172
Toxicity of silica-containing calcium phosphate glasses demonstrated in mice Mitsuo Nagase, Yoshihiro Abe, Masaki Chigira and Eiichi Udagawa Department Department
of Orthopaedic Surgery, Gunma University School of Medicine, Maebashi, Japan; and of Materials Science and Engineering, Nagoya institute of Technology, Nagoya, Japan
Suspensions of calcium phosphate glass containing various concentrations of silica {glass composition (moles): 100 Ca(PO,), to x SiO,,x = 0, 5, 10, 15 or 40) dispersed in normal saline were injected intraperitoneaily into C57BL16 mice to determine the mortality within 30 days. The mortality was O/10, 3/10, g/10, lo/10 and lOI16 at x = 0, 5, 10, 15 and 40 mol of silica, respectively. By means of inductively coupled plasma analysis, the amount of dissolved silica (Si4+) in water at 37°C from the calcium phosphate glass depended on the amount of silica in the glasses. The mortality of mice was directly proportional to the silica content of the glass injected intraperitoneally. These results clearly show that the dissolved silica (Si4’) from the glass, monomeric or low molecular silicic anion, is highly toxic. The SiO, component in
biomaterials has toxic potential when dissolved in the body. Keywords:
Glasses, toxicity,
silicon
Received 31 January 1991; revised 6 March 1991; accepted 20 May 1991
Hench first developed a bioactive glass, bioglassa, of composition SiOZ (45 wtW), CaO (24.5 wtW), Na,O (24.5 wt%), PZO, (6 wt%), for bone-graft substitutes*. Since then many silicate glasses, ceramics and glassceramics containing calcium and phosphorus have been developed ‘* 3. It is important to know whether dissolved SiO, exhibits toxicity or not. Our studies in 1987 showed for the first time that fine powders < 10 pm of the silicate glass-ceramics containing calcium and phosphorus, apatite-wollastonite glass ceramic (SiO,:34.2 wt%) had lethal effects on C57BL16 (H-Zb), C3H (H-Zk) and A/J (H-2’) mice*. However, the mechanism of the toxicity and what component of the silica-containing glassceramic is toxic remained unclear. The purpose of this study is to determine the relationship of the toxicity to the silica content in the calcium phosphate glasses.
quenching rate. The fine powders of these five kinds of calcium phosphate glass and pure silica glass itself (SiO, = 100%) were evaluated simultaneously. The protein content of these particles was checked by the direct Ninhydrin reaction and Bradford’s method using Coomassie Brilliant Blue5. No trace of protein appeared. These particles were dissimilar and irregular in shape. same
Particle measurements Particle sizes were determined using a particle size analyser (Horiba, Model CAPA-500). The surface areas were measured with a Quantasorb surface area analyser (Quantachrome Corp., Model MS-8). One-point BrunauerEmmett-Teller (B.E.T.) surface area determinations were made using nitrogen (adsorbate) in helium (carrier) gas mixtures.
Animals MATERIALS AND METHODS Materials For this study, we prepared five types of calcium phosphate glasses (CaO/P,O, = 1) in which 100 moles of Ca(PO,), was combined with x moles of SiOZ (x = 0, 5, lo,15 or 40 mol of silica). We tried to make glasses with x > 40 but clear glasses were not obtained owing to the higher melting temperatures with increasing SiO,. All those specimens were obtained from the melts with the Correspondence to Dr Mitsuo Nagase, Department of Orthopaedic Surgery, Gunma School of Medicine, 3-39-22 Showo, Maebishi, Gunma, 371 Japan. Biomaterials
1992, Vol. 13
No. 3
The Gbweek-old male C57BL/6 mice used in this study were acclimatized in our animal care facility for one week prior to use. They were housed ten per cage and maintained under controlled temperatu~, lighting and relative humidity. Animals were allowed free access to water and fed normal chow.
