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Corrosion of experimental magnesium alloys in blood and PBS: A gravimetric and microscopic evaluation
Materials Science and Engineering B 176 (2011) 1797–1801
Contents lists available at ScienceDirect
Materials Science and Engineering B journal homepage: www.elsevier.com/locate/mseb
Short communication
Corrosion of experimental magnesium alloys in blood and PBS: A gravimetric and microscopic evaluation Ch. Schille a,∗ , M. Braun b , H.P. Wendel b , L. Scheideler a , N. Hort c , H.-P. Reichel d , E. Schweizer a , J. Geis-Gerstorfer a a
University Hospital Tuebingen, Center for Dentistry, Oral Medicine and Maxillofacial Surgery, Section Medical Materials & Technology, Osianderstr. 2-8, D-72076 Tübingen, Germany University Hospital Tuebingen, Div. Congenital & Paediatric Cardiac Surgery, University Children’s Hospital, Tuebingen, Germany, Calwerstr. 7/1, D-72076 Tübingen, Germany c GKSS Research Centre, Institute of Materials Research, Max-Planck-Str. 1, D-21502 Geesthacht, Germany d Weissensee Company, Bürgermeister-Ebert-Str. 30-32, D-36124 Eichenzell, Germany b
a r t i c l e
i n f o
Article history: Received 3 November 2010 Received in revised form 11 March 2011 Accepted 10 April 2011 Available online 28 April 2011 Keywords: Gravimetry Magnesium alloys Static immersion test Chandler-loop
a b s t r a c t Corrosion tests for medical materials are often performed in simulated body fluids (SBF). When SBF are used for corrosion measurement, the open question is, how well they match the conditions in the human body. The aim of the study was to compare the corrosion behaviour of different experimental magnesium alloys in human whole blood and PBSminus (phosphate buffered saline w/o Ca and Mg) as a simulated body fluid by gravimetric weight measurements and microscopic evaluation. Eight different experimental magnesium alloys, containing neither Mn nor other additives, were manufactured. With these alloys, a static immersion test in PBSminus and a dynamic test using the Chandler-loop model with human whole blood over 6 h were performed. During the static immersion test, the samples were weighed every hour. During the dynamic test, the specimens were weighed before and after the 6 h incubation period in the Chandler-loop. From both tests, the total mass change was calculated for each alloy and the values were compared. Additionally, microscopic pictures from the samples were taken at the end of the test period. All alloys showed different corrosion behaviour in both tests, especially the alloys with high aluminium content, MgAl9 and MgAl9Zn1. Generally, alloys in PBS showed a weight gain due to generation of a microscopically visible corrosion layer, while in the blood test system a more or less distinct weight loss was observed. When alloys are ranked according to corrosion susceptibility, the results differ also between the test systems. The MgAl9 alloy, showing the most pronounced corrosion in PBS, was one of the least corroding alloys under simulated in vivo conditions in blood. Thus, the ranking concerning clinical suitability of the magnesium alloys tested in this study is different, depending on the used electrolyte and the kind of method. For a possible clinical use, the alloy MgAl9Zn1 might be preferable for further investigations. © 2011 Elsevier B.V. All rights reserved.
1. Introduction One problem of magnesium alloys for biomedical applications is their fast degradation. To test the degradation in vitro, different kinds of corrosion measurements can be applied, such as static immersion or electrochemical tests. Most in vitro corrosion studies of magnesium alloys were performed in different simulated body fluids (SBF) like phosphate buffered saline (PBS), Hanks or Ringer solution, artificial plasma, etc. Different pH values and composition of the simulated body fluids influence the corrosion behaviour, as well as kind of fabrication, composition, grain size, finishing, heat treatment, coating, etc. of magnesium alloys. The
∗ Corresponding author. Tel.: +49 7071 2983493/2986199; fax: +49 7071 295775. E-mail address: [email protected] (Ch. Schille). 0921-5107/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.mseb.2011.04.007
advantage of using simulated body fluids for in vitro studies is that they can be easily handled and provide fast first information [3,4]. The open question is whether these results correspond to results obtained under in vivo conditions [2], since the compositions of SBFs are rather simple and do not resemble the much more complex composition of body liquids like blood or interstitial fluid. For biomaterials with an intended application in the blood stream, the dynamic flow is an additional issue of the surrounding corrosion medium, so that static immersion tests are of limited value. There are only few publications about corrosion measurements under dynamic conditions. However, these measurements were also performed with simulated body fluids [3,4]. We therefore looked for a model to simulate clinical conditions, using a real body fluid instead of SBF under dynamic testing conditions and set out to compare this system with static immersion test conditions using SBF.
