Effectiveness of Dicalcium Phosphate Dihydrate as Biocompatible Coatings on 316L and 316LN Stainless Steel

During 1980s, biomedical implants were a feasible choice for patients undergoing joint replacement, but the orthopaedic market place failed to create cement less implant. As a consequence bioactive coatings were proposed as orthopedic implants. The biomedical industry used ceramic coating on stainless steel as they are wearresistant and also aid in Osseo-integration between bone and implant [1]. The physical and mechanical properties analogous to device design and functionality are the added advantages of these Coatings. Electrophoretic deposition process is unique as the physical and mechanical properties of these coatings are the point of focus in medical implants [2]. Dip coating technology facilitates in designing an implant with beneficial mechanical properties and also provides superb synthetic bone properties [3].


Introduction
During 1980s, biomedical implants were a feasible choice for patients undergoing joint replacement, but the orthopaedic market place failed to create cement less implant. As a consequence bioactive coatings were proposed as orthopedic implants. The biomedical industry used ceramic coating on stainless steel as they are wearresistant and also aid in Osseo-integration between bone and implant [1]. The physical and mechanical properties analogous to device design and functionality are the added advantages of these Coatings. Electrophoretic deposition process is unique as the physical and mechanical properties of these coatings are the point of focus in medical implants [2]. Dip coating technology facilitates in designing an implant with beneficial mechanical properties and also provides superb synthetic bone properties [3].
In view of their processability, weldability, satisfactory mechanical properties, Metallic biomaterials are found to be beneficial in the field of biomedical materials. They degrade when in synergy with body fluids, which is the main drawback of these metallic biomaterials. Hence, care must be taken while using metallic biomaterials for conventional metallic implants. Corrosion resistance; the capacity to generate a protective passive film is checked before selecting the materials for implants. Commercially, 316L and 316LN stainless steels are widely used in biomedical applications. An external oxide layer protects the materials, assuring a satisfactory corrosion resistance. The trouble associated with corrosion is, release of ions from metallic species, that are harmful to the organism [4]. This protective passive layer decreases the corrosion rate and also ceases the ion release. Corrosion behavior of 316L and 316LN is the determining factor for their success as biomaterials [5]. The primary step for developing new biomaterials is, evaluation of their corrosion parameters in vitro. Among the mechanical properties, UTS (Ultimate Tensile Strength) is mainly concerned. UTS of 316L is 558Mpa and 316LN is 685Mpa [6].
In this study various experiments were done to test whether 316L and 316LN steel can be used as orthopedic implant. The 316L and 316LN were coated with Dicalcium phosphate di-hydrate by Electrophoretic deposition method and with polyvinyl alcohol by Dip coating method. The corrosion behavior of these two coated 316L and 316LN stainless steel were evaluated by electrochemical techniques. OCP (Octacalcium phosphate) and DCPD (Dicalcium phosphate dihydrate) were deposited on Ti by ECR, whereas HAp was deposited on 316L alloy by EPD method [7]. So in this study DCPD has been deposited by electrophoresis. Samples were electrophoretically coated with Di-calcium phosphate di-hydrate, and dip coated with polyvinyl alcohol. Comparison of corrosion resistance among the coated samples revealed interesting characteristics. Coated 316LN showed better corrosion resistance than 316L. Dip coated 316LN shows better corrosion resistance than 316L. Coated samples were further studied by The Scanning Electron Microscope and Energy Dispersive X-Ray Spectroscopy. Though electrophoretic deposition gave much better coating and uniform variation of calcium compared to dip coating, E CORR , I CORR values of dip coated samples in Ringers's solution were better presumably because of formation of passive layer during dip coating. However stability of dip coated surface was poor.

