A xhs mu/ Bd. Vol. 21. pp. 165 to 769. Pergamon
Press 1976.Printed in Great
Britain.
A SCANNING ELECTRON MICROSCOPE STUDY OF FACTORS INFLUENCING ETCH PATTERNS OF HUMAN ENAMEL J. E. TYLER M.R.C. Dental Unit, Dental School, Bristol BSl 2LY, England
Summary-Facets were ground and polished in the outer third of the enamel thickness from 25 caries-free human molar teeth to produce enamel sub-surfaces cut perpendicularly to the prism direction. Under static conditions of acid dissolution, using 0.25 M formic, acetic or lactic acid solutions adjusted to pH 4.0, prism cores were preferentially dissolved. The predominant honeycomb acid-etch pattern, which showed an elevation of prism sheaths relative to adjacent unetched enamel surfaces, is interpreted as due to the precipitation of dissolution products. Vigorous agitation of the acid media caused preferential dissolution at prism junctions, a change of acid etch pattern attributed to the breakdown of diffusion barriers and accelerated dissolution rates which would tend to enhance the loss of enamel tissue from sites of greatest porosity, that is, the prism junctions. Therefore, the establishment of diffusion barriers with the potential for recrystallization of dissolution products may be a contributory factor in the formation of the honeycomb topography.
INTRODUCTION The preferential loss of prism core enamel by acidetching and boundary enamel with chelation has been described by Miiller and Schait, 1957; Poole and Johnson, 1967; Hoffman, McEwan and Drew, 1969a, b. Possible reasons for such variations have been given by Johnson, Poole and Tyler (1971), but the correct interpretation of the mechanism for the differential dissolution of enamel is of importance for two reasons. It should aid the appreciation of the structural changes in enamel caries; it is of importance in restorative dentistry because of the need to prepare surfaces for the attachment of sealant resins. Enamel dissolution studies have used static un>,tirred demineralizing systems in which reaction products accumulate at the surface of specimens being etched. The main objective of the experiments de?,cribed here was to discover the effect on etch patterns of the removal of reaction products by stirring. MATERIALSAND
RESULTS
Exposure of cut enamel surfaces to the non-stirred, dilute solutions of lactic acid produced a predominant honeycomb etch pattern as described by Poole and Johnson (1967) (Figs. 1 and 3). However, detailed examination of these specimens showed lines of enamel prisms with a greater depth of prism-core dissolution’compared with that in adjacent areas of honeycomb etch pattern. This modified etch topography was consistently associated with surface scratch marks of the order of l-2pm in depth and 24mm in length, which were fortuitiously produced by polishing enamel with 400 mesh grade alumina. Despite the loss of enamel along the scratches, prism boundary material was elevated and continuous within these marks (Figs. 1 and 3), forming ‘bridges’ to complete the acid honeycomb topography. Similar bridge-type etch patterns were observed using dilute acetic or formic acid, and this effect appeared to be independent of the organic acid used.
METHODS
Twenty-five freshly extracted human molar teeth ‘Merecarefully cleaned with fine pumice powder and 1he roots removed. Facets of enamel, free from caries, ‘vere prepared using a water-cooled diamond disc to l,roduce surfaces cut perpendicularly to the prism di.ection. These facets were polished with graded alu:nina and fractured into two parts. One part being exposed for 10min to a non-stirred 200ml aliquot of 0.25 molar acid test solution containing either formic, acetic or lactic acids adjusted to pH 4.0 with ,;odium hydroxide; the other part was etched under identical conditions, except that the acid was vigor,~sly stirred with a mechanical stirrer. To avoid ,nechanical damage to the enamel surfaces during stirred acid etching, specimens were fixed to glass plates before being placed into the acid test solutions. To compare polished enamel surfaces with adjacent 0.B. 21,12-L,
acid-etched areas, portions of the prepared surfaces were protected with thin films of polystyrene cement which, after acid etching under the prescribed static dissolution conditions, were peeled away in order to expose the un-etched polished enamel. Upon reuniting the etched fractured facets, all specimens were carefully immersed in distilled water for one minute before drying at room temperature and vacuum-coating with a layer of gold-palladium alloy to a thickness of approximately 50nm. Specimens were examined at a 45” tilt angle in a “Stereoscan” scanning electron microscope (Cambridge Instrument Co.), operating at 10 kV. Stereopair micrographs with a rotation of 4” between exposures were taken to assist in the interpretation of the acid-etched enamel surface. A simple mirror stereoscope (Fairey Surveys Ltd.) was used for the visualisation of stereopairs.
