Communication pubs.acs.org/IC
A Water-Soluble Cationic Zinc Lysine Precursor for Coating ZnO on Biomaterial Surfaces Shaotang Yuan,† Shiri Nawrocki,† Michael Stranick,† Ying Yang,† Chong Zheng,‡ James G. Masters,† and Long Pan*,† †
Colgate−Palmolive Company, 909 River Road, Piscataway, New Jersey 08854, United States Department of Chemistry and Biochemistry, Northern Illinois University, DeKalb, Illinois 60115, United States
‡
S Supporting Information *
homogeneously. Consequently, the development of a novel alternative method to firmly deposit insoluble ZnO on the surfaces of biomaterials at milder conditions, such as at room temperature and neutral pH, remains as a major challenge for researchers. It is known that Zn ions tend to bind to many proteins found in humans.24 Because most biological surfaces are negatively charged,25,26 the synthetic premise was to design a soluble cationic zinc coordination compound that can bind to surfaces of most biomaterials by electrostatic interaction and then generate ZnO in situ via hydrolysis. As building blocks of proteins, amino acids have been widely used in pharmaceutical, personal care, and oral care products. Therefore, understanding the structure of zinc−amino acid coordination compounds is of critical importance. Since Kretsinger and Cotton determined the first crystal structure of these types in 1963,27 many structures of other zinc−amino acid coordination compounds have been solved. However, it should be noted that a majority of these zinc−amino acid compounds are neutral. In fact, only a few cationic zinc compounds containing organic ligands have been reported.28−31 Alagha et al. reported the first cationic zinc− arginine coordination compound,32 which was synthesized by mixing zinc acetate [Zn(OAc)2] and L-arginine (L-Arg) in an anhydrous methanol solution. The resulting compound, [Zn(OAc)(L-Arg)2]OAc·3H2O, was insoluble in water. The insolubility not only renders uniform aqueous surface deposition onto biological surfaces nearly impossible but also leverages hydrolysis of the compound to form ZnO impractical. This finding prompted the need to investigate the synthesis of a watersoluble zinc−amino acid coordination compound that is compatible with biomaterials. In this Communication, we report the synthesis of a water-soluble, cationic, zinc−amino acid coordination compound by reacting ZnO with L-lysine hydrochloride to produce [Zn(C6H14N2O2)2Cl]Cl·2H2O (1). It will be further shown that 1 can generate ZnO in situ on the surfaces of biomaterials through a facile hydrolysis reaction. In addition, we also demonstrate its outstanding property as a precursor for depositing an antibacterial ZnO film on arbitrary biomaterial surfaces in situ utilizing a facile and safe route. The reaction equation to form 1 is
ABSTRACT: A novel water-soluble cationic zinc lysine coordination compound, [Zn[(C6H14N2O2)]2Cl]Cl·2H2O (1), has been designed and synthesized and its crystal structure determined. The aqueous solution of this coordination compound is not only transparent and stable at room temperature but it is also nearly neutral (pH ∼ 7). It is worth noting that zinc oxide (ZnO) forms in situ upon dilution of a solution of the compound. The bioactivity of ZnO has been confirmed using an Alarma Blue assay. These unique properties allow the coordination compound to gently grow ZnO coating with excellent antibacterial benefits onto biomaterial surfaces in a facile and safe manner.