Animal experimental
protocol
The adult mice were exposed to single doses of silica in the range 0-40 mol in calcium phosphate glass. Ten animals of each group received each of these doses. Animals were given an intraperitoneal dose of glass powder (5000 mglkg body wt) dispersed in 1.0 ml of normal saline. All of the animals were studied simulD 1992 Bu~e~o~h-Hei~mann 0142-~12/92/030172-~
Ltd
glass: M. Nagase et al.
Toxicity of silica-containing
taneously and were dosed intraperitoneally same single glass stock suspensions.
Quantitative
determination
of dissolved
from
173 100 Co ( P03& to X SiO,
the
X
ions
IO3
The amount of dissolved Si4+, Ca2+ and P5+ from the glass specimen in water was quantitatively determined using an inductively coupled plasma (ICP) analyser (Shimadzu, ICPS-500). A plate-like specimen of a sample of glass (surface area = 2 cm’) was used. The dissolved ions were determined after immersing the specimen in 300 ml of distilled water at 37°C for 50 h.
Mortality Intraperitoneal administration of calcium phosphate glass containing various concentrations of silica resulted in an increase in the mortality with increasing amounts of silica in the glass. The mortality was O/10, 3/10, 9/10, lo/10 and lo/10 at x = 0, 5, 10, 15 and 40 mol of silica, respectively (Figure 1). The mean time of death was 68.0, 29.3, 19.2 and 6.0 h after treatment at x = 5, 10, 15 and 40 mol of silica, respectively (Figure 1). The mortality in mice given pure silica glass powder itself was O/10. In contrast to the mortality with silica glass itself or calcium phosphate glass without silica, the silica in calcium phosphate glass decreased the mean time of death and increased the number of deaths within 30 days.
Quantitative
determination
of dissolved
ions
The dissolved amount of Si4+, Ca2+ and P5+ in the water from the calcium phosphate glasses increased with the increasing amount of silica in the glass (Figure 2). No dissolved Si4’, Cazt or Pst was detected by ICP after 50 h of treatment of pure silica glass [SiO,: molar fraction = 1) in water.
I.-
T102 9 5 E Z P Z 51 .u, 10 n
RESULTS
-
I
I 1
0.1
Ca(P4)2 Figure2
’
1
I
0
0.3
0.2
aI.0 SiO,
Molar fraction
Amount of dissolved ions in water at 37°C for 50 h
versus. glass composition. The dissolution of ions into the water depended on the glass composition. (0) Ca2+;( A) Si4+.
Effect of particle
P5+; (0)
measurements
The differences in grain sizes or surface areas among samples did not affect the amounts of dissolved ions from the samples. The diameters of grains of materials used in this experiment were 3.9-8.3 pm (lable 1). The surface areas of the materials were 2.3-3.2 m’/g, except for the pure silica glass itself, which was 13.0 m2/g (7Me 1).
DISCUSSION Silicon is, next to oxygen, the most abundant element in the earth’s crust. Although only trace amounts appear in most animal tissues, silicon is an essential element in the chick” and the rat’, required for normal growth and development. Silicon may function as a biological crosslinking agent and contribute to the architecture and resilience of connective tissue’. The ingestion of a small amount of amorphous silica (silicon dioxide] powder appears to be completely harmless. However, when Table1
24
48
72
96
120
144
168
Time (h)
Figure1 The mortality in C57BL/6 mice given a single intraperitoneal dose of the glass powders. Administration of calcium phosphate glass containing increasing concentrations of silica increased the mortality depending on the amount of silica in the glass. (Cl) 0 mol; (4) 5 mol; (A) 10 mol; (0) 15 mol; (0) 40 mol; (W) SiO,.
