Tae Geol Lee a,*, Jinmo Kim a,b, Hyun Kyong Shon a, Donggeun Jung b, Dae Won Moon a a
Nano-Surface Group, Korea Research Institute of Standards and Science (KRISS), P.O. Box 102, Daejeon 305-600, Republic of Korea b Department of Physics, Brain Korea 21 Physics Research Division and Institute of Basic Science, Sungkyunkwan University, Suwon 440-746, Republic of Korea Received 12 September 2005; accepted 15 February 2006 Available online 2 May 2006
Abstract A chemical derivatization technique in ToF-SIMS along with principal component analysis (PCA) were used to perform a quantitative study of the surface amine density of the plasma-polymerized ethylenediamine (PPEDA) thin film. We used the scores on principal component (PC) 1 from a PCA of ToF-SIMS data for the PPEDA films and their chemical-derivatized surfaces for comparison with the surface amine densities. These surface amine densities were independently determined by UV–visible spectroscopy. Our work found a good linear relationship between the surface amine densities and the scores on PC 1 from a PCA of the ToF-SIMS data for the chemical-derivatized PPEDA surfaces, but not for the PPEDA thin films themselves. In addition to quantification, our PCA results provided insights into the surface chemical composition of each surface. # 2006 Elsevier B.V. All rights reserved. Keywords: Chemical derivatization; ToF-SIMS; PCA; PPEDA; UV–visible spectroscopy
1. Introduction Controlling the surface density and selectivity of specific functional groups that exist on the surface is crucial in biomaterial applications since these functional groups control the immobilization of proteins or cells [1]. To fully control the surface density of a specific functional group, it is important to first have a useful surface analysis tool to quantify it. Compared to X-ray photoelectron spectroscopy (XPS) [2], the ToF-SIMS technique has played only an ancillary role in the quantification analysis of specific surface chemical compositions. This is mainly due to the complex matrix effect and low molecular secondary ion efficiency from organic/bio materials [3]. We have recently developed a chemical derivatization technique in ToF-SIMS to quantify the surface amine density of a plasma-polymerized ethylenediamine (PPEDA) thin film deposited on a glass surface [4]. The surface amine density was controlled as a function of deposition plasma power and quantified using UV–visible absorption spectrometry. Chemi-
* Corresponding author. Tel.: +82 42 868 5129; fax: +82 42 868 5032. E-mail address: [email protected] (T.G. Lee). 0169-4332/$ – see front matter # 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.apsusc.2006.02.214
cal tags were hybridized with the surface amine groups and were detected in ToF-SIMS spectra as characteristic molecular secondary ions. We found good linear correlations between the surface densities of the amine groups and the normalized ToFSIMS intensities of several individual secondary ions. In this work, we performed a principal component analysis (PCA) [5] on ToF-SIMS data obtained from PPEDA and their chemical-derivatized surfaces. The scores on principal component (PC) 1 of each surface were compared with their surface amine densities to obtain a correlation curve for the quantitative analysis. 2. Experimental 2.1. Preparation of PPEDA film The details of PPEDA films deposited on glass slides are reported in elsewhere [6]. The following is a brief description. PPEDA film was deposited at the substrate at room temperature with a deposition pressure of 30 mTorr, deposition time of 2 min, and an Ar flow rate of 30 sccm. Ethylenediamine was used as the monomer and put in a stainless steel bubbler, which was heated to 50 8C for vaporization. Two plasmas were used to
T.G. Lee et al. / Applied Surface Science 252 (2006) 6632–6635
6633