765
766
J. E. -ryler
Comparison of etched and polished enamel surfaces protected by polystyrene cement films showed apparent elevation of prism boundary material (Fig. 4) to levels approximating to that of the unaltered enamel surface. Stereopair micrographs (Fig. 5) showed that sections of prism boundary material were raised above the un-etched enamel surface in all specimens examined. Enamel etch patterns produced by stirred acid dissolution (Fig. 2) strongly suggested that prism junction locations were preferentially dissolved leaving the axial parts of the prism cores less affected. In contrast to the accepted honeycomb pattern, the peripheral regions of prisms, particularly those associated with prism ‘heads’ were severely etched compared to the central prism cores. DISCUSSION
To understand’the progress and formation of carious enamel lesions and enamel etch patterns, it is necessary to define those factors which contribute to the differential solubility of enamel. Studies of early enamel lesions by Darling (1958) indicated that the intial acid attack is associated with structural points of entry at the enamel surface related to the striae of Retzius followed by the involvement of “interprismatic” regions and prism cross-striations, with the subsequent loss of prism core material. Poole and Johnson (1967) observed d&ones to be more seriously affected than parazones, so that the general direction of prisms, and consequently the mean direction of enamel crystals, appear to have a role in determining the relative rates of dissolution (Johnson, 1966; Sharpe, 1967; Boyde, 1971; Simmelink, Nygaard and Scott, 1974). Crystal orientation changes abruptly at prism junctions where not -only more inter-crystal space would be present but possible variations in the distribution of organic matter may produce differential dissolution rates (Poole, 1966). Although it is established that the prism junction areas are particularly permeable (Poole, Tailby and Berry, 1963; Linden, 1968), the role of structural features of enamel in dissolution processes is not fully understood, nor is it clear whether a loss of mineral or organic matrix occurs as the initial stage of dental caries. Dilute mineral or organic acids produce the honeycomb type of etch pattern, which may be interpreted as the preferential loss of prism cores. However, this generalization is only true with prolonged dilute acid attack because, with brief dilute acid and concentrated acid etching of enamel, prism peripheries are preferentially dissolved (Boyde, 1969; Johnson et al., 1971; Gwinnett, 1971). The observation that brief exposure of enamel to dilute acids under static dissolution conditions produced narrow clefts within prism junctions is of considerable importance with respect to the pattern of destruction in dental caries. This loss of prism-periphery enamel is one of the earliest detectable ultrastructural features of dental caries (Gray and Francis, 1963; Johnson, 1967), and both polarized light and microradiographic studies of the advancing front of the carious lesions show loss of tissue at prism junctions (Darling, 1958; Silverstone, 1966). Prism cleft formation, regarded as the first stage of acid dissolution of en-
amel, may be related to the ease of penetration of etchants into the enhanced porous structure of prism peripheries and the selective loss of enamel from these sites (Johnson et al., 1971). Later stages of dilute acid dissolution of enamel, or advanced dental caries, are characterized by honeycomb topography, where prism cores have apparently dissolved faster than boundary enamel. Attempts to explain this differential solubility of enamel have relied upon static features of structure and composition, and little consideration has been given to the possibility that dynamic features of dissolution, such as diffusion processes (Holly and Gray, 1968; Gray, 1966), may contribute to the etch pattern. Dissolution of ionic crystals was considered by Napper and Smythe (1966) to occur in two consecutive stages. The initial process involved the disengagement of solute ions from steps in the crystal surface followed by a second stage concerned with the transfer of iqns into the bulk of the dissolution medium by diffusion and convection. They believed that mass transport processes are rate-limiting