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Ch. Schille et al. / Materials Science and Engineering B 176 (2011) 1797–1801
Table 1 Ion concentration of PBSminus compared to human whole blood [8]. Ion
Blood
PBSminus
+
142.0 5.0 1.5 2.5 103.0 27.0 1.0 0.5
146.0 4.1 – – 140.6 – 9.5 –
Na K+ Mg2+ Ca2+ Cl− (HCO3 )− (HPO4 )2− SO4 2− Concentration in mmol/l.
The Chandler-loop is a measurement system for testing the hemocompatibility of stents under dynamic flow conditions and in human whole blood [6,7]. The aim of this in vitro study was to compare the corrosion results of eight different experimental magnesium alloys after 6 h immersion in an artificial, simulated body fluid (PBSminus , phosphate buffered saline without Mg+ and Ca+ ), with the results from corrosion measurements in human blood. For comparison, the ion concentrations of blood and PBSminus are presented in Table 1 [8]. 2. Materials and methods The composition of the experimental magnesium alloys, which were used in this study can be seen in Table 2. The alloys resemble well-known technical alloys like e.g. AZ 91, but were manufactured without additives like e.g. Mn, they contain only the main alloying elements Mg, Al, Zn, Nd and Y [5]. From each alloy, round sample discs with a diameter of 10 mm and 1 mm height were prepared, polished with SiC 1200 in ethanol on both sides and ultrasonically cleaned in ethanol for 3 min.
were made with a rotation speed of 10 rpm and at a temperature of 37 ◦ C using a thermostat-controlled water bath (Q102, Haake, Berlin). After 6 h the tests were terminated and the samples were cleaned in different solutions (0,9% saline, 1% SDS solution) and stored in ethanol. Before and after the chandler-loop tests the weight of the two samples of each (dried) assembly was measured. 2.2. Gravimetric corrosion tests in PBS For the immersion test in PBSminus (DPBS w/o Ca, Mg; GIBCO (Invitrogen # 14190-169) six specimens of each alloy were investigated. Before onset of the immersion test, the weight of each sample was determined. Each sample was placed into a 15 ml PP tube, filled with 5 ml PBSminus and placed into a heating cabinet at 37 ◦ C. Change of weight (referring to original weight) was measured every hour after drying the samples for 20 min by exposure to air. Subsequently, samples were placed into new PP tubes with fresh electrolyte. 2.3. Microscopic evaluation and statistical calculation After the Chandler-loop and immersion tests, images were taken with a stereo microscope (Wild Heerbrugg) at magnifications of 10, 20 and 32. The gravimetric results of the Chandler-loop test were calculated per unit of area. Differences between means were pairwise tested for statistical significance using Student’s t-test (p < 0.05) for independent samples (Table 3). 3. Results and discussion
2.1. Gravimetric corrosion tests in blood
3.1. Gravimetric results
For the tests with the modified Chandler-loop model 24 specimens were used from each alloy. Samples were clamped pairwise in circular loops of PVC tubes (l = 50 cm), which were closed with a silicon tube. Two parallel sets of silicone loops were used in every experiment (4 samples per alloy). The experiments were repeated six times with blood from three different donors. Each tube was filled with 20 ml fresh human whole blood. The measurements
3.1.1. Immersion test over 6 h in PBSminus Fig. 1 shows the time-dependent gravimetric results over 6 h static immersion. For all magnesium alloys tested, it was found that in PBSminus a weight increase occurred under static immersion conditions. The increase is caused by the formation of a thick corrosion product layer [1], which can be seen in the microscopic pictures (Fig. 3). Fig. 1 also shows that after 4 h a kind of
Table 2 Chemical composition of the magnesium alloys tested. All values are given in wt.% and have been measured using a Spektrolab M (Spektro. Kleve. Germany). Al MgZn1 MgAl3 MgAl9 MgNd2 MgY4 MgAl3Zn1 MgAl9Zn1 MgY4Nd2
Zn
Y
Nd
0.966 3.07 8.43 1.99 4.95 3.01 9.03
1.01 1.16
0.00503 1.96
4.67
Fe
Cu
Ni
Si
Mg
0.00275 0.00311 0.00152 0.00410 0.00576 0.00387 0.00503 0.00480
0.00373 0.00202 0.00295 0.00384 0.00419 0.00249 0.00363 0.00413
0.00206 0.00096 0.00063 0.00193 <0.00020 0.00106 0.00081 0.00112
0.0316 0.0201 0.0154 0.0151 0.0339 0.0332 0.0229 0.0321
Bal. Bal. Bal. Bal. Bal. Bal. Bal. Bal.