Abstract
The corrosion behaviors of two materials 316L Stainless Steel and 316LN Stainless Steel have been investigated for use as biomaterials. These samples were electrophoretically coated with Dicalcium phosphate dihydrate, and dip coated with polyvinyl alcohol. Time, current, concentration and voltage were the variables during electrophoresis. Dip coating was done for the same periods of time as was done during electrophoresis. Corrosion resistance properties were measured in Ringer's solution by Gamry Potentiostat. The I CORR and E CORR values were estimated using Gamry Echem Software and Tafel's extrapolation method. Coated samples were immersed in SBF solution for different periods of time, viz., 1 second, 24hours, 72hours and 1week and then further I CORR and E CORR values were estimated in Ringer's solution. For coated samples Electrochemical Impedance Spectroscopy were also done. Different parameters like Rp, alpha, Wd of EIS were used to evaluate the effectiveness of the coatings. Comparison of corrosion resistance among the coated samples revealed a few interesting characteristics. While DCPD coated Stainless Steel showed considerable improvement in corrosion resistance compared to as received sample, dip coated samples did not show appreciable improvement. Coated 316L shows better corrosion resistance than 316LN. Dip coated 316LN shows better corrosion resistance than 316L. So Electrophoretic Deposition gave much better coating in comparison to Dip coating. Coated samples were further studied by The Scanning Electron Microscope and Energy Dispersive X-Ray Spectroscopy. While SEM was done to ascertain uniformity of coating, EDAX was done to see the variation of calcium deposition as a function of different deposition parameters. Electrophoretic deposition gave much better coating and uniform variation of calcium compared to dip coating.

Experimental Procedure
Compositions of the samples are given in table 1.

Coating procedure
Two types of coatings were done: Calcification with electrophoretic deposition: For EPD Dicalcium phosphate dihydrate (CaHPO 4 ,2H 2 O) was used. It is practically insoluble in water, with a solubility of 0.02 g per 100 ml at 25°C. The sample to be coated was properly polished. Then the sample was used as anode and graphite plate was used as cathode. EPD experiments were done by varying the current, time, concentration of DCPD and voltage. The parameters are given in table 2. Calcification experiment was conducted by immersing the 316L and 316LN substrates in a phosphate-buffered solution, prepared by 8 gm Disodium hydrogen phosphate (Na 2 HPO 4 ) and 0.1(M) HCl at around neutral pH at room temperature. The solution was then adjusted to slight super saturation with respect to Dicalcium Phosphate Di-Hydrate (DCPD), which potentially promotes the nucleation and calcification of the calcium phosphate crystals. After the calcium phosphate was deposited on the substrates, the substrates were washed with double distilled water and dried.
Dip coating procedure: For dip coating polyvinyl alcohol was used. After proper polishing, samples were dipped in a phosphate-buffer solution added with polyvinyl alcohol for three different periods of time, viz., 30 minutes, 45 minutes and 60 minutes. After the deposition, substrates were dried.

Stability of DCPD deposit in SBF solution: 316L and 316LN
substrates after calcification were immersed in SBF at 37°C for 1second, 24hours, 72hours and 1week. After the immersion, the comparison of E CORR , I CORR values in SBF solution of the two different samples immersed for different periods of time were estimated.

Corrosion testing
Standard Electrochemical Corrosion Cell was used to perform the electrochemical potentiostatic polarization tests on standard flat metal specimens. Polarization experiments were carried out as per ASTM ST72 using Gamry Potentiostat. The software used was Gamry Echem Analyst. Potentiodynamic experiment in Ringer's solution with a scan rate of 1mV/sec was done with the as received, DCPD coated, Dip coated, and SBF solution immersed samples. I CORR -E CORR values were estimated from the polarization curves by Tafel's extrapolation method and are given in table 3 and a few typical polarization diagrams in Ringer's solution are given in figures 1-7. The composition of 250ml Ringer's solution is: NaCl -2.15gm/l, CaCl 2-0.0825gm/l and KCl -0.0759gm/l with pH 7.4 maintained throughout the experiment The Electrochemical impedance spectroscopy was done with the coated samples. Then the Bode plot, Nyquist plot and Rp and Ru values were obtained and the generated data from these curves are presented in table 3. Stability test at SBF solution curves (Figures 8,9).