and, therefore, in unstirred systems where dissolution products accumulate, the dissolution rate becomes diffusion-controlled. However, in a stirred system which markedly facilitates the mass transfer of dissolution products, ionic disengagement from crystal surfaces becomes the principal rate-limiting process and, as a consequence, the dissolution rate becomes controlled by surface area availability. Considering how this applies to different etching systems it is likely that, because of the enhanced porosity, all etching agents preferentially penetrate prism junctions, where disengagement of ions begins. With ethylene diamine tetra-acetic acid, released calcium is continuously bound as a chelate complex and, with stirred dilute acids, calcium is continuously removed by rapid diffusion and convection. In both cases, therefore, diffusion barriers are never created and the etch patterns are determined solely by the relative areas of enamel exposed to the etchant at different sites. As the surface area in the junctions are greater than the area of the etched ends of the prisms, preferential dissolution continues in the junctions to produce rod-shaped elements. However, in the case of unstirred dilute acids, the initial preferential dissolution within junctions would slow and perhaps cease because the limited diffusion of ions causes an accumulation of reaction products with possible re-precipitation of various phosphates. If so, the dissolution at the cut ends of the prisms would overtake that at junctions. Nevertheless, the problem of the survival of the prism junction remains. Johansen (1963) has proposed that survival could be due to greater concentrations of certain elements or of organic material, which reduce the solubility of the prism peripheries. The findings described here, suggest the possibility that another important factor is the reprecipitation of mineral in the prism junctions. On acid-etched surfaces, there appears to be an actual elevation of prism sheaths above the original level of the cut surface and foreign crystals, seen in electron micrographs (Johnson, 1967), appear to persist until the last stages of both acid dissolution and natural caries. With solubility isotherms and carious enamel pyrolyses, Brown, Pate1 and Chow (1975) identified
Etch patterns of human enamel dicalcium phosphate dihydrate in enamel treated with dilute acids and showed that at least three per cent of the phosphate in carious enamel was this calcium compound. Concentrated acid-etching of enamel, the current preliminary conditioning process prior to the application of fissure sealants or resins, produces a variable etch topography as shown by the coexistence of prism cleft formation and loss of prism cores (Gwinnett, 1971; Silverstone et al., 1975), and the change of structure associated with surface enamel may contribute to etch pattern irregularity. The possibility exists that surface deposition of extraneous calcium phosphates [Chow and Brown, 1973), as mentioned, may interfere with the enamel-adhesive interface and so produce ;t mechanically weaker bond. Acknowledgements-I am grateful to the staff of the Long .ishton Research Station Stereoscan Unit-for their assist.mce with the scanning electron microscopy and for advice given by Dr. D. F. G. Poole.
REFERENCES goyde A. 1969. Electron microscopic observations relating to the nature and development of prism decussation in mammalian dental enamel. Bull. Grpmt int. Rech. scient. Stomat. 12, 151-207. Boyde A. 1971. Second fnr. Symp. on Tooth Enamel, London 1969 (Edited by Fearnhead R. W. and Stack M. V.) pp. 50 Wright, Bristol. Brown W. E.. Pate1 P. R. and Chow L. C. 1975. Formation of CaHPd,.2H20 from enamel mineral and its relationship to caries mechanism. J. dent. Res. 54, 471-481. “how L. C. and Brown W. E. 1973. Phosphoric acid conditioning of teeth for pit and fissure sealants. J. dent. Res. 52, 1158. Darling A. I. 1958. Studies on the early lesion of enamel caries, its nature, mode of spread and points of entry. Br. dent. J. 105, 119-135. Gray J. A. and Francis M. D. 1963. Physical chemistry of enamel dissolution. In: Mechanisms of Hard Tissue Destruction (Edited by Sognnaes R. F.) pp. 213-260. A.A.A.S., Washington. Gray J. A. 1966. Kinetics of enamel dissolution during formation of incipient caries-like lesions. Archs oral Biol. 11, 397421. Gwinnett A. J. 1971. Histologic changes in human enamel following treatment with acidic adhesive conditioning agents. Archs oral Biol. 16, 731-738. Hoffman S., McEwan W. S. and Drew C. M. 1969a. Scan-