Table 3 Statistical results from paired Student’s t-test (p < 0.05) of the gravimetric weight loss data after 6 h Chandler-loop in human whole blood for each magnesium alloy tested. MgZn1 MgZn1 MgAl3 MgAl9 MgNd2 MgY4 MgAl3Zn1 MgAl9Zn1 MgY4Nd2
Mg3Al3
MgAl9
MgNd2
Mg4Y4
Mg3Al3Zn1
MgAl9Zn1
MgY4Nd2
–
+ +
+ + +
+ – + –
– – + + +
+ + – + + +
+ + + – – + +
Ch. Schille et al. / Materials Science and Engineering B 176 (2011) 1797–1801
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Fig. 2. Comparison of the mean gravimetric weight change (gain or loss, respectively) after 6 h immersion in PBSminus , cleaning after 6 h immersion in PBSminus and after 6 h Chandler-loop in human whole blood for each magnesium alloy tested. Bars represent means and s.d. from 6 samples per group for the PBSminus data and 24 samples for blood data, respectively.
Fig. 1. Calculated weight gain during 6 h immersion in PBSminus for each magnesium alloy tested. Each data point represents means and standard deviation from 6 samples.
peel-off-effect of these corrosion layers could be detected on MgAl9 and MgNd2 samples. 3.1.2. Comparison of results between the static immersion test in PBSminus and the dynamic with Chandler-loop in human whole blood In Fig. 2, data for the gravimetric weight loss after (a) 6 h in PBSminus , (b) after cleaning the specimen from this test with distilled water for 10 s and, subsequently, for 3 min with ethanol in an ultrasonic bath, and (c) after 6 h Chandler-loop are presented. It is obvious, that the gravimetric results after 6 h Chandler-loop in human whole blood are completely different compared to the static immersion test in PBSminus . With the Chandler-loop, in human whole blood a real weight loss of all magnesium alloys tested could be measured. In contrast to the static immersion test in PBSminus , the alloys MgAl9 and MgAl9Zn1 showed the lowest weight loss with −260 ± 81 and −202 ± 96 g/cm2 after 6 h Chandler-loop in human whole blood. Similar results were found for the ion release data [6], which were additionally measured for all magnesium alloys tested. On the other hand, after 6 h static immersion test in PBSminus and cleaning, the difference in weight before and after cleaning for these two alloys was 3674 ± 1154 and 573 ± 64 g/cm2 . By cleaning the specimens from the static immersion test in PBSminus the corrosion product layer was partly dissolved from the surface of all magnesium alloys tested, which can also be seen in the microscopic pictures (Fig. 3). This is obviously the reason why the gravimetric values after cleaning reach nearly the starting weight
and a real weight loss of the samples, with exception of MgAl9, could not be measured. The magnesium alloy MgAl9 showed a very high weight loss after cleaning. But this alloy also showed a very thick corrosion layer after 6 h (Fig. 3) and a peel-off-effect after 4 h (Fig. 1), which has an influence on these results. After cleaning, the corrosion layer was significantly reduced. This alloy was the only one, where the solution turned white coloured after 2 h of immersion, indicating an intense corrosion process. After 4 h immersion, peel-off-effects could be seen, because small parts of the alloys and the precipitates appeared in the solution. Additional analytical measurements of the electrolyte after every hour of immersion, which were performed for all magnesium alloys tested, were not possible with this alloy due to the strong reaction with PBSminus [6]. These results showed that a ranking of all these magnesium alloys based on data from in vitro corrosion tests is strongly influenced by the electrolyte and the kind of measurement (static or dynamic) used in this study. Xu et al. [9,10] investigated in vitro and in vivo three different Mn-containing magnesium alloys. For the in vitro study a phosphate buffer solution for the immersion test was used. With this simulated body fluid a weight increase was measured, after cleaning with chromic acid a weight loss could be measured. However, the in vivo animal study showed only degradation with the same alloys. In this in vitro study precipitates also could be found, but they were not specified. 3.2. Surface morphology After the Chandler-loop tests, a colour change of all alloys could be observed. Generally, the surfaces could be divided into two groups. The magnesium alloys containing rare earth elements and the alloy MgZn1 were black stained, and the alloys containing Al were light-coloured. This qualitative observation differed from the immersion tests in PBSminus revealing grey–green discolouration with all alloys tested. Surface morphology of the samples differed also. From all eight magnesium alloys tested, the microscopic pictures (magnification: 32×) of the alloys Mg1Zn, MgAl9 and MgAl9Zn1 were chosen as representatives and are presented in Fig. 3. All alloys showed a different metal surface morphology after 6 h Chandler-loop. The metal surface of MgAl9 was inhomogeneous and rough, on the contrary, for MgAl9Zn1 the metal surface was homogenous, smooth and the grains were smaller. The metal surface of MgZn1 is black
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Ch. Schille et al. / Materials Science and Engineering B 176 (2011) 1797–1801
Fig. 3. Microscopic pictures (magnification: 32×) of the magnesium alloys MgZn1, MgAl9 and MgAl9Zn1 with respect to the used test methods: (A) after 6 h Chandler-loop in human whole blood, (B) after 6 h immersion in PBSminus , (C) after cleaning (10 s with distilled water, 3 min in ethanol in an ultrasonic bath).
