Block copolymers for tooth enamel protection

Abstract

Described herein are block copolymers having hydrophobic blocks and hydrophilic blocks which are effective in binding to the surface of hard tissue; compositions comprising the same, as well as methods of making and using the same.

Claims

1. An oral hygienic composition comprising: an orally acceptable carrier; fluoride, and a block copolymer having at least one hydrophobic block and at least one hydrophilic block in an amount effective to bind hydroxyapatite; and wherein the oral hygienic composition is selected from the group consisting of: toothpaste, mouthwash, strips, and gel containing trays; and wherein the composition comprises effective amounts of fluoride, the block copolymer, and the carrier to reduce citric acid erosion of hydroxyapaptite by 15-30% as compared to the same composition without fluoride; wherein the block copolymer is a poly methyl methacrylate-poly methacryloyloxyethyl phosphate block copolymer or a poly methyl methacrylate-poly acrylate acid block copolymer.

2. The oral hygienic composition of claim 1, wherein the block copolymer is effective to protect the hydroxyapatite from loss of calcium by at least about 10 percent after exposure of the hydroxyapatite to the copolymer and subsequent exposure of the copolymer coated hydroxyapatite to citric acid.

3. The oral hygienic composition of claim 1, wherein the block copolymer has a molecular weight in the range of 1,000 to 1,000,000, individual hydrophilic blocks having a molecular weight in the range of 200 to 1,000,000, and individual hydrophobic blocks having a molecular weight in the range of 200 to 1,000,000.

4. The oral hygienic composition of claim 1, wherein the hydrophilic blocks comprise from 10 to 90 weight percent of the block copolymer and the hydrophobic blocks comprise from 10 to 90 weight percent of the block copolymer.

5. The oral hygienic composition of claim 1, wherein the block has a molecular weight and the polymers have a total molecular weight effective to provide a solubility in water of 0.001 to 100 g/l.

6. The oral hygienic composition of claim 1, wherein the block copolymer has a molecular weight in a range of 1,000 to 1,000,000.

7. The oral hygienic composition of claim 1, wherein the block copolymer has a molecular weight in a range of 1,000 to 10,000.

8. A method for protecting tooth enamel from acid erosion, the method comprising: applying a block copolymer according to claim 1 to tooth enamel, the block copolymer having at least one hydrophobic block and at least one hydrophilic block which are effective to bind to hydroxyapatite.

9. The method of claim 8, wherein the block copolymer is effective to protect the hydroxyapatite from loss of calcium by at least about 10 percent after exposure of the hydroxyapatite to the copolymer and subsequent exposure of the copolymer coated hydroxyapatite to citric acid.

10. The method of claim 8, wherein the block copolymer has a molecular weight in a range of from about 1,000 to have 1,000,000.

11. The method of claim 8, wherein the block copolymer has a molecular weight in the range of from about 1,000 to about 1,000,000, individual hydrophilic blocks having a molecular weight in the range of from about 200 to about 1,000,000, and individual hydrophobic blocks having a molecular weight in the range of from about 200 to about 1,000,000.

12. The method of claim 8, wherein the hydrophilic blocks comprise from about 10 to about 90 weight percent of the block copolymer and the hydrophobic blocks comprise from about 10 to about 90 weight percent of the block copolymer.

13. The method of claim 8, wherein the tooth enamel is exposed to a citric acid solution before and/or after applying the block copolymer.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) The present invention will become more fully understood from the detailed description and the accompanying drawings, wherein:

(2) FIG. 1: Controlled synthesis of hydrophobic block via RAFT method by using different monomer/CTA/initiator ratios. MMA stands for methyl methacrylate, CTA stands for chain transfer agent, AIBN stands for azoisobutyronitrile, and MOEP stands for methacryloyloxyethyl phosphate.

(3) FIGS. 2a-2f: Illustration of Fourier transform Infrared (FTIR) spectra of enamel treated with polymers at different pHs and concentrations. FIGS. 2a, 2c and 2e relate to aqueous compositions where the polymers are solubilized at 1 g/L. FIGS. 2b, 2d and 2f relate to aqueous compositions where the polymers are solubilized at 0.2 g/L. The pH of the treatment solution is 3.1 in FIG. 2a, 3.7 in FIG. 2b, 4.2 in FIGS. 2c and 2d and 7.0 in FIGS. 2e and 2f. The insert in FIG. 2a is a FTIR spectrum of a phosphate copolymer.