Grain sizes and surface areas
Materials*
Grain sizes (pm)
Surface areas (m2/g)
0 mol 5 mol 10 mol 15 mol 40 mol SiO,
5.9 4.3 3.9 4.3 6.3 4.1
3.2 2.3 3.0 2.5 3.1 13.0
*O,5,lO. 15 and 40 mol = calcium phosphate glass (Ca0/P20, = 1) with the composition of 100 Ca(P03)2 to x 90, in moles in which x = 0, 5, 10, 15 or 40 mol of silica, and SiO, = pure silica glass itself.
Biomaterials
1992, Vol. 13 No. 3
174
silica enters the body in any other way than by ingestion, some type of toxicity almost invariably resultsg. We showed that a calcium phosphate glass-ceramic containing 34.2 wt% of silica was lethal in C57BLl6 and A/J mice4. The LD,, of the silica containing glass-ceramic after intraperitoneal administration in C57BL/6 mice was 750 mg/kg .(Ref. 4). The mice died from 2 to 6 days after administration of the glass-ceramic4. Significant footpad swelling occurred for a week after challenge in mice challenged with the glass-ceramic4. The lethality of the material administered weekly did not change during 5 weeks4. A special kind of inflammation, not an immunological one4, plays an important role in the lethal reaction. The present study was undertaken to examine the intraperitoneal toxicity of calcium phosphate glass containing various concentrations of silica in C57BLl6 mice. Calcium phosphate glasses containing large amounts of silica are extremely toxic. The Si4+, Ca2+ and P5+ in water dissolved from the calcium phosphate glass depends on the amount of silica in the glasses, but the amounts of Ca2+ and P5+ were much lower than the amounts normally existing in serum. The dissolved silica (Si4+) in this study may be monomeric or may not be highly polymerized lo . It is suggested that the concentration of monomeric or low molecular silicic anions determines the toxicity of the materials, but the total content of non-dissolved silica had no relation to its toxicity. In determining the biological consequences of dissolved or monomeric silica, what kind of reaction may play an important role in the lethal reaction will require further study. Under these circumstances it is interesting to speculate on the mechanism of the biological effects of dissolved or monomeric silica. There have been a great many studies to explore the pathogenesis of silicosis’1-‘6. But almost all silicosis researchers studied the biological reactions of various forms of solid silica, e.g. crystalline silica [quartz, tridymite, cristobalite, coesite and stishovite)16, silica powder without an organophilic hydrophobic monolayer of hydrocarbon groups15, silica gelI and the the finest amorphous silica *I . Nobody has investigated toxicity of monomeric or low molecular silicic anions dissolved from composition materials. We showed in this study that there was no toxicity in pure silica (SiO,) glass or in calcium phosphate (Ca[PO,),) glass without silica. Monomeric silicic anions dissolved from silica-containing glasses may induce the toxicity. The difference in toxicity among the various forms of solid silica may depend on the amounts of monomeric silica dissolved from the silica, The difference in the mean time of death (6.0-68.0 h after administration in this study, 75.9 h after administration in our previous study41 may depend on the different rates at which monomeric silica is dissolved from composition materials. Silica is also released from ‘bioglass’, and the dissolved amount depends on the glass composition’7. It is hoped that future researchers and engineers will be mindful of this toxicity due to silica dissolved in calcium phosphate glass. Note also that we have not addressed any variation in sensitivity to silica with respect to age, sex, diet, route of exposure, or treatment regimen. These factors may also affect the 36day mortality in mice. The marked physiological response at the high doses Biomaterials 1992. Vol. 13 No. 3
Toxicity
of silica-containing
glass: M. Nagase et al.
of silica may well be symptomatic of the detrimental biochemical changes that follow silica exposure, even long after implantation, since glasses and ceramics are susceptible to both general corrosion and pitting attack in aqueous solutions, and silica is 20 times more soluble in organisms than in saline solution”. Several types of glass-ceramic in systems with calcium phosphate without silica have been developed’” lg. That kind of glassceramic would be expected to be much better for implant systems than silica-containing glass-ceramics although the former have some disadvantages such as the weak mechanical strength in water and low chemical stability.