coloured. With all alloys, no corrosion layer could be seen microscopically (32×) after blood contact. On the contrary, after 6 h immersion in PBSminus , all metal surfaces were covered with a corrosion layer of different thickness for each alloy. After cleaning, the corrosion layer was only partly dissolved. Obviously, in contrast to the results obtained with blood, dissolution of Mg alloys in PBSminus leads to precipitation of Mg-containing corrosion product on the surface. Thus, dissolving of magnesium by the reaction between magnesium alloy and PBSminus solution followed by precipitating of Mg-containing phosphate layer or product from solution on the surface strongly influences the overall corrosion behaviour [9]. It is proposed, that due to the reaction of magnesium hydroxide a pH increase occurs at the Mg alloy surface promoting the formation of insoluble magnesium phosphate precipitates, Mg3 (PO4 )2 [11]. 3.3. Statistics From the gravimetric results after 6 h Chandler-loop in human whole blood, t-tests with a level of p < 0.05 were done with all magnesium alloys tested. The results can be seen in Table 3. No significant difference was found between MgAl9 and MgAl9Zn1, MgNd2 and MgY4 compared with MgY4Nd2. All the other combinations of the alloys were significantly different. 4. Conclusion The results in the Chandler-loop system using fresh human blood indicate that static immersion tests in artificial simulated body fluids like PBSminus are not suitable to simulate corrosion behaviour of Mg alloys under in vivo conditions.
While the Mg alloys tested in PBSminus under static conditions revealed weight gain due to the formation of insoluble corrosion layers, the same alloys showed a weight loss when blood was used as corrosion medium and the test was performed under dynamic conditions. Generally, in the blood test system weight loss occurred of the Mg alloys investigated. In contrast to blood contact, the Mg alloys tested in PBSminus under static condition revealed weight gain due to the formation of corrosion layers. These layers could be removed partly by ultrasonic cleaning in water. With regard to the different chemical compositions of the magnesium alloys investigated here, it was found that in contact with human whole blood the higher Al content in MgAl9 type alloys reduced the weight loss considerably compared to MgAl3 type alloys. On the other hand, the addition of Zn to MgAl3 changed corrosion stability for the worse. This could not be found with MgAl9Zn1. Rare-earth elements appeared in the middle range under the test condition described here. Whereas in blood MgAl9 showed low corrosion, this alloy showed the highest weight loss in PBSminus . Thus, a corrosionpromoting effect of Al is indicated in this test system. This is also supported by the finding of higher weight loss of MgAl9 compared to MgAl3 in PBSminus after cleaning In summary, it could be demonstrated that there are significant differences in the influence alloying elements on corrosion behaviour of Mg alloys between human whole blood and the simulated body fluid used here. This study revealed that it is difficult to evaluate the corrosion behaviour of magnesium alloys using just standard tests in simply composed simulated body fluids as they are used often in degradation studies. In future studies, a more
Ch. Schille et al. / Materials Science and Engineering B 176 (2011) 1797–1801
physiologic kind of simulated body fluid and a flow corrosion cell should be preferred for corrosion measurements of magnesium alloys for biomedical applications. Acknowledgement This study was partly supported by AiF (project no: KF0548101PK7). References [1] N.T. Kirkland, J. Lespagnol, N. Birbilis, M.P. Staiger, Corros. Sci. 52 (2010) 287–294. [2] W.-D. Mueller, L. Nascimento, L. de Mele, M. Fernandez, Acta Biomater. 6 (2010) 1749–1755.
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[3] Y. Chen, S. Zhang, J. Li, Y. Song, C. Zhao, X. Zhang, Mater. Lett. 64 (2010) 1996–1999. [4] J. Levesque, D. Dube, M. Fiset, D. Mantovani, Mater. Sci. Forum 426–432 (2003) 521–526. [5] C. Schille, H.-P. Reichel, N. Hort, C. Füger, J. Geis-Gerstorfer, in: K. Kainer (Ed.), Magnesium, Wiley, 2009, pp. 1195–1200, ISBN 978-3-527-32732-4. [6] J. Geis-Gerstorfer, Ch. Schille, E. Schweizer, F. Rupp, L. Scheideler, H.-P. Reichel, N. Hort, A. Nolte, H.-P. Wendel,: Blood triggered Corrosion of Magnesium Alloys, Mater. Sci. Eng. B (2011), same issue. [7] S. Sinn, T. Scheuermann, G. Ziemer, H.P. Wendel, Biomaterialien 10 (3/4) (2010) 148. [8] P. Schmutz, N.-C. Quach-Vu, I. Gerber, Electrochem. Soc. Interface (Summer) (2008) 35–40. [9] L. Xu, E. Zhang, D. Yin, S. Zheng, K. Yang, J. Mater. Sci.: Mater. Med. 19 (2008) 1017–1025. [10] L. Xu, G. Yu, E. Zhang, F. Pan, K. Yang, J. Biomed. Mater. Res. 83A (2007) 703– 711. [11] Y. Xin, C. Liu, X. Xang, G. Tang, X. Chu, J. Mater. Res. 22 (2007) 2004–2011.