(4) FIGS. 3-1 to 3-7: FIGS. 3-1 through 3-3 indicate the UV spectra of a phosphate block copolymer before and after binding with HA powder at different conditions. 3-1: UV spectra of P(MMA).sub.19-b-P(MOEP).sub.9 with known concentrations. 3-2: UV spectra of P(MMA).sub.19-b-P(MOEP).sub.9 after binding with HA powder at pH=4. 3-3: UV spectra of P(MMA).sub.19-b-P(MOEP).sub.9 after binding with HA powder at pH=7. FIG. 3-4 through FIG. 3-6 illustrate the UV spectra of carboxylic block copolymers before and after binding with HA at different conditions. 3-4: UV spectra of P(MMA).sub.17-b-P(AA).sub.35 with standard concentrations. 3-5: UV spectra of P(MMA).sub.17-b-P(AA).sub.35 after binding with HA powder at pH=4. 3-6: UV spectra of P(MMA).sub.17-b-P(AA).sub.35 after binding with HA powder at pH=7. FIG. 3-7 shows dependence of phosphorylated block copolymer and carboxylated adsorption on concentration and pH. Lower pH indicates more polymer adsorbed to HA, while the phosphorylated polymer can be adsorbed onto HA more than the carboxylated polymer.

(5) FIGS. 4-1 & 4-2: Show the enamel surface morphology after exposing to acid. 4-1: SEM images of untreated enamel surface before (a) and after (b) acid erosion. 4-2: is the enamel surface morphology which was treated by phosphate block copolymer and then exposed to acid erosionSEM images of P(MMA).sub.19-b-P(MOEP).sub.9 treated-enamel surface before and after acid erosion.

DETAILED DESCRIPTION

(6) The following description of the preferred embodiment(s) is merely exemplary in nature and is in no way intended to limit the invention, its application, or uses.

(7) As used throughout, ranges are used as shorthand for describing each and every value that is within the range. Any value within the range can be selected as a terminus of the range and is encompassed by the invention. In addition, all references, patents, patent application publications and books cited herein are hereby incorporated by referenced in their entireties. In the event of a conflict in a definition in the present disclosure and that of a cited reference, the present disclosure controls.

(8) Unless otherwise specified, all percentages and amounts expressed herein and elsewhere in the specification should be understood to refer to percentages by weight. The amounts given are based on the active weight of the material.

EXAMPLES AND TESTS

(9) 1. Controlled synthesis of hydrophobic blocks

(10) 2. Block copolymer synthesis

(11) 3. Polymer/enamel binding

(12) 4, Quantitative analysis of polymer/HA binding

(13) 5. Anti erosion test by phosphate block copolymer

(14) 6. Anti erosion test by phosphate block copolymer in presence of fluoride

(15) 7. SEM observation on the surface morphology of enamel

(16) 1. Controlled Synthesis of Hydrophobic Blocks

(17) Typically, 10 mmol MMA, 0.25 mmol RAFT CTA agent (e.g. 4-cyano-4-[(dodecylsulfanylthiocarbonyl)sulfanyl]pentanoic acid) and 0.1 mmol AIBN were dissolved in 10 ml 1,4-dioxane. After purging with Argon for 1 h, the system was heated to 70 C. for a period of time. Gel permeation chromatography (GPC) was used to monitor the average macromolecular weight (Mn) of hydrophobic block. For example, Mn of polymethyl methacrylate (PMMA) can be well controlled using different monomer/CTA/initiator ratios as shown in FIG. 1.

(18) 2. Block Copolymer Synthesis

(19) ##STR00011##

(20) The synthesis of PMMA-b-PMOEP is shown in Scheme 1. Once the targeted Mn of PMMA segment was achieved, certain amounts of methacryloyloxyethyl phosphate (MOEP) in 1,4-dioxane was then injected into the system with syringe and the reaction was further allowed to continue for different reaction times. The composition of PMMA-b-PMOEP could be adjusted by using different feeding ratios and different polymerization times as shown in Table 1.

(21) TABLE-US-00001 TABLE 1 Composition of PMMA-b-PMOEP using different polymerization time and feeding ratios RAFT -chain Number of Number of MMA transfer agent AIBN MOEP MMA in MOEP in (mol) (mol) (mol) (mol) copolymer copolymer 100 2.5 1 50 17 9 100 2.5 1 50 19 14 100 2.5 1 80 17 14 100 2.5 1 80 20 35

(22) The synthesis of PMMA-b-PAA by RAFT polymerization is shown in Scheme 2-1. Once the targeted Mn of PMMA segment was achieved, certain amounts of acrylic acid (AA) in 1,4-dioxane was then injected into the system with syringe and the reaction was further allowed to continue for different reaction times. The composition of PMMA-b-PAA could be adjusted by using different feeding ratios and different polymerization times as shown in Table 2. PAA stands for poly acrylate acid.