767
ning electron microscope studies of dental enamel. J. dent. Res. 48, 242-250. Hoffman S., McEwan W. S. and Drew C. M. 1969b. Scanning electron microscope studies of EDTA-treated enamel. J. dent. Res. 48, 12341242. Holly F. J. and Gray J. A. 1968. Mechanism for incipient carious lesion growth utilizing a physical model based on diffusion concepts. Archs oral Biol. 13, 319-334. Johansen E. 1963. Ultrastructural and chemical observations on dental caries. In: Mechanism of Hard Tissue Destruction (Edited by Sognnaes R. F.) pp. 187-211. A.A.A.S., Washington. Johnson N. W. 1966. Differences in the shape of human enamel crystallites after partial destruction by caries. EDTA and various acids. Archs oral Biol. 11, 1421-1424. Johnson N. W. 1967. Some aspects of the ultrastructure of early human enamel caries seen with the electron microscope. Archs oral Biol. 12, 1505-1521. Johnson N. W., Poole D. F. G. and Tyler J. E. 1971. Factors affecting the differential dissolution of human enamel in acid and EDTA. A scanning electron microscope study. Archs oral Biol. 16, 385-396. LindCn L. A. 1968. Microscopic observations of fluid flow through enamel in vitro. Odont. Revy 9, 349-367. Miiller G. and Schait A. 1957. Morphologic differences in replicas of intact enamel decalcified in acid or EDTA. H>lv. odonr. Acta 1, 5-8. Napper D. H. and Smithe B. M. 1966. The dissolution kinetics of hydroxyapatite in the presence of kink poisons. J. dent. Res. 45, 1775-1783. Poole D. F. G. 1966. The use of the microscope in dental research. Br. dent. J. 121, 71-79. Poole D. F. G. and Johnson N. W. 1967. The effects of different demineralizing agents on human enamel surfaces studied by scant&g electron microscopy. Archs oral Biol. 12. 1621-1634. Poole D. F. G:, Tailby P. W. and Berry D. C. 1963. The movement of water and other molecules through human enamel. Archs oral Biol. 8, 771-772. Sharpe A. N. 1967. Influence of crystal orientation in human enamel on its reactivity to acid as shown by high resolution microradiography. Archs oral Biol. 12, 581-591.
Silverstone L. M. 1966. The primary translucent zone of enamel caries and of artifical caries-like lesions. Br. dent. J. 120, 461471. Silverstone L. M., Saxton C. A., Dogon, I. L. and Fejerskov 0. 1975. Variation in the pattern of acid etching of human dental enamel examined by scanning electron microscopy. Caries Res. 9, 373-387. Simmelink J. W., Nygaard V. K. and Scott D. B. 1974. Theory for the sequence of human and rat enamel dissolution- by acid and by EDTA: A correlated scanning and transmission electron microscope study. Archs orul Biol. 19, 183-197.
768
J. E. Tyler
Fig. 1. Control specimen of sound for ten minutes with 0.25 molar preferential prism core dissolution of prism boundary material across
Plate 1 human enamel surface cut transverse to prism direction and etched lactic acid adjusted to pH 4.0. The honeycomb structure due to is at an early stage of formation and illustrates the continuation scratch marks (approximately 1 pm wide and deep) produced during specimen preparation. x 1350
Fig. 2. Portion of the same enamel specimen as in Fig. 1, etched under the same conditions but with vigorous agitation of the acid dissolution media. A reverse etch pattern to that of the honeycomb indicates preferential dissolution occurring at peripheral areas of the prisms. x 1350 Fig. 3. Honeycomb-etch pattern produced under static dissolution conditions identical to Fig. 1, illustrating prism boundary material ‘bridging’ a cross-shaped scratch mark. x 2400 Fig. 4. Polystyrene film protection of a section of polished enamel (left) shows, after static acid dissolution and removal of this film, the relationship of the honeycomb topography to surface scratch marks. Continuation of prism peripheries are observed despite the initial mechanical loss of enamel within scratch marks. x 2400 Fig. 5. Stereopair micrographs of boundary areas created on polished enamel surfaces by etching as in Fig. 4, showing the depth of acid etch and the elevation of parts of the honeycomb topography slightly above that of the unetched enamel surface. x 5000.