Contents lists available at ScienceDirect
Materials Science and Engineering B journal homepage: www.elsevier.com/locate/mseb
Short communication
Corrosion of experimental magnesium alloys in blood and PBS: A gravimetric and microscopic evaluation Ch. Schille a,∗ , M. Braun b , H.P. Wendel b , L. Scheideler a , N. Hort c , H.-P. Reichel d , E. Schweizer a , J. Geis-Gerstorfer a a
University Hospital Tuebingen, Center for Dentistry, Oral Medicine and Maxillofacial Surgery, Section Medical Materials & Technology, Osianderstr. 2-8, D-72076 Tübingen, Germany University Hospital Tuebingen, Div. Congenital & Paediatric Cardiac Surgery, University Children’s Hospital, Tuebingen, Germany, Calwerstr. 7/1, D-72076 Tübingen, Germany c GKSS Research Centre, Institute of Materials Research, Max-Planck-Str. 1, D-21502 Geesthacht, Germany d Weissensee Company, Bürgermeister-Ebert-Str. 30-32, D-36124 Eichenzell, Germany b
a r t i c l e
i n f o
Article history: Received 3 November 2010 Received in revised form 11 March 2011 Accepted 10 April 2011 Available online 28 April 2011 Keywords: Gravimetry Magnesium alloys Static immersion test Chandler-loop
a b s t r a c t Corrosion tests for medical materials are often performed in simulated body fluids (SBF). When SBF are used for corrosion measurement, the open question is, how well they match the conditions in the human body. The aim of the study was to compare the corrosion behaviour of different experimental magnesium alloys in human whole blood and PBSminus (phosphate buffered saline w/o Ca and Mg) as a simulated body fluid by gravimetric weight measurements and microscopic evaluation. Eight different experimental magnesium alloys, containing neither Mn nor other additives, were manufactured. With these alloys, a static immersion test in PBSminus and a dynamic test using the Chandler-loop model with human whole blood over 6 h were performed. During the static immersion test, the samples were weighed every hour. During the dynamic test, the specimens were weighed before and after the 6 h incubation period in the Chandler-loop. From both tests, the total mass change was calculated for each alloy and the values were compared. Additionally, microscopic pictures from the samples were taken at the end of the test period. All alloys showed different corrosion behaviour in both tests, especially the alloys with high aluminium content, MgAl9 and MgAl9Zn1. Generally, alloys in PBS showed a weight gain due to generation of a microscopically visible corrosion layer, while in the blood test system a more or less distinct weight loss was observed. When alloys are ranked according to corrosion susceptibility, the results differ also between the test systems. The MgAl9 alloy, showing the most pronounced corrosion in PBS, was one of the least corroding alloys under simulated in vivo conditions in blood. Thus, the ranking concerning clinical suitability of the magnesium alloys tested in this study is different, depending on the used electrolyte and the kind of method. For a possible clinical use, the alloy MgAl9Zn1 might be preferable for further investigations. © 2011 Elsevier B.V. All rights reserved.
1. Introduction One problem of magnesium alloys for biomedical applications is their fast degradation. To test the degradation in vitro, different kinds of corrosion measurements can be applied, such as static immersion or electrochemical tests. Most in vitro corrosion studies of magnesium alloys were performed in different simulated body fluids (SBF) like phosphate buffered saline (PBS), Hanks or Ringer solution, artificial plasma, etc. Different pH values and composition of the simulated body fluids influence the corrosion behaviour, as well as kind of fabrication, composition, grain size, finishing, heat treatment, coating, etc. of magnesium alloys. The
∗ Corresponding author. Tel.: +49 7071 2983493/2986199; fax: +49 7071 295775. E-mail address: [email protected] (Ch. Schille). 0921-5107/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.mseb.2011.04.007
advantage of using simulated body fluids for in vitro studies is that they can be easily handled and provide fast first information [3,4]. The open question is whether these results correspond to results obtained under in vivo conditions [2], since the compositions of SBFs are rather simple and do not resemble the much more complex composition of body liquids like blood or interstitial fluid. For biomaterials with an intended application in the blood stream, the dynamic flow is an additional issue of the surrounding corrosion medium, so that static immersion tests are of limited value. There are only few publications about corrosion measurements under dynamic conditions. However, these measurements were also performed with simulated body fluids [3,4]. We therefore looked for a model to simulate clinical conditions, using a real body fluid instead of SBF under dynamic testing conditions and set out to compare this system with static immersion test conditions using SBF.
