Role of Sintering Temperature Dependent Crystallization Of
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Processing and Application of Ceramics 13 [1] (2019) 12–23 https://doi.org/10.2298/PAC1901012C Role of sintering temperature dependent crystallization of bioactive glasses on erythrocyte and cytocompatibility Shivalingam Chitra1, Purushothaman Bargavi1, Dhinasekaran Durgalakshmi2, Padmanaban Rajashree3, Subramanian Balakumar1,∗ 1National Centre for Nanoscience and Nanotechnology University of Madras, Guindy Campus, Chennai-600025, India 2Department of Medical Physics, Anna University, Chennai-600025, India 3Centre for Advanced Study in Crystallography and Biophysics University of Madras, Guindy Campus, Chennai-600025, India Received 10 August 2018; Received in revised form 14 November 2018; Accepted 11 December 2018 Abstract Bioglass (BG) was preparedby sol-gel method and the role of sintering temperatures (600, 700 and 800°C) on crystalline phase changes, bioactivity, erythrocyte and MG-63 cell line compatibility was investigated. Increase in sintering temperature from 600 to 800°C led to the secondary phase formation that was confirmed through structural analysis. Micrographics revealed the formation of nanorods (700°C) and nanoflake like (800°C) morphologies. Biocompatibility assay showed that, BG sintered at 600°C had optimal biocompatibility while better mechanical property was noted at 700°C. Altogether, the study demonstrated that increasing the sinter- ing temperature will result in increased crystallinity which in turn resulted in the optimal biomineralization but decreased the biocompatibility. Hence, we demonstrated the importance of temperature during the processing of BG for various applications, as it affects many properties including bioactivity and compatibility. Keywords: bioactive glass, sol-gel, sintering, bioactivity, biocompatibility I. Introduction to gain better bioactive as well as biocompatible mate- rials. Even though several varieties of commercial ma- The Bioglass 45S5 particulates have been used in the ® terials have evolved; still research is needed to improve name of NovaBone (NovaBone products LLC, Jack- the aspect of dissolution/degradation resulting from the sonville, FL) [1] to cure maxillofacial and orthopaedic bioactivity/biocompatibility. defects. The Endosseous Ridge Maintenance Implant The advantage of bioglass is its easy bonding with (ERMI) introduced a bioglass (BG) into the tooth ex- both bone and tissue [6,7] and faster response and in- tracted sites in the form of cone and putty as im- teraction between implant material and host tissue than plants [2]. PerioGlas (US Biomaterials at present sold other biomaterials such as hydroxyapatite and trical- to NovaBone) was used to treat periodontal diseases cium phosphate [8]. However, several processing routes, [3]. Biogran is one of other forms of the Bioglass 45S5 such as the production of scaffolds or the deposition of (Biomet 3i, palm Beach Gardens, FL) which is used coatings, include a thermal treatment to apply or sinter for jaw bone defects [4]. The current issue is that in the glass. The exposure to high temperature may induce all the commercial materials particle sizes are greater a devitrification phenomenon, altering the properties than 50 µm, which may cause the toxicity at the ge- and, in particular, the bioactivity of the glass. During nomic level [5]. Hence with the advent of nanotech- sintering, the material becomes more compact, whereas nology, the research has moved towards optimisation of the degree of porosity decreases and interconnecting crystallinity and morphological characteristic features, porosity may be lost. Thus, osteoconductivity will be re- ∗ ffi Corresponding author: tel: +44 22202749, duced and biodegradability becomes more di cult [9]. e-mail: [email protected] Often, ceramic particles are still visible, even after sev- 12 S. Chitra et al. / Processing and Application of Ceramics 13 [1] (2019) 12–23 eral years. They have a negative influence on the me- II. Experimental procedure chanical properties of the bone and favour chronic in- flammation [10]. 2.1. Materials and methods The bioglass 45S5 system contains 45wt.% of SiO2, All the chemicals and reagents used in the present 24.5wt.% Na2O, 24.5wt.% CaO and 6wt.