The authors are grateful to Dr Hiroyasu Takeuchi, Research and Development Center, Mitsubishi Material Co., Ltd for his assistance in measurements of grain size and surface area; and Mr Kiichi Oosawa, Department of Orthopaedic Surgery, Gunma University School of Medicine for his assistance in photography. This work was supported in part by Grant of Japan Sports Medicine Foundation Inc. No. 1.
REFERENCES 1
Hench, L.L., Splinter, R. J., Allen, W.C. and Greenlee, T.K. Bonding mechanism at the interface of ceramic prosthetic materials, 1. Biomed. Mater. Res. Symp.
2
Blenke, B.A., Pfeil, E., Bromer, H. and Kas, H.H., Implantate aus Glas-Keramik in der Knochenchirurgie (Tierexperimentelle Untersuchungen), Langenbechs Arch. Chir. Suppl. Forum 1973, 107-110 Kitsugi, T., Yamamuro, T., Nakamura, T., Higashi, S., Kakutani, Y., Hyakuna, K., Ito, S., Kokubo, T., Takagi, M. and Shibuya, T., Bone bonding behavior of three kinds of apatite containing glass ceramics, I. Biomed.
1971,2,117-141
3
Mater.
4
5
6 7 a
Res.
1986, 28, 1295-1307
Nagase, M., Shimizu, T., Chigira, M. and Udagawa, E., [An analytical study of the inflammation induced by a calcium phosphate ceramic,] Kitakanto Zgaku 1967, 37, 101-106 Bradford, M.M., A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding, Anal. Biocbem. 1976, 72, 246-254 Carlisle, E.M., Silicon: an essential element for the chick, Science 1972, 178, 619-621 Schwartz, K. and Milne, D.B., Growth-promoting effects of silicon in rats, Nature 1972, 239, 333-334 Schwartz, K., A bound form of silicon in glycosaminoglycans and polyuronides, Proc. Natl Acad. Sci. USA 1973, 70, 1608-1612
9
10
Michon, R., Sue, P. and Merinis, J., Metabolisme de la Silice et des Silicates Inhales par l’Anima1, Suiva a 1’Aide de %i. C.R. Acad. Sci. 1956, 243, 2194-2195 Abe, Y., Yoshikawa, Y., Naruse, A. and Saito, H. [Effect of SiO, additive on the rates of crystallization of Mg(PO,), and Ca(PO,), glasses,] I. Ceram. Sot. Jpn 1977, 85.151-153
11
Lungren, K.D., Experimental investigations on the significance of the size of particles in the reaction of the
Toxicity of silica-containing
12
13
14
15
glass: M. Nagase et al.
peritoneum to amorphous silica, Acta Med. Stand. 1953, 145, 84-90 Minova, K. F., Respredelennie Kremniia v Organizme Belykh Krys Pri Eksperimental’ nom Silikoze, Gig. fl. Prof. ZaboZ. 1989, 13,50-51 Oghiso, Y. and Kubota, Y., Interleukin 1 production and accessory cell function of rat alveolar macrophages exposed to mineral dust particles, Microbial. Zmmunol. 1987,31,275-287 Policard, A., Collet, A., Daniel-Moussard, H. and Pregermain, S., Sur les premiers stades des alterations experimentales du rein par l’acide silicique._Etude au microscope electronique, J. Ural. Med. Chir. 1980, 85, 585-800 Schepers, G.W.H., Hypertension due to inhaled sub-
175
18
17
18
19
micron amorphous silica, ‘Ibxicol. Appl. Pharmacol. 1959,1,487 Stalder, K. and Stober, W., Haemolytic activity of suspensions of different silica modifications and inert dusts, Nature 1965,207,874-875 Ravaglioli, A., Krajewski, A., Zini, Q. and Venturi, R., Short and long range silicate release from doped bioglass into 199 medium, Biomaterials 1986,7, 76-78 Abe, Y., Kasuga, T. and Hosono, H., Preparation of highstrength calcium phosphate glass-ceramics by unidirectional crystallization, J. Am. Ceram. Sot. 1984, 67, 142-144 Abe, Y., Hosono, H. and Hosoe, M., Development of calcium phosphate glass-ceramics for surgical and dental use, Phosphorus 81 Sulfur 1987, 30, 337-340
Biomaterials
1992,Vol. 13 No. 3
Toxicity of silica-containing calcium phosphate glasses demonstrated in mice Mitsuo Nagase, Yoshihiro Abe, Masaki Chigira and Eiichi Udagawa Department Department