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Ch. Schille et al. / Materials Science and Engineering B 176 (2011) 1797–1801
Table 1 Ion concentration of PBSminus compared to human whole blood [8]. Ion
Blood
PBSminus
+
142.0 5.0 1.5 2.5 103.0 27.0 1.0 0.5
146.0 4.1 – – 140.6 – 9.5 –
Na K+ Mg2+ Ca2+ Cl− (HCO3 )− (HPO4 )2− SO4 2− Concentration in mmol/l.
The Chandler-loop is a measurement system for testing the hemocompatibility of stents under dynamic flow conditions and in human whole blood [6,7]. The aim of this in vitro study was to compare the corrosion results of eight different experimental magnesium alloys after 6 h immersion in an artificial, simulated body fluid (PBSminus , phosphate buffered saline without Mg+ and Ca+ ), with the results from corrosion measurements in human blood. For comparison, the ion concentrations of blood and PBSminus are presented in Table 1 [8]. 2. Materials and methods The composition of the experimental magnesium alloys, which were used in this study can be seen in Table 2. The alloys resemble well-known technical alloys like e.g. AZ 91, but were manufactured without additives like e.g. Mn, they contain only the main alloying elements Mg, Al, Zn, Nd and Y [5]. From each alloy, round sample discs with a diameter of 10 mm and 1 mm height were prepared, polished with SiC 1200 in ethanol on both sides and ultrasonically cleaned in ethanol for 3 min.
were made with a rotation speed of 10 rpm and at a temperature of 37 ◦ C using a thermostat-controlled water bath (Q102, Haake, Berlin). After 6 h the tests were terminated and the samples were cleaned in different solutions (0,9% saline, 1% SDS solution) and stored in ethanol. Before and after the chandler-loop tests the weight of the two samples of each (dried) assembly was measured. 2.2. Gravimetric corrosion tests in PBS For the immersion test in PBSminus (DPBS w/o Ca, Mg; GIBCO (Invitrogen # 14190-169) six specimens of each alloy were investigated. Before onset of the immersion test, the weight of each sample was determined. Each sample was placed into a 15 ml PP tube, filled with 5 ml PBSminus and placed into a heating cabinet at 37 ◦ C. Change of weight (referring to original weight) was measured every hour after drying the samples for 20 min by exposure to air. Subsequently, samples were placed into new PP tubes with fresh electrolyte. 2.3. Microscopic evaluation and statistical calculation After the Chandler-loop and immersion tests, images were taken with a stereo microscope (Wild Heerbrugg) at magnifications of 10, 20 and 32. The gravimetric results of the Chandler-loop test were calculated per unit of area. Differences between means were pairwise tested for statistical significance using Student’s t-test (p < 0.05) for independent samples (Table 3). 3. Results and discussion
2.1. Gravimetric corrosion tests in blood
3.1. Gravimetric results
For the tests with the modified Chandler-loop model 24 specimens were used from each alloy. Samples were clamped pairwise in circular loops of PVC tubes (l = 50 cm), which were closed with a silicon tube. Two parallel sets of silicone loops were used in every experiment (4 samples per alloy). The experiments were repeated six times with blood from three different donors. Each tube was filled with 20 ml fresh human whole blood. The measurements
3.1.1. Immersion test over 6 h in PBSminus Fig. 1 shows the time-dependent gravimetric results over 6 h static immersion. For all magnesium alloys tested, it was found that in PBSminus a weight increase occurred under static immersion conditions. The increase is caused by the formation of a thick corrosion product layer [1], which can be seen in the microscopic pictures (Fig. 3). Fig. 1 also shows that after 4 h a kind of
Table 2 Chemical composition of the magnesium alloys tested. All values are given in wt.% and have been measured using a Spektrolab M (Spektro. Kleve. Germany). Al MgZn1 MgAl3 MgAl9 MgNd2 MgY4 MgAl3Zn1 MgAl9Zn1 MgY4Nd2
Zn
Y
Nd
0.966 3.07 8.43 1.99 4.95 3.01 9.03
1.01 1.16
0.00503 1.96
4.67
Fe
Cu
Ni
Si
Mg
0.00275 0.00311 0.00152 0.00410 0.00576 0.00387 0.00503 0.00480
0.00373 0.00202 0.00295 0.00384 0.00419 0.00249 0.00363 0.00413
0.00206 0.00096 0.00063 0.00193 <0.00020 0.00106 0.00081 0.00112
0.0316 0.0201 0.0154 0.0151 0.0339 0.0332 0.0229 0.0321
Bal. Bal. Bal. Bal. Bal. Bal. Bal. Bal.
