Comments on "Determination of Bisphenol A and Related Aromatic Compounds Released from Bis-GMA-Based Composites and Sealants by High Performance Liquid Chromatography".

I was very much disappointed in the paper by Pulgar et al. (1) because their laboratory once again presented unreliable and confusing HPLC data, regardless of my comments (2) on their previous paper (3). They also have provided no scientific response to my criticism (2).

The peak designated as BADGE in Figure 2A and C of their paper (1) is not bisphenol A diglycidylether (BADGE) because its retention time reads approximately 6 min from those chromatograms, but it is given as 7.10 min in their Table 1. This is supported further by the gas chromatogram (GC) presented in Figure 3A, which shows no peak for BADGE at 27.9 min. Thus, it is reasonable to assume that BADGE was not contained in Delton.

In addition, the peaks designated as bisphenol A propoxylate (PBPA) and bisphenol A etoxylate (EBPA) in Figure 2C (1) are doubtful because no peaks are noticeable at 27.7 and 23.9 min in the GC presented in Figure 3A and no mass spectra are shown. The presence of bisphenol A diglycidylether methacrylate (Bis-GMA) in a polymerized sample of Delton is not clear because Figure 3C shows no molecular ion peak for Bis-GMA, whereas Pulgar et al. (1) identified a GC peak at 22.8 min in Figure 3A as Bis-GMA.

Pulgar et al. (1) also present curious data concerning the elution of bisphenol A (BPA) and bisphenol A dimethacrylate (Bis-DMA) in Tables 6, 8, and 10 and Tables 5, 6, and 9, respectively. BPA eluted more in the polymerized samples than in the nonpolymerized ones at all pHs indicated. This should be reversed because the diffusion of BPA is usually more limited in polymerized samples than in the nonpolymerized ones. The increase in elution of Bis-DMA at pH 9 and pH 12 compared to pH 1 and pH 7 is unlikely because Bis-DMA is hydrolyzed more easily at alkaline conditions than at acidic or neutral conditions.

Therefore, based on these reasons, I suggest that the designation of each peak shown in Figure 2 of Pulgar et al. (1) is not reliable, and therefore all data shown in Tables 3-10 are also doubtful. I also suggest that the HPLC analysis be performed more carefully and thoroughly. Each peak should be carefully examined to establish whether it originates from a single pure compound or from a mixture of different compounds. It is difficult to separate the BPA peak from the those of other compounds contained in Bis-GMA or Bis-GMA-based resins under the HPLC conditions used by Pulgar et al. (1). In their report it appears that commercial Bis-GMA monomer is a pure compound; however, commercially available Bis-GMA is not pure but a mixture of many compounds.

My laboratory (4,5) has established that commercial Bis-GMA is composed of many minor components and four major components: Bis-GMA, 2,2-[4-(2-hydroxy-3-methacryloyloxy-1 -propoxy)-4'-(3-hydroxy-2-methacryloyloxy-1 -propoxy)]diphenylpropane (Iso-bis-GMA), 2,2-[4-(2-hydroxy-3-methacryloyloxy -1-propoxy)-4'-(2,3-dihydroxy-l-propoxy)] diphenylpropane (Bis-GMA-H), and 2,2-[4-(2-hydroxy-3-methacryloyloxy-1-propoxy) -4'-(2,3-dimethacryloyloxy-1-propoxy)]diphenylpropane (Bis-GMAM). We analyzed three commercial Bis-GMA monomers and six Bis-GMA-based composite resins including Z-100, Charisma, and Tetric under the following HPLC conditions: we used a C18 column (flow of 1 mL/min) with acetonitrile/water (50/50) as the solvent, a temperature of 40 [degrees] C, a duration of 55 min (isocratic mode), UV detection (at 230 nm), and fluorescence (excitation 275 nm; emission 300 nm) (5). Using this HPLC analysis, we separated the peak of BPA from that of Bis-GMA-H (which has a peak at a retention time close to that of BPA) and other impurities. Moreover, we also confirmed that BPA and Bis-GMA-H are included in the same peak under certain HPLC conditions (4).

Quantification of the BPA content was only possible when a fluorescence detector was used, and it was impossible using a UV detector because of the extremely low BPA content (5). We found 43-130 [micro]g BPA/g Bis-GMA and 1.5-10.2 [micro]g BPA/g unpolymerized composite resin. The contents of Bis-GMA-H were measured at 18-50 mg/g Bis-GMA and 1.1-1.4 mg/g composite resin (5). The elution of Bis-GMA-H from polymerized Z-100 composite was 58 mg/g composite in 37 [degrees] C distilled water during a 24-hr period (6). This value is within the range of 3-165 [micro]g BPA/g polymerized composite at pH 7 shown by Pulgar et al. (1) in their Tables 4, 5, 7, 8, and 9. This suggests that most of the "BPA" reported by Pulgar et al. was probably Bis-GMA-H from the five Bis-GMA-based resins. Regarding BPA content in composite resins, Manabe et al. (7) reported 6.4 [micro]g BPA/g unpolymerized resin for a commercial resin analyzed by GC-mass spectrometry. This value is quite similar to our data. Thus, we sugggest that the BPA content in commercial dental resins is usually a maximum of 10 [micro]g/g resin. Moreover, long-term leaching of BPA from a polymerized resin in water has been predicted to be slight and slow (6). Therefore, little or no long-term estrogenic effect due to BPA contained in Bis-GMA-based dental resins can be expected.

Finally, I suggest that Pulgar et al. (1) replace the chemical structures for BADGE and Bis-GMA shown in their Figure 1 with correct ones. In conclusion, I suggest that Pulgar et al. (1) withdraw the confusing data presented in the paper or totally revise their paper.

REFERENCES AND NOTES

(1.) Pulgar R, Olea-Serrano MF, Novillo-Fertrell A, Rivas A, Pazos P, Pedraza V, Navajas J-M, Olea N. Determination of bisphenol A and related aromatic compounds released from Bis-GMA-based composites and sealants by high performance liquid chromatography. Environ Health Perspect 108:21-27 (2000).

(2.) Imai Y. Comments on "Estrogenicity of resin-based composites and sealants used in dentistry." Environ Health Perspect 107:A290 (1999).

(3.) Olea N, Pulgar R, Perez P, Olea-Serrano F, Rivas A, Novillo-Fertrell A, Pedraza V, Soto AM, Sonnenschein C. Estrogenicity of resin-based composites and sealants used in dentistry. Environ Health Perspect 104:298-305 (1996).

(4.) Ohsaki A, Imai Y. Analysis of major components contained in bis-GMA monomer. Dent Mater J 18:425-429 (1999).

(5.) Imai Y, Watanabe M, Ohsaki A. Analysis of major components and bisphenol A in commercial bis-GMA and bis-GMA-based resins using high performance liquid chromatography. Dent Mater J 19:263-269 (2000).

(6.) Imai Y, Komabayashi T. Elution of bisphenol A from a composite resin: a model experiment. Dent Mater J 19:133-138 (2000).

(7.) Manabe A, Kaneko S, Numazawa S, Itoh K, Inoue M, Hisamitu H, Sasa R, Yoshida T. Detection of bisphenol-A in dental materials by gas chromatography-mass spectrometry. Dent Mater J 19:75--86 (2000).

Yohji Imai Institute of Biomaterials and Bioengineering Tokyo Medical and Dental University Tokyo, Japan E-mail: [e-mail]