of Orthopaedic Surgery, Gunma University School of Medicine, Maebashi, Japan; and of Materials Science and Engineering, Nagoya institute of Technology, Nagoya, Japan
Suspensions of calcium phosphate glass containing various concentrations of silica {glass composition (moles): 100 Ca(PO,), to x SiO,,x = 0, 5, 10, 15 or 40) dispersed in normal saline were injected intraperitoneaily into C57BL16 mice to determine the mortality within 30 days. The mortality was O/10, 3/10, g/10, lo/10 and lOI16 at x = 0, 5, 10, 15 and 40 mol of silica, respectively. By means of inductively coupled plasma analysis, the amount of dissolved silica (Si4+) in water at 37°C from the calcium phosphate glass depended on the amount of silica in the glasses. The mortality of mice was directly proportional to the silica content of the glass injected intraperitoneally. These results clearly show that the dissolved silica (Si4’) from the glass, monomeric or low molecular silicic anion, is highly toxic. The SiO, component in
biomaterials has toxic potential when dissolved in the body. Keywords:
Glasses, toxicity,
silicon
Received 31 January 1991; revised 6 March 1991; accepted 20 May 1991
Hench first developed a bioactive glass, bioglassa, of composition SiOZ (45 wtW), CaO (24.5 wtW), Na,O (24.5 wt%), PZO, (6 wt%), for bone-graft substitutes*. Since then many silicate glasses, ceramics and glassceramics containing calcium and phosphorus have been developed ‘* 3. It is important to know whether dissolved SiO, exhibits toxicity or not. Our studies in 1987 showed for the first time that fine powders < 10 pm of the silicate glass-ceramics containing calcium and phosphorus, apatite-wollastonite glass ceramic (SiO,:34.2 wt%) had lethal effects on C57BL16 (H-Zb), C3H (H-Zk) and A/J (H-2’) mice*. However, the mechanism of the toxicity and what component of the silica-containing glassceramic is toxic remained unclear. The purpose of this study is to determine the relationship of the toxicity to the silica content in the calcium phosphate glasses.
quenching rate. The fine powders of these five kinds of calcium phosphate glass and pure silica glass itself (SiO, = 100%) were evaluated simultaneously. The protein content of these particles was checked by the direct Ninhydrin reaction and Bradford’s method using Coomassie Brilliant Blue5. No trace of protein appeared. These particles were dissimilar and irregular in shape. same
Particle measurements Particle sizes were determined using a particle size analyser (Horiba, Model CAPA-500). The surface areas were measured with a Quantasorb surface area analyser (Quantachrome Corp., Model MS-8). One-point BrunauerEmmett-Teller (B.E.T.) surface area determinations were made using nitrogen (adsorbate) in helium (carrier) gas mixtures.
Animals MATERIALS AND METHODS Materials For this study, we prepared five types of calcium phosphate glasses (CaO/P,O, = 1) in which 100 moles of Ca(PO,), was combined with x moles of SiOZ (x = 0, 5, lo,15 or 40 mol of silica). We tried to make glasses with x > 40 but clear glasses were not obtained owing to the higher melting temperatures with increasing SiO,. All those specimens were obtained from the melts with the Correspondence to Dr Mitsuo Nagase, Department of Orthopaedic Surgery, Gunma School of Medicine, 3-39-22 Showo, Maebishi, Gunma, 371 Japan. Biomaterials
1992, Vol. 13
No. 3
The Gbweek-old male C57BL/6 mice used in this study were acclimatized in our animal care facility for one week prior to use. They were housed ten per cage and maintained under controlled temperatu~, lighting and relative humidity. Animals were allowed free access to water and fed normal chow.