(23) The synthesis of PMMA-b-PAA can also be prepared by an indirect method shown in Scheme 2-2.

(24) ##STR00012##

(25) ##STR00013##

(26) The hydrophobic and hydrophilic block chain lengthen can be adjusted by monomer/CTA/initiator ratio and polymerization time. Different block copolymers with different compositions are shown in Table 2.

(27) TABLE-US-00002 TABLE 2 Compositions of carboxylate block copolymers Code Mn AA fraction tBE1 9.7k PMMA.sub.77-b-PAA.sub.23 0.23 tBE2 9.7k PMMA.sub.73-b-PAA.sub.28 0.28 tBE3 11.7k PMMA.sub.67-b-PAA.sub.64 0.49 tBE4 21.5k PMMA.sub.69-b-PAA.sub.198 0.74 tBE5 29.9k PMMA.sub.67-b-PAA.sub.318 0.83

(28) 3. Polymer/Enamel Binding

(29) The structure of block copolymer used in this test is P(MMA).sub.19-b-P(MOEP).sub.14. Before polymer treatment, the surface of bovine enamel was pre-conditioned by immersing the enamel in 1% citric acid solution (pH=3.8) for 5 min. Polymer solution with different concentrations (0.2 and 1.0 g/L) and different pHs (3.1, 4.2 and 7.0) were used to treat the bovine enamel surface for 5 min at 50 rpm. Then the treated surface was washed with phosphate buffer solution (pH=7.0) and acid solution (pH=3.8) for three cycles (5 min/cycle). The treated and etched enamel was characterized by FTIR spectroscopy after air dry. The FTIR spectra are shown in FIG. 2. The peaks at 1452, 1407, and 869 cm.sup.1 could be assigned to the existence of carbonated hydroxyapatite on the surface. The peak at 1730 cm.sup.1 could be ascribed to the characteristic absorption peak of CO in block copolymers. Both the effects of polymer concentration and pH on the binding were evaluated. When increasing the polymer concentration from 0.2 to 1.0 g/L, the relative intensity of peak at 1730 cm.sup.1 was increased, indicating higher polymer concentration could facilitate the binding efficiency. This could be ascribed to the strong interaction between phosphate groups in the block copolymer and the active site on enamel. Also, from the FIGS. 2a, 2c and 2e, when the pH is increased from 3.1 to 7.0 and the polymer concentration is kept constant as 1.0 g/L, less polymer could be adsorbed onto the enamel surface. The first and second dissociation constants, pK.sub.a1 and pK.sub.a2 for phosphoric acid are 2.12 and 7.21, respectively. The phosphate groups of the copolymer are believed to exist in the form of RHPO.sub.4.sup. and RPO.sub.4.sup.2, where R stands for the polymer side chains attached to the backbone. The former moiety (RHPO.sub.4.sup.) will be dominant over the latter one at the pH range (3.1-7.0) in this test. The phosphate block copolymer with negative charge could bind with the calcium domains on HA surface via electrostatic interaction. A lower pH value of the polymer solutions appears to facilitate the binding.

(30) 4. Quantitative Analysis of Polymer/Hydroxyapatite (HA) Binding

(31) The structures of block copolymers used in this test are P(MMA).sub.19-b-P(MOEP).sub.9 and P(MMA).sub.17-b-P(AA).sub.35. Polymer solutions of 5 ml with different concentrations and different pH values were mixed with 100 mg HA powder for 2 h at room temperature. After centrifuging for 10 min at 10000 rpm, the solution was used tested by UV-vis spectroscopy. The absorbance of thiocarbonyl group (CS) before and after binding were utilized to calculate the adsorbed polymer onto HA powder. The calibration curve was performed by using polymer solution with known concentrations. The UV spectra of phosphorylated or carboxylated block copolymer before and after binding are shown in FIG. 3-1 to FIG. 3-6. The calculated adsorbed polymer bound to HA is shown in FIG. 3-7. It can be seen that when the polymer concentration is gradually increased from 0.06 to 1.0 g/L, more and more polymer could be adsorbed onto the HA surface.