Z
inc oxide (ZnO) is a remarkable semiconductor and piezoelectric material that has attracted immense interest because of its many applications in the electronic and optoelectronic field.1−12 In addition, ZnO is also a very important active ingredient in pharmaceutical and cosmetic products because of its antibacterial and antimicrobial benefits, as well as for providing UV protection.13−16 In many commercial zinc-containing products, insoluble ZnO particles are suspended in the formula and applied to the surfaces of interest in the form of an ointment, cream, suspension, etc. In such cases, ZnO cannot be retained on the surfaces for extended periods of time because of its easy removal by water and/or other external factors. This low retention of ZnO on the surfaces provides impetus for us to seek a new method to firmly deposit ZnO on surfaces, particularly those of biomaterials. Over the past few decades, significant efforts have been devoted to obtaining uniform, high-quality ZnO films and then depositing them onto other materials. These techniques include metal−organic chemical vapor deposition,17,18 plasma-enhanced chemical vapor deposition,19 pulsed-laser deposition,20 atomic layer deposition,21 etc.22,23 However, all of these techniques require high energy input, and the deposition process normally occurs at high temperatures (>300 °C). As a result, these severe conditions hinder the practicality of such methods for the deposition of ZnO onto biomaterials, such as human skin, teeth, and hair. On the other hand, the wet chemistry method of reacting soluble zinc salts with base can produce zinc hydroxide, which, at low temperatures, gradually dissociates to ZnO. Nevertheless, the basic pH of such a solution may cause irritation, which also inhibits its practical use on biomaterials © XXXX American Chemical Society
Received: July 12, 2016
A
DOI: 10.1021/acs.inorgchem.6b01663 Inorg. Chem. XXXX, XXX, XXX−XXX
Communication
Inorganic Chemistry
In order to reveal the unique properties of 1, we explored and compared its hydrolysis reaction to that of other zinc−amino acid mixtures containing similar compositions under the same conditions. Because the hydrochloride salt forms of arginine (Arg), cysteine (Cys), and glycine (Gly) are commercially available, we prepared ZnO-Arg·HCl, ZnO-Cys·HCl, and ZnOGly·HCl solutions for comparison with 1. Details of the preparations are summarized in the Supporting Information. These solutions were diluted to a zinc concentration of 2% by weight. The pH values of the solutions were 7.4, 7.3, 2.9, and 3.9 for 1, ZnO-Arg·HCl, ZnO-Cys·HCl, and ZnO-Gly·HCl, respectively (see Table S2 for details). Among all of the samples, the ZnO-Arg·HCl solution was the only one that appeared turbid, although both arginine and lysine fall into the same amino acid category. The instability of the ZnO-Arg·HCl solution differentiates it from that of stable 1 even though both share a similar final pH. 1, ZnO-Cys·HCl, and ZnO-Gly·HCl were then further diluted 2-, 4-, 8-, 16-, and 32-fold by deionized water, and each individual solution was placed in the turbidity meter to monitor the formation of precipitates over a period of 30 min at 37 °C. The pH of each dilution is provided in Table S3. No precipitation was found in any of the ZnO-Cys·HCl or ZnO-Gly· HCl dilutions because of the low pH. Figure S2 shows a comparison of the percent transmission of each 1 dilution. As the figure shows, there was an increase in precipitation as the solution became more diluted up to 16-fold. Further dilution to 32-fold revealed a higher percent transmission due to the excess amount of water with the precipitate. The 16-fold dilution product of 1 was relatively stable in the first 15 min; however, a rapid decrease in the percent transmission in the next 10 min suggested that a large amount of precipitate was formed in a short period of time. The white precipitate was isolated and collected by filtration. The PXRD pattern of this precipitate, shown in Figure 2,
ZnO + 2C6H15N2O2 Cl + H 2O → [Zn(C6H14N2O2 )2 Cl]Cl ·2H 2O
An aqueous solution of 1 is transparent and has a nearly neutral pH (7.0−7.4). The crystal structure of 1 is determined using single-crystal X-ray diffraction and shown in Figure 1. The
Figure 1. Molecular structure of complex 1. Color code: Zn, green; O, red; N, blue; C, gray; Cl, yellow; H, neon green.
selected crystallographic data and bond lengths and angles of 1 are summarized in Table 1 and Table S1. In this complex, the Zn Table 1. Crystallographic Data for Compound 1 empirical formula fw temperature (K) wavelength (Å) cryst syst space group unit cell dimens a (Å) b (Å) c (Å) volume (Å3) Z calcd density (g/cm3) abs coeff (mm−1) θ range for data collection (deg) data/restraints/param GOF on F2 final R indices [I > 2σ(I)] R indices (all data)
C12H32N4O6Cl2Zn 464.69 293(2) 0.71073 monoclinic P21 5.275(2) 17.040(5) 11.407(3) 1022.5(5) 2 1.509 1.496 1.79−24.99 3468/1/230 1.180 R1 = 0.0318, wR2 = 0.0816 R1 = 0.0340, wR2 = 0.0830
ion is coordinated by two lysine ligands with two N atoms from its amine groups and O atoms from the carboxylic groups, which are in a trans position from each other in an equatorial plane. It is interesting to discover that a Cl ligand is apically coordinated to the Zn ion, which leads to the distorted square-pyramidal geometry. This novel structure gives rise to a positive cation moiety as expected, to which a second Cl− ion acts as the counterion to form an ionic salt. The powder X-ray diffraction (PXRD) pattern of the product powder agrees well with simulated results from the single-crystal structure (Figure S1). In addition, elemental analyses of C, H, N, and Zn ions from the crystal match with the calculated values (see the Characterization section in the Supporting Information). These instrumental analyses indicate the high purity of the crystal. In contrast to the insoluble zinc(II) arginine complex reported by Alagha et al.,30 the water-soluble nature of complex 1 provides great opportunities for potential applications.