Animal experimental
protocol
The adult mice were exposed to single doses of silica in the range 0-40 mol in calcium phosphate glass. Ten animals of each group received each of these doses. Animals were given an intraperitoneal dose of glass powder (5000 mglkg body wt) dispersed in 1.0 ml of normal saline. All of the animals were studied simulD 1992 Bu~e~o~h-Hei~mann 0142-~12/92/030172-~
Ltd
glass: M. Nagase et al.
Toxicity of silica-containing
taneously and were dosed intraperitoneally same single glass stock suspensions.
Quantitative
determination
of dissolved
from
173 100 Co ( P03& to X SiO,
the
X
ions
IO3
The amount of dissolved Si4+, Ca2+ and P5+ from the glass specimen in water was quantitatively determined using an inductively coupled plasma (ICP) analyser (Shimadzu, ICPS-500). A plate-like specimen of a sample of glass (surface area = 2 cm’) was used. The dissolved ions were determined after immersing the specimen in 300 ml of distilled water at 37°C for 50 h.
Mortality Intraperitoneal administration of calcium phosphate glass containing various concentrations of silica resulted in an increase in the mortality with increasing amounts of silica in the glass. The mortality was O/10, 3/10, 9/10, lo/10 and lo/10 at x = 0, 5, 10, 15 and 40 mol of silica, respectively (Figure 1). The mean time of death was 68.0, 29.3, 19.2 and 6.0 h after treatment at x = 5, 10, 15 and 40 mol of silica, respectively (Figure 1). The mortality in mice given pure silica glass powder itself was O/10. In contrast to the mortality with silica glass itself or calcium phosphate glass without silica, the silica in calcium phosphate glass decreased the mean time of death and increased the number of deaths within 30 days.
Quantitative
determination
of dissolved
ions
The dissolved amount of Si4+, Ca2+ and P5+ in the water from the calcium phosphate glasses increased with the increasing amount of silica in the glass (Figure 2). No dissolved Si4’, Cazt or Pst was detected by ICP after 50 h of treatment of pure silica glass [SiO,: molar fraction = 1) in water.
I.-
T102 9 5 E Z P Z 51 .u, 10 n
RESULTS
-
I
I 1
0.1
Ca(P4)2 Figure2
’
1
I
0
0.3
0.2
aI.0 SiO,
Molar fraction
Amount of dissolved ions in water at 37°C for 50 h
versus. glass composition. The dissolution of ions into the water depended on the glass composition. (0) Ca2+;( A) Si4+.
Effect of particle
P5+; (0)
measurements
The differences in grain sizes or surface areas among samples did not affect the amounts of dissolved ions from the samples. The diameters of grains of materials used in this experiment were 3.9-8.3 pm (lable 1). The surface areas of the materials were 2.3-3.2 m’/g, except for the pure silica glass itself, which was 13.0 m2/g (7Me 1).
DISCUSSION Silicon is, next to oxygen, the most abundant element in the earth’s crust. Although only trace amounts appear in most animal tissues, silicon is an essential element in the chick” and the rat’, required for normal growth and development. Silicon may function as a biological crosslinking agent and contribute to the architecture and resilience of connective tissue’. The ingestion of a small amount of amorphous silica (silicon dioxide] powder appears to be completely harmless. However, when Table1
24
48
72
96
120
144
168
Time (h)
Figure1 The mortality in C57BL/6 mice given a single intraperitoneal dose of the glass powders. Administration of calcium phosphate glass containing increasing concentrations of silica increased the mortality depending on the amount of silica in the glass. (Cl) 0 mol; (4) 5 mol; (A) 10 mol; (0) 15 mol; (0) 40 mol; (W) SiO,.