Etch patterns of human enamel
Plate 1
169
Press 1976.Printed in Great
Britain.
A SCANNING ELECTRON MICROSCOPE STUDY OF FACTORS INFLUENCING ETCH PATTERNS OF HUMAN ENAMEL J. E. TYLER M.R.C. Dental Unit, Dental School, Bristol BSl 2LY, England
Summary-Facets were ground and polished in the outer third of the enamel thickness from 25 caries-free human molar teeth to produce enamel sub-surfaces cut perpendicularly to the prism direction. Under static conditions of acid dissolution, using 0.25 M formic, acetic or lactic acid solutions adjusted to pH 4.0, prism cores were preferentially dissolved. The predominant honeycomb acid-etch pattern, which showed an elevation of prism sheaths relative to adjacent unetched enamel surfaces, is interpreted as due to the precipitation of dissolution products. Vigorous agitation of the acid media caused preferential dissolution at prism junctions, a change of acid etch pattern attributed to the breakdown of diffusion barriers and accelerated dissolution rates which would tend to enhance the loss of enamel tissue from sites of greatest porosity, that is, the prism junctions. Therefore, the establishment of diffusion barriers with the potential for recrystallization of dissolution products may be a contributory factor in the formation of the honeycomb topography.
INTRODUCTION The preferential loss of prism core enamel by acidetching and boundary enamel with chelation has been described by Miiller and Schait, 1957; Poole and Johnson, 1967; Hoffman, McEwan and Drew, 1969a, b. Possible reasons for such variations have been given by Johnson, Poole and Tyler (1971), but the correct interpretation of the mechanism for the differential dissolution of enamel is of importance for two reasons. It should aid the appreciation of the structural changes in enamel caries; it is of importance in restorative dentistry because of the need to prepare surfaces for the attachment of sealant resins. Enamel dissolution studies have used static un>,tirred demineralizing systems in which reaction products accumulate at the surface of specimens being etched. The main objective of the experiments de?,cribed here was to discover the effect on etch patterns of the removal of reaction products by stirring. MATERIALSAND
RESULTS
Exposure of cut enamel surfaces to the non-stirred, dilute solutions of lactic acid produced a predominant honeycomb etch pattern as described by Poole and Johnson (1967) (Figs. 1 and 3). However, detailed examination of these specimens showed lines of enamel prisms with a greater depth of prism-core dissolution’compared with that in adjacent areas of honeycomb etch pattern. This modified etch topography was consistently associated with surface scratch marks of the order of l-2pm in depth and 24mm in length, which were fortuitiously produced by polishing enamel with 400 mesh grade alumina. Despite the loss of enamel along the scratches, prism boundary material was elevated and continuous within these marks (Figs. 1 and 3), forming ‘bridges’ to complete the acid honeycomb topography. Similar bridge-type etch patterns were observed using dilute acetic or formic acid, and this effect appeared to be independent of the organic acid used.
METHODS
Twenty-five freshly extracted human molar teeth ‘Merecarefully cleaned with fine pumice powder and 1he roots removed. Facets of enamel, free from caries, ‘vere prepared using a water-cooled diamond disc to l,roduce surfaces cut perpendicularly to the prism di.ection. These facets were polished with graded alu:nina and fractured into two parts. One part being exposed for 10min to a non-stirred 200ml aliquot of 0.25 molar acid test solution containing either formic, acetic or lactic acids adjusted to pH 4.0 with ,;odium hydroxide; the other part was etched under identical conditions, except that the acid was vigor,~sly stirred with a mechanical stirrer. To avoid ,nechanical damage to the enamel surfaces during stirred acid etching, specimens were fixed to glass plates before being placed into the acid test solutions. To compare polished enamel surfaces with adjacent 0.B. 21,12-L,
acid-etched areas, portions of the prepared surfaces were protected with thin films of polystyrene cement which, after acid etching under the prescribed static dissolution conditions, were peeled away in order to expose the un-etched polished enamel. Upon reuniting the etched fractured facets, all specimens were carefully immersed in distilled water for one minute before drying at room temperature and vacuum-coating with a layer of gold-palladium alloy to a thickness of approximately 50nm. Specimens were examined at a 45” tilt angle in a “Stereoscan” scanning electron microscope (Cambridge Instrument Co.), operating at 10 kV. Stereopair micrographs with a rotation of 4” between exposures were taken to assist in the interpretation of the acid-etched enamel surface. A simple mirror stereoscope (Fairey Surveys Ltd.) was used for the visualisation of stereopairs.