produce a high-quality amine surface. The inductively coupled plasma power varied from 3 to 70 W and was generated around the shower ring by a circular coil, which was connected to a 13.56 MHz radio frequency (RF) generator through a matching box. The 3 W of fixed substrate bias was generated from another RF generator and put into a substrate holder for plasma around the glass slide. The wall of the deposition chamber was grounded, and the base pressure of the deposition chamber was <10 6 Torr when pumped with a turbo molecular pump. 2.2. Chemical derivatization and UV–visible spectroscopy To determine the surface amine density of PPEDA film, UV– visible absorption spectrometry was used [4,7]. In a nitrogen atmosphere, a PPEDA-coated glass slide (1.5 cm 2.5 cm size) was allowed to react with excess 4-nitrobenzaldehyde (4NBA, 10 mg) anhydrous ethanol solution (25 mL) overnight at 50 8C. After the Schiff base reaction, the substrate was thoroughly washed and sonicated with absolute ethanol, methylene chloride, acetone and hexane for 3 min each sequentially and dried in a vacuum. At this stage, the chemically tagged substrate was submitted for ToF-SIMS analysis before hydrolysis. For hydrolysis, the imine-formed substrate was immersed in 1 mL volume of water overnight at 50 8C (Scheme 1). The aqueous solution of hydrolyzed 4-NBA (emax = 1.45 104 M 1 cm 1) was measured with a HP 8453 UV–visible spectrophotometer (HP, USA). All spectra were recorded after baseline correction and converted to surface amine densities in accordance with Beer’s law. 2.3. ToF-SIMS and PCA ToF-SIMS measurements were obtained with a TOF-SIMS V instrument (ION-TOF GmbH, Germany) using 25 keV Au+ primary ions (average current of 0.8 pA, pulse width of 16.8 ns, repetition rate of 5 kHz) at high-current bunched mode. The analysis area of 100 mm 100 mm was randomly rastered by primary ions and was charge compensated by low-energy electron flooding. The primary ion dose was kept below 1012 ions/cm2 to ensure static SIMS condition. Mass resolution was usually higher than 5000 at positive and negative modes. Mass calibration of positive and negative ion spectra was internally performed by using H+, H2+, CH3+, C2H3+ and C3H4+
Fig. 1. Positive ion ToF-SIMS spectra of PPEDA thin film deposited at 10 W of deposition plasma power (a) before and (b) after chemical tagging with 4-NBA.
peaks and H , C , CH , C2 and C2H peaks, respectively. A PCA of ToF-SIMS data was performed using PLS_Toolbox v. 3.5 (Eigenvector Research, Manson, WA) for MATLAB (MathWorks, Inc., Natick, MA). Raw data were normalized to the total secondary ion counts and mean-centered before the PCA process. 3. Results and discussion 3.1. ToF-SIMS analysis Six different PPEDA-coated thin films were left to react with 4-NBA and were measured using ToF-SIMS. Full spectra of PPEDA films obtained before and after the chemical tagging reaction are shown in Fig. 1a and b, respectively. We have included the full spectra for PPEDA made at 10 W plasma power as a typical case. Many peaks relating to aliphatic hydrocarbon (CxHy+) and aliphatic hydrocarbon containing nitrogen (CxHyNz+) are evident in a ToF-SIMS spectrum of the PPEDA film as shown in Fig. 1a. After a chemical tagging reaction with 4-NBA, several new peaks appear in the mass range from m/z 170 to 230, namely m/z 177, 191, 207 and 221. These new secondary ions originated from the chemical tag molecules 4-NBA and were tentatively assigned as C9H9N2O2+ (exact m/z 177.066, observed m/z 176.980), C10H11N2O2+ (exact m/z 191.082, observed m/z 190.996), C9H11N4O2+ (exact
Scheme 1.
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T.G. Lee et al. / Applied Surface Science 252 (2006) 6632–6635
Fig. 2. Plot of the surface amine densities of PPEDA film vs. scores on PC 1 from PCA of ToF-SIMS data for PPEDA and their chemical-derivatized surfaces (a and c) and their corresponding plots of m/z vs. loadings on PC 1 (b and d).