% of P2O5 work are analytical grade without additional purifica- and mostly two methods are used for the synthesis: tion. TEOS (tetraethyl orthosilicate (98% pure) was pur- melt quenching and sol-gel method. We used the sol- chased from Alfa Aesar, orthophosphoric acid (88% gel method to prepare bioglass, because this tech- pure), calcium nitrate (99% pure) and nitric acid (70% nique takes lower preparation temperature, better con- pure) were purchased from Spectrum Reagents and trol, good quality, minimal cost and produces BG with Chemicals Pvt. Ltd., sodium hydroxide (98% pure) was high surface area accelerating the rate of hydroxyap- purchased from Sisco research laboratory. These chem- atite formation [11,12]. It is evident from the literature icals were utilized to prepare bioglass, through sol-gel that mechanical stability increases with increase in crys- method. Firstly, 45% of TEOS (tetraethyl orthosilicates) tallinity [13]. Massera et al. [14] studied BG crystal- was added with double distilled water, nitric acid and lization mechanism and its structural properties and re- ethanol for silica hydrolysis. It was stirred for 1 h to ported that better bioactivity results as a consequence of form a complete gel. Then reagents were dissolved in- partial crystallisation. Furthermore, Thomas et al. [15] dividually for 45 min and added to silica matrix in the expressed that crystallization mechanism depends on following order and amount: 6% orthophosphoric acid, sintering kinetics which contributes to the understand- 24.5% calcium nitrate and finally sodium hydroxide ing of the structural behaviour and phase transition in 24.5%. The resultant product was a white sol that was BG. Thus, as it is shown in Table 1 a few reports are kept for ageing under stirring for 12h to form a highly available on the structure stabilisation of BG at various ordered gel matrix and then it was dried in a hot air oven temperatures, however the relation between biocompat- for 24h at 120°C. Finally, in order to stabilize the phase ibility and crystallization temperature are sparsely re- formation, heat treatment was adopted at different tem- ported. Hence, we intend are intended to bring out the peratures such as 600, 700 and 800°C for 3h in a box significance of various sintering temperatures on the furnace under ambient atmosphere. Thereafter, they will erythrocyte compatibility, ALP activity, and cytocom- be designated as BG600, BG700 and BG800, respec- patibility in addition to bio mineralization of BG. tively. Table 1. Literature survey of bioglass crystallinity with respect to biocompatibility Crystalline phase and sintering Biocompatibility/ Method of synthesis Ref. temperature Bioactivity Bioglass® powderbymelt Na CaSi O – 800 °C Assessed biodegradation in the 2 2 6 [16] quenchingmethod Na2Ca4(PO4)2SiO4 – 800to 950°C termsof preferentialdissolution Bioglass® structural transformationsreportedby Na2CaSi2O6 –600to750°C – [17] melt quenching method Bioglass® (45S5) powder from Na Ca Si O –1050°Cfor140min – [18] NovaMin USA 2 2 3 9 Bioglass prepared by sol-gel Wollastonite, apatite and pseudo Elevated in vitro apatite formation [19] process wollastonitephases–1000°C at1000°C 45S5 bioglass purchased by U.S.BiomaterialsCorp.andball Na Ca Si O – 1000°C Wollastonite phase was formed [20] 2 2 3 9 after mineralization milled for 4h Na2CaSi2O6, Ca5(PO4)2SiO4, CaSiO3 In vitro dissolution kinetics was Bioglass synthesised by sol-gel –700to1000°C foundinthepresenceof [21] method Wollastonite phase – 1100 °C crystalline phase Bioglass® powderusingspark Amorphous–550°C BG(600°C)showedimproved [22] plasma sintering Na2CaSi2O6 – 600 °C bioactivity Melt derived 45S5 powder Investigated cell proliferation and sinteredbysparkplasma Na2CaSi3O8 –500to600°C alkalinephosphataseactivityby [23] sintering (SPS) MG-63 and L929 cell line 13 S. Chitra et al. / Processing and Application of Ceramics 13 [1] (2019) 12–23 2.2. Material characterization used FESEM to examine the morphological changes The prepared bioglass was characterized to study due to gradual dissolution and the formation of bone like its thermal properties using thermogravimetric analysis hydroxyl carbonate apatite layer on the material surface. ff and di erential thermal analysis (SII Nanotechnology- 2.4. Haemocompatibility studies TG/DTA-6300, Japan). Structural and phase conforma- tions were examined using X-ray diffraction (XRD, Haemocompatibility is one of the essential methods PANalytical Instruments, The Netherlands) using Cu- to evaluate the compatibility of any biomaterial using Kα1 radiation (λ = 0.154nm) with the scanning rate erythrocytes. Initially, blood was collected from volun- of 2θ = 3°/min. The morphological changes of BG teer donor retained by using EDTA to inhibit the co- before and after immersion were examined using field agulation and centrifuged for 10min at 4°C, and again emission scanning electron microscope (FESEM, HI- washed 2 to 3 times using phosphate buffer saline (PBS) TACHI SU-6600, Japan) with the electron beam en- to separate plasma. The obtained blood sample (50 µl) ergy of 15keV. Further on, to confirm the molecular and PBS (950 µl) were mixed with bioglass samples in vibrations, we utilized Raman and Fourier transform Eppendorf tubes. All bioglass samples were analysed infrared spectroscopy (Nanophoton Raman 11i, Japan in triplicates (n = 3) with