(32) 5. Anti Erosion Test of Phosphate Block Copolymer

(33) The structure of block copolymer used in this test is P(MMA).sub.17-b-P(MOEP).sub.12 and P(MMA).sub.18-b-P(AA).sub.29. Atomic absorption (AA) spectrometry is one of the most reliable and sensitive methods on evaluating the dental erosion by monitoring the mineral loss. The typical testing procedure used was as follows. First, sintered hydroxyapatite (HA) discs were immersed in 1% citric acid (pH=2.5) for 15 min at room temperature, then soaked in water and sonicated for 30 min. HA discs were fixed on a 6 well plate by using KERR compounds. Note that only the top surface of HA was exposed to the solutions. After air drying, the fixed HA discs were challenged by 1% citric acid (pH=3.8) for 15 min at 37 C. with a shaking speed of 50 rpm. The solution was collected and the calcium concentration was designated as [Ca].sub.ref. The HA discs were washed with phosphate buffer solution (PBS, pH=7.0) and then treated with polymer solution (1 g/L) or PBS (as blank) for 2 min. After another washing with PBS, the HA was again challenged with citric acid for another 15 min. The solution was collected and the calcium concentration was measured by AA spectrometry [Ca].sub.treat. Because of the heterogeneity among HA samples, the relative calcium level (Ca level), calculated as the following equation (S1), was utilized as an index to assess the protecting efficiency against acid erosion.

(34) $\begin{array}{cc}\mathrm{Ca}\mathrm{level}=\frac{{\left[\mathrm{Ca}\right]}_{\mathrm{treat}}}{[{\mathrm{Ca}}_{]\mathrm{ref}}}*100\%& \mathrm{Equation}\mathrm{S1}\end{array}$

(35) The different polymer treatments on HA surface could influence the calcium level as shown in Table 3. The calcium level after phosphorylated polymer treatments with different polymer treating times was decreased from 91% for blank (non-polymer treated) to 50%, 48%, 34%, 17% for 0.5, 1, 2, or 5 minutes polymer treatment, respectively. The calcium level after carboxylated polymer treatments with different polymer treating time was decreased from 91% for blank (non-polymer treated) to 56%, 60%, 64%, 31% for 0.5, 1, 2, or 5 minutes polymer treatment, respectively. The possible reason is that the adsorbed polymer onto enamel/HA could form a protective layer and prevent the mineral from release.

(36) TABLE-US-00003 TABLE 3 Calcium released level (%) following acid erosion challenge Treating time Treatment 0 30 s 1 min 2 min 5 min Blank (PBS buffer) 90.7 n/a n/a n/a n/a P(MMA).sub.18-b-P(AA).sub.29, n/a 56.4 59.6 63.9 31.2 pH = 4.2 P(MMA).sub.17-b-P(MOEP).sub.12, n/a 50.3 48.3 34.1 16.7 pH = 4.2

(37) The treatment with phosphate monomer and block copolymer on HA surface could influence on the calcium level as shown in Table 4. The calcium level without treatment is 90%. With treatment with phosphate monomer, the calcium level is still around that level, indicating phosphate monomer treatment has a negligible effect on inhibiting mineral loss during acid challenge. Once the HA is treated by phosphate block copolymer, the calcium level is significantly decreased to 43%, meaning that phosphate block copolymer could protect tooth by lowering down the mineral loss during acid challenge. The possible reason is that the adsorbed phosphate block copolymer onto enamel/HA via its phosphate groups and the hydrophobic groups could obstruct the acid attack by forming a protective layer.

(38) TABLE-US-00004 TABLE 4 The effect of phosphate monomer and phosphate block copolymer (P(MMA).sub.20-b-P(MOEP).sub.35) on calcium released level Concentration, Calcium Level Treatment g/L pH (%) Blank (PBS buffer) n/a 7.0 90.4 13.6 Phosphate monomer 1.0 4.2 88.5 12.6 P(MMA).sub.20-b-P(MOEP).sub.35 1.0 4.2 42.5 7.5

(39) The carboxlyate block copolymers' protecting effect is similarly evaluated based on the protocol above and the result is shown in Table 5, where the carboxylic monomer, AA, and its homopolymer, polyacrylic acid (PAA), are also included for comparison. The calcium level was also decreased most for the block co-polymer. The pH value doesn't show a significant influence on the anti erosion behavior of the carboxylate block copolymers.