m/z 207.088, observed m/z 207.036) and C10H13N4O2+ (exact m/z 221.104, observed m/z 221.109) [4]. 3.2. PCA for quantitative analysis For comprehensive ToF-SIMS data interpretation, a PCA was performed on each set of raw data obtained from the PPEDA and the 4-NBA tagged surfaces. Two PCs, PC 1 (accounting for 78.26/82.79% of the variance) and PC 2 (accounting for 15.19/ 13.24% of the variance), were sufficient to explain the PPEDA/4NBA tagged PPEDA data set. Fig. 2a and b show the scores plot of PC 1 from PCA of the ToF-SIMS data for PPEDA as a function of the surface amine densities and the corresponding loadings plot of PC 1 as a function of m/z, respectively. For comparison, the scores and loading plots of PC 1 for 4-NBA tagged PPEDA are shown in Fig. 2c and d, respectively. Interestingly, there was a good linear correlation between the scores on PC 1 for 4-NBA tagged PPEDA and the surface amine densities. This is in comparison to the poor linear correlation between the scores on PC 1 for the PPEDA and their surface amine densities. Similar behaviors were observed when individual characteristic secondary ions signals from PPEDA and their chemical-derivatized surfaces were used for quantitative analysis [4]. The failure of the quantitative analysis with pure PPEDA may be due to the complex plasma-polymerization mechanism as a function of plasma deposition power, which results in non-linear correlations between the ToF-SIMS intensities and the surface amine densities for each individual peaks. In addition to the quantitative study, our PCA results gave us insights into the surface chemical compositions of each surface. For example, the negative scores on PC 1 for PPEDA film (i.e.,
4.4 amine groups/nm2 at 5 W, 3.6 amine groups/nm2 at 10 W, 3.2 amine groups/nm2 at 30 W, 2.9 amine groups/nm2 at 50 W and 2.7 amine groups/nm2 at 70 W) corresponded well to the peaks of m/z > 109. The positive scores on PC 1 for PPEDA film (i.e., 5.4 amine groups/nm2 at 3 W) corresponded well to the peaks of m/z < 67, as shown in Fig. 2a and b. This suggests that molecular cross-linking on the PPEDA film occurred due to the increased deposition plasma power [8]. To the contrary, the formation of small species containing intact amine groups occurred more frequently on the PPEDA surface at a lower deposition plasma power. Regarding the PCA results from the chemical-derivatized surfaces, the positive scores on PC 1 for the 4-NBA tagged PPEDA film (i.e., 5.4 amine groups/nm2 at 3 W and 4.4 amine groups/nm2 at 5 W) corresponded well to the peaks of m/ z > 177, as shown in Fig. 2c and d, suggesting that the more surface amine groups are on PPEDA surface, the more chemical tagging molecules are hybridized on the PPEDA surface. 4. Conclusions We have shown that a chemical derivatization technique in ToF-SIMS along with PCA is a useful method of studying the surface chemical composition of PPEDA thin films in a quantitative and systematic manner. There was a good linear relationship between the surface amine densities and scores on PC 1 from PCA of ToF-SIMS data for the chemical-derivatized PPEDA surface, compared to the poor linear relationship between the surface amine densities and scores on PC 1 for the PPEDA itself. We also found that PCA gave us insights into molecular cross-linking on plasma-polymerized thin films.
T.G. Lee et al. / Applied Surface Science 252 (2006) 6632–6635
Acknowledgement This work was supported by the R&D Program of Fusion Strategies for Advanced Technologies of MOCIE. References [1] D.G. Castner, B.D. Ratner, Surf. Sci. 500 (2002) 28. [2] D.S. Everhart, C.N. Reilley, Anal. Chem. 53 (1981) 665.
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[3] X.V. Eynde, in: J.C. Vickerman, D. Briggs (Eds.), ToF-SIMS: Surface Analysis by Mass Spectrometry, IM Publications and SurfaceSpectra Limited, Manchester, 2001, p. 543. [4] J. Kim, H.K. Shon, D. Jung, D.W. Moon, S.Y. Han, T.G. Lee, Anal. Chem. 77 (2005) 4137. [5] M.S. Wagner, D.J. Graham, B.D. Ratner, D.G. Castner, Surf. Sci. 570 (2004) 78. [6] J. Kim, H. Park, D. Jung, S. Kim, Anal. Biochem. 313 (2003) 41. [7] J.H. Moon, J.W. Shin, S.Y. Kim, J.W. Park, Langmuir 12 (1996) 4621. [8] N. Inagaki, Plasma Surface Modification and Plasma Polymerization, Technomic Publishing Co. Inc., Lancaster, USA, 1996, p. 153.