Table 3 Statistical results from paired Student’s t-test (p < 0.05) of the gravimetric weight loss data after 6 h Chandler-loop in human whole blood for each magnesium alloy tested. MgZn1 MgZn1 MgAl3 MgAl9 MgNd2 MgY4 MgAl3Zn1 MgAl9Zn1 MgY4Nd2
Mg3Al3
MgAl9
MgNd2
Mg4Y4
Mg3Al3Zn1
MgAl9Zn1
MgY4Nd2
–
+ +
+ + +
+ – + –
– – + + +
+ + – + + +
+ + + – – + +
Ch. Schille et al. / Materials Science and Engineering B 176 (2011) 1797–1801
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Fig. 2. Comparison of the mean gravimetric weight change (gain or loss, respectively) after 6 h immersion in PBSminus , cleaning after 6 h immersion in PBSminus and after 6 h Chandler-loop in human whole blood for each magnesium alloy tested. Bars represent means and s.d. from 6 samples per group for the PBSminus data and 24 samples for blood data, respectively.
Fig. 1. Calculated weight gain during 6 h immersion in PBSminus for each magnesium alloy tested. Each data point represents means and standard deviation from 6 samples.
peel-off-effect of these corrosion layers could be detected on MgAl9 and MgNd2 samples. 3.1.2. Comparison of results between the static immersion test in PBSminus and the dynamic with Chandler-loop in human whole blood In Fig. 2, data for the gravimetric weight loss after (a) 6 h in PBSminus , (b) after cleaning the specimen from this test with distilled water for 10 s and, subsequently, for 3 min with ethanol in an ultrasonic bath, and (c) after 6 h Chandler-loop are presented. It is obvious, that the gravimetric results after 6 h Chandler-loop in human whole blood are completely different compared to the static immersion test in PBSminus . With the Chandler-loop, in human whole blood a real weight loss of all magnesium alloys tested could be measured. In contrast to the static immersion test in PBSminus , the alloys MgAl9 and MgAl9Zn1 showed the lowest weight loss with −260 ± 81 and −202 ± 96 g/cm2 after 6 h Chandler-loop in human whole blood. Similar results were found for the ion release data [6], which were additionally measured for all magnesium alloys tested. On the other hand, after 6 h static immersion test in PBSminus and cleaning, the difference in weight before and after cleaning for these two alloys was 3674 ± 1154 and 573 ± 64 g/cm2 . By cleaning the specimens from the static immersion test in PBSminus the corrosion product layer was partly dissolved from the surface of all magnesium alloys tested, which can also be seen in the microscopic pictures (Fig. 3). This is obviously the reason why the gravimetric values after cleaning reach nearly the starting weight
and a real weight loss of the samples, with exception of MgAl9, could not be measured. The magnesium alloy MgAl9 showed a very high weight loss after cleaning. But this alloy also showed a very thick corrosion layer after 6 h (Fig. 3) and a peel-off-effect after 4 h (Fig. 1), which has an influence on these results. After cleaning, the corrosion layer was significantly reduced. This alloy was the only one, where the solution turned white coloured after 2 h of immersion, indicating an intense corrosion process. After 4 h immersion, peel-off-effects could be seen, because small parts of the alloys and the precipitates appeared in the solution. Additional analytical measurements of the electrolyte after every hour of immersion, which were performed for all magnesium alloys tested, were not possible with this alloy due to the strong reaction with PBSminus [6]. These results showed that a ranking of all these magnesium alloys based on data from in vitro corrosion tests is strongly influenced by the electrolyte and the kind of measurement (static or dynamic) used in this study. Xu et al. [9,10] investigated in vitro and in vivo three different Mn-containing magnesium alloys. For the in vitro study a phosphate buffer solution for the immersion test was used. With this simulated body fluid a weight increase was measured, after cleaning with chromic acid a weight loss could be measured. However, the in vivo animal study showed only degradation with the same alloys. In this in vitro study precipitates also could be found, but they were not specified. 3.2. Surface morphology After the Chandler-loop tests, a colour change of all alloys could be observed. Generally, the surfaces could be divided into two groups. The magnesium alloys containing rare earth elements and the alloy MgZn1 were black stained, and the alloys containing Al were light-coloured. This qualitative observation differed from the immersion tests in PBSminus revealing grey–green discolouration with all alloys tested. Surface morphology of the samples differed also. From all eight magnesium alloys tested, the microscopic pictures (magnification: 32×) of the alloys Mg1Zn, MgAl9 and MgAl9Zn1 were chosen as representatives and are presented in Fig. 3. All alloys showed a different metal surface morphology after 6 h Chandler-loop. The metal surface of MgAl9 was inhomogeneous and rough, on the contrary, for MgAl9Zn1 the metal surface was homogenous, smooth and the grains were smaller. The metal surface of MgZn1 is black
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Fig. 3. Microscopic pictures (magnification: 32×) of the magnesium alloys MgZn1, MgAl9 and MgAl9Zn1 with respect to the used test methods: (A) after 6 h Chandler-loop in human whole blood, (B) after 6 h immersion in PBSminus , (C) after cleaning (10 s with distilled water, 3 min in ethanol in an ultrasonic bath).