Grain sizes and surface areas
Materials*
Grain sizes (pm)
Surface areas (m2/g)
0 mol 5 mol 10 mol 15 mol 40 mol SiO,
5.9 4.3 3.9 4.3 6.3 4.1
3.2 2.3 3.0 2.5 3.1 13.0
*O,5,lO. 15 and 40 mol = calcium phosphate glass (Ca0/P20, = 1) with the composition of 100 Ca(P03)2 to x 90, in moles in which x = 0, 5, 10, 15 or 40 mol of silica, and SiO, = pure silica glass itself.
Biomaterials
1992, Vol. 13 No. 3
174
silica enters the body in any other way than by ingestion, some type of toxicity almost invariably resultsg. We showed that a calcium phosphate glass-ceramic containing 34.2 wt% of silica was lethal in C57BLl6 and A/J mice4. The LD,, of the silica containing glass-ceramic after intraperitoneal administration in C57BL/6 mice was 750 mg/kg .(Ref. 4). The mice died from 2 to 6 days after administration of the glass-ceramic4. Significant footpad swelling occurred for a week after challenge in mice challenged with the glass-ceramic4. The lethality of the material administered weekly did not change during 5 weeks4. A special kind of inflammation, not an immunological one4, plays an important role in the lethal reaction. The present study was undertaken to examine the intraperitoneal toxicity of calcium phosphate glass containing various concentrations of silica in C57BLl6 mice. Calcium phosphate glasses containing large amounts of silica are extremely toxic. The Si4+, Ca2+ and P5+ in water dissolved from the calcium phosphate glass depends on the amount of silica in the glasses, but the amounts of Ca2+ and P5+ were much lower than the amounts normally existing in serum. The dissolved silica (Si4+) in this study may be monomeric or may not be highly polymerized lo . It is suggested that the concentration of monomeric or low molecular silicic anions determines the toxicity of the materials, but the total content of non-dissolved silica had no relation to its toxicity. In determining the biological consequences of dissolved or monomeric silica, what kind of reaction may play an important role in the lethal reaction will require further study. Under these circumstances it is interesting to speculate on the mechanism of the biological effects of dissolved or monomeric silica. There have been a great many studies to explore the pathogenesis of silicosis’1-‘6. But almost all silicosis researchers studied the biological reactions of various forms of solid silica, e.g. crystalline silica [quartz, tridymite, cristobalite, coesite and stishovite)16, silica powder without an organophilic hydrophobic monolayer of hydrocarbon groups15, silica gelI and the the finest amorphous silica *I . Nobody has investigated toxicity of monomeric or low molecular silicic anions dissolved from composition materials. We showed in this study that there was no toxicity in pure silica (SiO,) glass or in calcium phosphate (Ca[PO,),) glass without silica. Monomeric silicic anions dissolved from silica-containing glasses may induce the toxicity. The difference in toxicity among the various forms of solid silica may depend on the amounts of monomeric silica dissolved from the silica, The difference in the mean time of death (6.0-68.0 h after administration in this study, 75.9 h after administration in our previous study41 may depend on the different rates at which monomeric silica is dissolved from composition materials. Silica is also released from ‘bioglass’, and the dissolved amount depends on the glass composition’7. It is hoped that future researchers and engineers will be mindful of this toxicity due to silica dissolved in calcium phosphate glass. Note also that we have not addressed any variation in sensitivity to silica with respect to age, sex, diet, route of exposure, or treatment regimen. These factors may also affect the 36day mortality in mice. The marked physiological response at the high doses Biomaterials 1992. Vol. 13 No. 3
Toxicity
of silica-containing
glass: M. Nagase et al.
of silica may well be symptomatic of the detrimental biochemical changes that follow silica exposure, even long after implantation, since glasses and ceramics are susceptible to both general corrosion and pitting attack in aqueous solutions, and silica is 20 times more soluble in organisms than in saline solution”. Several types of glass-ceramic in systems with calcium phosphate without silica have been developed’” lg. That kind of glassceramic would be expected to be much better for implant systems than silica-containing glass-ceramics although the former have some disadvantages such as the weak mechanical strength in water and low chemical stability.