765
766
J. E. -ryler
Comparison of etched and polished enamel surfaces protected by polystyrene cement films showed apparent elevation of prism boundary material (Fig. 4) to levels approximating to that of the unaltered enamel surface. Stereopair micrographs (Fig. 5) showed that sections of prism boundary material were raised above the un-etched enamel surface in all specimens examined. Enamel etch patterns produced by stirred acid dissolution (Fig. 2) strongly suggested that prism junction locations were preferentially dissolved leaving the axial parts of the prism cores less affected. In contrast to the accepted honeycomb pattern, the peripheral regions of prisms, particularly those associated with prism ‘heads’ were severely etched compared to the central prism cores. DISCUSSION
To understand’the progress and formation of carious enamel lesions and enamel etch patterns, it is necessary to define those factors which contribute to the differential solubility of enamel. Studies of early enamel lesions by Darling (1958) indicated that the intial acid attack is associated with structural points of entry at the enamel surface related to the striae of Retzius followed by the involvement of “interprismatic” regions and prism cross-striations, with the subsequent loss of prism core material. Poole and Johnson (1967) observed d&ones to be more seriously affected than parazones, so that the general direction of prisms, and consequently the mean direction of enamel crystals, appear to have a role in determining the relative rates of dissolution (Johnson, 1966; Sharpe, 1967; Boyde, 1971; Simmelink, Nygaard and Scott, 1974). Crystal orientation changes abruptly at prism junctions where not -only more inter-crystal space would be present but possible variations in the distribution of organic matter may produce differential dissolution rates (Poole, 1966). Although it is established that the prism junction areas are particularly permeable (Poole, Tailby and Berry, 1963; Linden, 1968), the role of structural features of enamel in dissolution processes is not fully understood, nor is it clear whether a loss of mineral or organic matrix occurs as the initial stage of dental caries. Dilute mineral or organic acids produce the honeycomb type of etch pattern, which may be interpreted as the preferential loss of prism cores. However, this generalization is only true with prolonged dilute acid attack because, with brief dilute acid and concentrated acid etching of enamel, prism peripheries are preferentially dissolved (Boyde, 1969; Johnson et al., 1971; Gwinnett, 1971). The observation that brief exposure of enamel to dilute acids under static dissolution conditions produced narrow clefts within prism junctions is of considerable importance with respect to the pattern of destruction in dental caries. This loss of prism-periphery enamel is one of the earliest detectable ultrastructural features of dental caries (Gray and Francis, 1963; Johnson, 1967), and both polarized light and microradiographic studies of the advancing front of the carious lesions show loss of tissue at prism junctions (Darling, 1958; Silverstone, 1966). Prism cleft formation, regarded as the first stage of acid dissolution of en-
amel, may be related to the ease of penetration of etchants into the enhanced porous structure of prism peripheries and the selective loss of enamel from these sites (Johnson et al., 1971). Later stages of dilute acid dissolution of enamel, or advanced dental caries, are characterized by honeycomb topography, where prism cores have apparently dissolved faster than boundary enamel. Attempts to explain this differential solubility of enamel have relied upon static features of structure and composition, and little consideration has been given to the possibility that dynamic features of dissolution, such as diffusion processes (Holly and Gray, 1968; Gray, 1966), may contribute to the etch pattern. Dissolution of ionic crystals was considered by Napper and Smythe (1966) to occur in two consecutive stages. The initial process involved the disengagement of solute ions from steps in the crystal surface followed by a second stage concerned with the transfer of iqns into the bulk of the dissolution medium by diffusion and convection. They believed that mass transport processes are rate-limiting and, therefore, in