coloured. With all alloys, no corrosion layer could be seen microscopically (32×) after blood contact. On the contrary, after 6 h immersion in PBSminus , all metal surfaces were covered with a corrosion layer of different thickness for each alloy. After cleaning, the corrosion layer was only partly dissolved. Obviously, in contrast to the results obtained with blood, dissolution of Mg alloys in PBSminus leads to precipitation of Mg-containing corrosion product on the surface. Thus, dissolving of magnesium by the reaction between magnesium alloy and PBSminus solution followed by precipitating of Mg-containing phosphate layer or product from solution on the surface strongly influences the overall corrosion behaviour [9]. It is proposed, that due to the reaction of magnesium hydroxide a pH increase occurs at the Mg alloy surface promoting the formation of insoluble magnesium phosphate precipitates, Mg3 (PO4 )2 [11]. 3.3. Statistics From the gravimetric results after 6 h Chandler-loop in human whole blood, t-tests with a level of p < 0.05 were done with all magnesium alloys tested. The results can be seen in Table 3. No significant difference was found between MgAl9 and MgAl9Zn1, MgNd2 and MgY4 compared with MgY4Nd2. All the other combinations of the alloys were significantly different. 4. Conclusion The results in the Chandler-loop system using fresh human blood indicate that static immersion tests in artificial simulated body fluids like PBSminus are not suitable to simulate corrosion behaviour of Mg alloys under in vivo conditions.
While the Mg alloys tested in PBSminus under static conditions revealed weight gain due to the formation of insoluble corrosion layers, the same alloys showed a weight loss when blood was used as corrosion medium and the test was performed under dynamic conditions. Generally, in the blood test system weight loss occurred of the Mg alloys investigated. In contrast to blood contact, the Mg alloys tested in PBSminus under static condition revealed weight gain due to the formation of corrosion layers. These layers could be removed partly by ultrasonic cleaning in water. With regard to the different chemical compositions of the magnesium alloys investigated here, it was found that in contact with human whole blood the higher Al content in MgAl9 type alloys reduced the weight loss considerably compared to MgAl3 type alloys. On the other hand, the addition of Zn to MgAl3 changed corrosion stability for the worse. This could not be found with MgAl9Zn1. Rare-earth elements appeared in the middle range under the test condition described here. Whereas in blood MgAl9 showed low corrosion, this alloy showed the highest weight loss in PBSminus . Thus, a corrosionpromoting effect of Al is indicated in this test system. This is also supported by the finding of higher weight loss of MgAl9 compared to MgAl3 in PBSminus after cleaning In summary, it could be demonstrated that there are significant differences in the influence alloying elements on corrosion behaviour of Mg alloys between human whole blood and the simulated body fluid used here. This study revealed that it is difficult to evaluate the corrosion behaviour of magnesium alloys using just standard tests in simply composed simulated body fluids as they are used often in degradation studies. In future studies, a more
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physiologic kind of simulated body fluid and a flow corrosion cell should be preferred for corrosion measurements of magnesium alloys for biomedical applications. Acknowledgement This study was partly supported by AiF (project no: KF0548101PK7). References [1] N.T. Kirkland, J. Lespagnol, N. Birbilis, M.P. Staiger, Corros. Sci. 52 (2010) 287–294. [2] W.-D. Mueller, L. Nascimento, L. de Mele, M. Fernandez, Acta Biomater. 6 (2010) 1749–1755.
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[3] Y. Chen, S. Zhang, J. Li, Y. Song, C. Zhao, X. Zhang, Mater. Lett. 64 (2010) 1996–1999. [4] J. Levesque, D. Dube, M. Fiset, D. Mantovani, Mater. Sci. Forum 426–432 (2003) 521–526. [5] C. Schille, H.-P. Reichel, N. Hort, C. Füger, J. Geis-Gerstorfer, in: K. Kainer (Ed.), Magnesium, Wiley, 2009, pp. 1195–1200, ISBN 978-3-527-32732-4. [6] J. Geis-Gerstorfer, Ch. Schille, E. Schweizer, F. Rupp, L. Scheideler, H.-P. Reichel, N. Hort, A. Nolte, H.-P. Wendel,: Blood triggered Corrosion of Magnesium Alloys, Mater. Sci. Eng. B (2011), same issue. [7] S. Sinn, T. Scheuermann, G. Ziemer, H.P. Wendel, Biomaterialien 10 (3/4) (2010) 148. [8] P. Schmutz, N.-C. Quach-Vu, I. Gerber, Electrochem. Soc. Interface (Summer) (2008) 35–40. [9] L. Xu, E. Zhang, D. Yin, S. Zheng, K. Yang, J. Mater. Sci.: Mater. Med. 19 (2008) 1017–1025. [10] L. Xu, G. Yu, E. Zhang, F. Pan, K. Yang, J. Biomed. Mater. Res. 83A (2007) 703– 711. [11] Y. Xin, C. Liu, X. Xang, G. Tang, X. Chu, J. Mater. Res. 22 (2007) 2004–2011.