Polarization behavior of DCPD coated 316L and 316LN samples
There were four variable in the electrophoretic depositions, viz., time, current, voltage, and concentration. Polarization study revealed a definite pattern of these variables on the I CORR and E CORR values. The pattern of variation was different for 316L and 316LN.
Effect on 316L: With increasing time of deposition E CORR becomes nobler but I CORR remains almost same. At 60mins E CORR is of -420mV vs. SCE and I CORR is of .1μA/cm 2 . It gives good passivity. With increasing current E CORR becomes nobler and I CORR keeps on decreasing. At 60mA current, E CORR is of -462mV vs SCE and ICORR are of 1 μA/cm 2 . This combination gives good passivity. In case of voltage variation, Figure  1 shows 5volt curve gives the noblest E CORR i.e. -242mV vs. SCE and minimum I CORR i.e. 9μA/cm 2 . It shows the best passivity. At 10volt E CORR and I CORR are not up to the mark. At 2 volts E CORR is almost same as  5volt. Concentration variation shows at 0.02g concentration E CORR is of -900mV vs. SCE and I CORR is of 90 μA/cm 2 . It also shows good passivity. At 0.06g concentration E CORR and I CORR are not up to the mark. From the discussion of the effect of different parameters of Electrophoretic deposition on 316L it appeared that carrying out the deposition at 5 Volts with maximum current with 0.06 g concentration for 45 minutes would give the best coating. So the deposition was carried out with these parameters and polarization tests were done in Ringers solution. Figure 6 shows that E CORR is of -242mV vs SCE and I CORR is of 0.9 μA/ cm 2 . Interestingly this is the best corrosion resistance as was thought.      Similarly for 316LN it appeared that carrying out the deposition at 60 mA and .06g concentration with maximum current for 45 minutes would give the best coating. So the deposition was carried out with these parameters and polarization tests were done in Ringers solution. Figure 6 shows E CORR (-171.4 mV vs. SCE) and I CORR (0.45 μA/cm 2 ). Interestingly this too reflects the best corrosion resistance. Moreover temperature also plays a very important role (Tables 3-5). The deposition experiments were carried out over the period of September to May at room temperature, which varied substantially.

Stability of the DCPD coated 316L and 316LN samples in SBF solution
SBF (pH 7.25) [8] is a metastable solution containing calcium and phosphate ions already supersaturated with respect to the apatite.
In case of 316L from the figure 8 at different time periods (1second, 24hours, 72hours, 1week) it can be seen that for all immersion times, i.e., for 1 second, 24 hours and 72 hours of immersion give almost same E CORR and I CORR . But 1week curve gives much active E CORR of -631 mV vs SCE and much higher I CORR of 200 μA/cm 2 . Beyond particular time period stability decreases. Figure 9 shows the corresponding behavior for 316LN at 1second, 24 hours, 72 hours and 1 week time periods. E CORR and I CORR vary sinusoidally. The stability of 316LN is found to be better than that of 316L.

Polarization behavior of Dip coated 316L and 316LN samples
In case of 316L, figure 7 shows, dip coated sample at 45 mins gives noblest E CORR of -328.1 mV vs. SCE and minimum I CORR of .3 μA/cm 2 in comparison with DCPD coating. In case of 316LN, figure 7 shows dip coated sample with active E CORR of -351 mV vs. SCE and minimum I CORR of .07 μA/cm 2 which is also better with respect to DCPD coated sample. Though dip coated samples exhibit much superior corrosion resistance I CORR in the nano range but on immersing the dipcoated sample in SBF coats come off ( Table 6).