unstirred systems where dissolution products accumulate, the dissolution rate becomes diffusion-controlled. However, in a stirred system which markedly facilitates the mass transfer of dissolution products, ionic disengagement from crystal surfaces becomes the principal rate-limiting process and, as a consequence, the dissolution rate becomes controlled by surface area availability. Considering how this applies to different etching systems it is likely that, because of the enhanced porosity, all etching agents preferentially penetrate prism junctions, where disengagement of ions begins. With ethylene diamine tetra-acetic acid, released calcium is continuously bound as a chelate complex and, with stirred dilute acids, calcium is continuously removed by rapid diffusion and convection. In both cases, therefore, diffusion barriers are never created and the etch patterns are determined solely by the relative areas of enamel exposed to the etchant at different sites. As the surface area in the junctions are greater than the area of the etched ends of the prisms, preferential dissolution continues in the junctions to produce rod-shaped elements. However, in the case of unstirred dilute acids, the initial preferential dissolution within junctions would slow and perhaps cease because the limited diffusion of ions causes an accumulation of reaction products with possible re-precipitation of various phosphates. If so, the dissolution at the cut ends of the prisms would overtake that at junctions. Nevertheless, the problem of the survival of the prism junction remains. Johansen (1963) has proposed that survival could be due to greater concentrations of certain elements or of organic material, which reduce the solubility of the prism peripheries. The findings described here, suggest the possibility that another important factor is the reprecipitation of mineral in the prism junctions. On acid-etched surfaces, there appears to be an actual elevation of prism sheaths above the original level of the cut surface and foreign crystals, seen in electron micrographs (Johnson, 1967), appear to persist until the last stages of both acid dissolution and natural caries. With solubility isotherms and carious enamel pyrolyses, Brown, Pate1 and Chow (1975) identified
Etch patterns of human enamel dicalcium phosphate dihydrate in enamel treated with dilute acids and showed that at least three per cent of the phosphate in carious enamel was this calcium compound. Concentrated acid-etching of enamel, the current preliminary conditioning process prior to the application of fissure sealants or resins, produces a variable etch topography as shown by the coexistence of prism cleft formation and loss of prism cores (Gwinnett, 1971; Silverstone et al., 1975), and the change of structure associated with surface enamel may contribute to etch pattern irregularity. The possibility exists that surface deposition of extraneous calcium phosphates [Chow and Brown, 1973), as mentioned, may interfere with the enamel-adhesive interface and so produce ;t mechanically weaker bond. Acknowledgements-I am grateful to the staff of the Long .ishton Research Station Stereoscan Unit-for their assist.mce with the scanning electron microscopy and for advice given by Dr. D. F. G. Poole.
REFERENCES goyde A. 1969. Electron microscopic observations relating to the nature and development of prism decussation in mammalian dental enamel. Bull. Grpmt int. Rech. scient. Stomat. 12, 151-207. Boyde A. 1971. Second fnr. Symp. on Tooth Enamel, London 1969 (Edited by Fearnhead R. W. and Stack M. V.) pp. 50 Wright, Bristol. Brown W. E.. Pate1 P. R. and Chow L. C. 1975. Formation of CaHPd,.2H20 from enamel mineral and its relationship to caries mechanism. J. dent. Res. 54, 471-481. “how L. C. and Brown W. E. 1973. Phosphoric acid conditioning of teeth for pit and fissure sealants. J. dent. Res. 52, 1158. Darling A. I. 1958. Studies on the early lesion of enamel caries, its nature, mode of spread and points of entry. Br. dent. J. 105, 119-135. Gray J. A. and Francis M. D. 1963. Physical chemistry of enamel dissolution. In: Mechanisms of Hard Tissue Destruction (Edited by Sognnaes R. F.) pp. 213-260. A.A.A.S., Washington. Gray J. A. 1966. Kinetics of enamel dissolution during formation of incipient caries-like lesions. Archs oral Biol. 11, 397421. Gwinnett A. J. 1971. Histologic changes in human enamel following treatment with acidic adhesive conditioning agents. Archs oral Biol. 16, 731-738. Hoffman S., McEwan W. S. and Drew C. M. 1969a. Scan-