(40) TABLE-US-00005 TABLE 5 The effect of carboxylic monomer (M-AA), acrylic acid homopolymer (AA), and tBE4 (PMMA-b-PAA block copolymer) on calcium released level Concentration, Calcium level Treatment g/L pH (%) Blank (PBS buffer) n/a 7.0 90.4 13.6 Acrylic acid monomer 1.0 4.2 77.6 5.6 PAA 1.0 4.2 86.1 14.7 PMMA-b-PAA (tBE4) 1.0 4.2 65.3 9.4

(41) In order to make a comprehensive comparison, some commercially available copolymers with random structure as well as other carboxylic block copolymers as shown in Table 6. It can be shown that both phosphate and carboxylate block copolymers exhibited a lower value of calcium level released, implying the importance of block structure in protecting tooth from acid challenge. Also, it should be addressed that phosphate block copolymer can more significantly inhibit the mineral loss possibly due to its higher binding strengthen onto HAP surface.

(42) TABLE-US-00006 TABLE 6 The effect of different polymers on calcium released level Concentration, Calcium level Treatment g/L pH (%) Blank (PBS buffer) n/a 7.0 90.4 13.6 P(MMA).sub.20-b-P(MOEP).sub.35 1.0 4.2 42.5 7.5 PAA 1.0 4.2 86.1 14.7 Carbopol 1.0 4.2 82.5 12.8 Gantrez 1.0 4.2 98.2 4.9 PMMA-b-PAA (tBE4) 1.0 4.2 65.3 9.4

(43) Another anti erosion test completed for the phosphorylated or carboxylated block copolymers was performed using the pH stat instrument. In this experiment, HAP discs were immersed in 15 ml 0.3% citric acid solution (pH 3.8) for 15 minutes before and after 2-minute treatment. The amount of the 10 mM HCl added over time to keep a pH 3.8 was recorded. The % reduction (anti-erosion efficiency) is calculated as

(44) $\left(1-\frac{\left(\frac{\mathrm{acid}\mathrm{addtion}}{\mathrm{time}}\right)\mathrm{after}}{\left(\frac{\mathrm{acid}\mathrm{addtion}}{\mathrm{time}}\right)\mathrm{before}}\right)*100.$
The higher reduction indicates better protection on erosion. The corresponding results are shown in Table 7. Similar to the findings obtained for the calcium release experiments, the PMAA homopolymer offered almost no protection (0.65%) while the PMMA-b-PAA block copolymers provided greater protection benefits and the PMMA-b-PMOEP block co-polymer provided the greatest benefits, a 30% reduction in erosion. In addition, increasing the molecular weight of the block co-polymer increases the anti-erosion efficacy

(45) TABLE-US-00007 TABLE 7 Anti-erosion efficiency of homo and block co-polymers Concentration, Reduction, Treatment g/L % PAA (MW = 50,000, Polysciences, Inc) 1.00 0.65 (PMMA)69-b-(PAA)198 1.00 15.16 (PMMA)20-b-P(AA)19 1.00 10.00 (PMMA)19-b-(PMOEP)9 1.00 30.37

(46) 6. Anti Erosion Test of Phosphate Block Copolymer in Presence of Fluoride

(47) Since the fluoride ion is widely used in oral care to protect enamel against acid attack, phosphorylated copolymers can greatly enhance the efficiency of this traditional treatment based on the pH stat assessment. It is clearly shown that the anti-erosion efficiency of the mixture of NaF and polymer is increased by 15-30% compared with the copolymer or NaF alone. This result clearly indicates that it's highly promising to enhance the benefits of fluoride when combining with those claimed block copolymer as oral products. Table 8 shows the anti-erosion protection benefits of the PMMA-b-PMOEP block copolymers (1 g/L) in the presence of 500 ppm F.

(48) TABLE-US-00008 TABLE 8 Anti-erosion efficiency of NaF and NaF + PMMA-b-PMOEP Treatment Reduction, % NaF 30.21 NaF + (PMMA).sub.19-b-(PMOEP).sub.9 54.29 (PMMA).sub.19-b-(PMOEP).sub.9 30.37

(49) 7. Surface Morphology

(50) The protective layer that is formed on the enamel surface could prevent the mineral loss as indicated by previous data. This layer could also protect the surface morphology of enamel surface by obstructing the diffusion of external acid. Without any treatment, enamel could be easily etched by acid as shown in FIG. 4. When the surface was treated by phosphate block copolymer first, the surface morphology before and after acid erosion, the tooth surface was largely preserved as shown in FIG. 4-2.