Analysis of EIS data
EIS study of a few selected coated samples was done. The study revealed CPE with Diffusion. Barring a few in most of the cases fits were good. This suggests very effective coating or formation of continuous layer over the surface during coating. Even the dip coated samples which showed good corrosion resistance shows good fit suggesting that corrosion resistance obtained maybe due to the inherent nature of the alloy. In case of 316L DCPD coated at 5volt gives best result interms of higher Ru (uncompensated resistance) i.e. 34.67 ohms and Rp value is     18.04 ohms. In case of 316LN DCPD coated sample 60mA gives best result interms of higher Ru i.e. 73.75 ohms. As we all know higher the value of Ru and Rp higher the stability of coating (Table 7).

SEM of 316L SS DCPD coated samples
Scanning Electron Microscopic and EDX study of DCPD and Dip coated 316L and 316LN samples: Morphology of the coatings was investigated by scanning electron microscopy with associated energy dispersive spectroscopy analysis (SEM-EDS). Figure 10 shows coating morphology of 316L DCPD coated samples at x10,000 magnification. The coating is uniform and there are no cracks. The satisfactory adhesion between the coating and substrate suggests its suitability for load-bearing capability. Figure 11 shows coating surface of 316LN sample, which shows better uniformity in coating than 316L at the same magnification. Figure 12 shows dip coated surface of 316L at x200 and x500 magnification. It shows porosity in coated surface. Figure 13 shows layered structure of 316LN dip coated surfaces at x1,000 and x2,000 magnifications. Since these specimens exhibited very good corrosion resistance and passivity, these layers are presumably oxide (passive) layers. The comparative EDX data (atomic%) of different coated samples is given in Here from the equation it can be seen that Di-calcium phosphate dihydrate reacts with Di-sodium hydrogen phosphate and Hydrochloric acid and produces Mono-calcium hydrogen phosphate (anhydrous MCPA) [2,6], and Sodium hypo-chloride. From the products of the     above equation it can be seen stoichiometrically Calcium ratio is half of the phosphorous. In case of DCPD coated (at 60mA current) 316L sample, position 1 shows PK α is 1.55 atomic% and CaK α is .67 atomic%, which satisfies the coating equation stoichiometricaly. DCPD coated (at 60mA current) 316L sample, position 2 shows PK α is .53 atomic% and CaK α is .79 atomic%, which somehow differs the equation. From the above discussion it is apparent that by Electrophoretic route it is mono calcium phosphate (anhydrous) that gets deposited rather than Dicalcium phosphate dihydrate.

Dip coating equation:
The possible reaction that takes place during dip coating is given below (C 2 H 4 O) n + Na 2 HPO 4 + 2HCl  coated material + H 2 O There is OHgroup associated with PVA .So there is some unsaturated charge with this oxygen of this OHgroup which can form bond either with metal oxide or metal. For elements with unsaturated 3-D orbits the bond is quite stronger. So it is expected it will form either metal hydroxide or Me-O hydroxide with 316L and 316LN SS. This acts as passive layer and may improves the corrosion resistance of the dip coated SS. In case 316L, dip coating at 45 mins (position2) CrK α is 14.53 atomic% and O is 17.98 atomic% .This indicates formation of Cr 2 O 3 . Presence of chlorine can also be seen here. However, average oxygen is lower. In case of 316LN dip-coating at 45mins (position1) shows 30.35 atomic% of O. But in case of 316LN at position 2 there is a mere presence of chlorine.
The EDX data corroborates the SEM finding that oxygen content in the dip coated specimens are higher. So the inference drawn earlier to explain better polarization behavior of dip coated samples the passivity is the cause for better corrosion resistance is substantiated by the SEM-EDX study.

316LN showed greater improvement in corrosion resistance
property than 316L and also 316LN showed better passivity after coating.
2. Electrophoretic deposition of DCPD could be done though improvement in corrosion resistance was not up to a high level.
3. Dip coated samples showed remarkable improvement corrosion resistance property and passivity.