767
ning electron microscope studies of dental enamel. J. dent. Res. 48, 242-250. Hoffman S., McEwan W. S. and Drew C. M. 1969b. Scanning electron microscope studies of EDTA-treated enamel. J. dent. Res. 48, 12341242. Holly F. J. and Gray J. A. 1968. Mechanism for incipient carious lesion growth utilizing a physical model based on diffusion concepts. Archs oral Biol. 13, 319-334. Johansen E. 1963. Ultrastructural and chemical observations on dental caries. In: Mechanism of Hard Tissue Destruction (Edited by Sognnaes R. F.) pp. 187-211. A.A.A.S., Washington. Johnson N. W. 1966. Differences in the shape of human enamel crystallites after partial destruction by caries. EDTA and various acids. Archs oral Biol. 11, 1421-1424. Johnson N. W. 1967. Some aspects of the ultrastructure of early human enamel caries seen with the electron microscope. Archs oral Biol. 12, 1505-1521. Johnson N. W., Poole D. F. G. and Tyler J. E. 1971. Factors affecting the differential dissolution of human enamel in acid and EDTA. A scanning electron microscope study. Archs oral Biol. 16, 385-396. LindCn L. A. 1968. Microscopic observations of fluid flow through enamel in vitro. Odont. Revy 9, 349-367. Miiller G. and Schait A. 1957. Morphologic differences in replicas of intact enamel decalcified in acid or EDTA. H>lv. odonr. Acta 1, 5-8. Napper D. H. and Smithe B. M. 1966. The dissolution kinetics of hydroxyapatite in the presence of kink poisons. J. dent. Res. 45, 1775-1783. Poole D. F. G. 1966. The use of the microscope in dental research. Br. dent. J. 121, 71-79. Poole D. F. G. and Johnson N. W. 1967. The effects of different demineralizing agents on human enamel surfaces studied by scant&g electron microscopy. Archs oral Biol. 12. 1621-1634. Poole D. F. G:, Tailby P. W. and Berry D. C. 1963. The movement of water and other molecules through human enamel. Archs oral Biol. 8, 771-772. Sharpe A. N. 1967. Influence of crystal orientation in human enamel on its reactivity to acid as shown by high resolution microradiography. Archs oral Biol. 12, 581-591.
Silverstone L. M. 1966. The primary translucent zone of enamel caries and of artifical caries-like lesions. Br. dent. J. 120, 461471. Silverstone L. M., Saxton C. A., Dogon, I. L. and Fejerskov 0. 1975. Variation in the pattern of acid etching of human dental enamel examined by scanning electron microscopy. Caries Res. 9, 373-387. Simmelink J. W., Nygaard V. K. and Scott D. B. 1974. Theory for the sequence of human and rat enamel dissolution- by acid and by EDTA: A correlated scanning and transmission electron microscope study. Archs orul Biol. 19, 183-197.
768
J. E. Tyler
Fig. 1. Control specimen of sound for ten minutes with 0.25 molar preferential prism core dissolution of prism boundary material across
Plate 1 human enamel surface cut transverse to prism direction and etched lactic acid adjusted to pH 4.0. The honeycomb structure due to is at an early stage of formation and illustrates the continuation scratch marks (approximately 1 pm wide and deep) produced during specimen preparation. x 1350
Fig. 2. Portion of the same enamel specimen as in Fig. 1, etched under the same conditions but with vigorous agitation of the acid dissolution media. A reverse etch pattern to that of the honeycomb indicates preferential dissolution occurring at peripheral areas of the prisms. x 1350 Fig. 3. Honeycomb-etch pattern produced under static dissolution conditions identical to Fig. 1, illustrating prism boundary material ‘bridging’ a cross-shaped scratch mark. x 2400 Fig. 4. Polystyrene film protection of a section of polished enamel (left) shows, after static acid dissolution and removal of this film, the relationship of the honeycomb topography to surface scratch marks. Continuation of prism peripheries are observed despite the initial mechanical loss of enamel within scratch marks. x 2400 Fig. 5. Stereopair micrographs of boundary areas created on polished enamel surfaces by etching as in Fig. 4, showing the depth of acid etch and the elevation of parts of the honeycomb topography slightly above that of the unetched enamel surface. x 5000.
Etch patterns of human enamel
Plate 1
169