The authors are grateful to Dr Hiroyasu Takeuchi, Research and Development Center, Mitsubishi Material Co., Ltd for his assistance in measurements of grain size and surface area; and Mr Kiichi Oosawa, Department of Orthopaedic Surgery, Gunma University School of Medicine for his assistance in photography. This work was supported in part by Grant of Japan Sports Medicine Foundation Inc. No. 1.
REFERENCES 1
Hench, L.L., Splinter, R. J., Allen, W.C. and Greenlee, T.K. Bonding mechanism at the interface of ceramic prosthetic materials, 1. Biomed. Mater. Res. Symp.
2
Blenke, B.A., Pfeil, E., Bromer, H. and Kas, H.H., Implantate aus Glas-Keramik in der Knochenchirurgie (Tierexperimentelle Untersuchungen), Langenbechs Arch. Chir. Suppl. Forum 1973, 107-110 Kitsugi, T., Yamamuro, T., Nakamura, T., Higashi, S., Kakutani, Y., Hyakuna, K., Ito, S., Kokubo, T., Takagi, M. and Shibuya, T., Bone bonding behavior of three kinds of apatite containing glass ceramics, I. Biomed.
1971,2,117-141
3
Mater.
4
5
6 7 a
Res.
1986, 28, 1295-1307
Nagase, M., Shimizu, T., Chigira, M. and Udagawa, E., [An analytical study of the inflammation induced by a calcium phosphate ceramic,] Kitakanto Zgaku 1967, 37, 101-106 Bradford, M.M., A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding, Anal. Biocbem. 1976, 72, 246-254 Carlisle, E.M., Silicon: an essential element for the chick, Science 1972, 178, 619-621 Schwartz, K. and Milne, D.B., Growth-promoting effects of silicon in rats, Nature 1972, 239, 333-334 Schwartz, K., A bound form of silicon in glycosaminoglycans and polyuronides, Proc. Natl Acad. Sci. USA 1973, 70, 1608-1612
9
10
Michon, R., Sue, P. and Merinis, J., Metabolisme de la Silice et des Silicates Inhales par l’Anima1, Suiva a 1’Aide de %i. C.R. Acad. Sci. 1956, 243, 2194-2195 Abe, Y., Yoshikawa, Y., Naruse, A. and Saito, H. [Effect of SiO, additive on the rates of crystallization of Mg(PO,), and Ca(PO,), glasses,] I. Ceram. Sot. Jpn 1977, 85.151-153
11
Lungren, K.D., Experimental investigations on the significance of the size of particles in the reaction of the
Toxicity of silica-containing
12
13
14
15
glass: M. Nagase et al.
peritoneum to amorphous silica, Acta Med. Stand. 1953, 145, 84-90 Minova, K. F., Respredelennie Kremniia v Organizme Belykh Krys Pri Eksperimental’ nom Silikoze, Gig. fl. Prof. ZaboZ. 1989, 13,50-51 Oghiso, Y. and Kubota, Y., Interleukin 1 production and accessory cell function of rat alveolar macrophages exposed to mineral dust particles, Microbial. Zmmunol. 1987,31,275-287 Policard, A., Collet, A., Daniel-Moussard, H. and Pregermain, S., Sur les premiers stades des alterations experimentales du rein par l’acide silicique._Etude au microscope electronique, J. Ural. Med. Chir. 1980, 85, 585-800 Schepers, G.W.H., Hypertension due to inhaled sub-
175
18
17
18
19
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1992,Vol. 13 No. 3