Figure 2. Comparison of the PXRD pattern of the 1 precipitate (blue) to that of the ZnO standard (red). The result clearly shows that the precipitate formed from hydrolysis of 1 is ZnO.
matches that of wurtzite ZnO very well. This finding strongly indicates the formation of ZnO by the hydrolysis reaction of 1. Additionally, the filtrate gave rise to colorless column-shaped single crystals, which were analyzed crystallographically. A lysine hydrochloride dihydrate crystal structure was confirmed using single-crystal X-ray diffraction (Figure S4). When the findings from the PXRD pattern of the precipitate and the X-ray diffraction pattern of the crystal found in the filtrate are B
DOI: 10.1021/acs.inorgchem.6b01663 Inorg. Chem. XXXX, XXX, XXX−XXX
Communication
Inorganic Chemistry
applied after the fingers were air-dried. The changing of the indicator color from red-orange to blue-purple indicates the detection of zinc. Figure S4 clearly demonstrates that both ZnO and ZnCl2 were easily removed from the skin by simple rubbing under running water because of the lack of blue color after Zincon was sprayed. In contrast, the treatment with 1 showed a firm deposition of zinc on the skin, as the prominent blue-purple color confirmed. In order to compare the zinc bioactivity of deposited zinc from 1 against regular ZnO, the antibacterial effects of the zinc activity were also examined. An in vitro study was conducted on pig skin using an Alamar Blue assay.33 The percentage of reduction was defined based on nontreatment as the control. As shown in Table 2, the results suggest that, after using the 1 formula, the residue
combined, the hydrolysis reaction of 1 can be described as follows: [Zn(lysine)2 Cl]Cl + H 2O ⇄ ZnO + 2lysine ·HCl
It is hypothesized that, in this reversible reaction, the aqua ligands are competing with both lysinate and chloride in 1 to form a zinc−aqua coordination compound, which, in turn, converts to zinc hydroxide and further hydrolyzes to ZnO. The equilibrium of the reaction depends on the amount of water in the solution. As a result, a stock solution of 1 with 2.4% zinc appeared transparent, indicating no formation of ZnO. However, as the solution was diluted, aqua ligands became dominant and were able to move the equilibrium to the right and trigger the formation of ZnO. At 16-fold dilution, with about 0.13% zinc, there was a sufficient amount of water to replace the coordinated chloride and lysinate. As a result, optimal amounts of precipitate at this dilution were observed. Further dilution completed the hydrolysis reaction; however, a lower amount of ZnO was detected because of excess dilution. The unique process of forming ZnO in situ by diluting precursor 1 in water provides a novel, facile, safe, and costeffective method to grow ZnOs coating on the surfaces of any inorganic or biological substance. This process has inherent advantages over previously published methods because it occurs at neutral pH and ambient conditions. Instead of triggering the deposition reaction by heat or pH change, simple dilution of the precursor solution under ambient conditions leads to ZnO deposition. As a result, 1 has the propensity for many potential applications in pharmaceutical and cosmetic products; e.g., it can readily deposit ZnO on human skin, hair, and teeth. A conceptual explanation for the ZnO deposition mechanism is presented in Scheme 1. It is proposed that (1) a positively charged 1 molecule
Table 2. Effect of Deposited Zinc on S. aureusa
1 ZnO placebo a
concn of Zn ions (%)
mean of % reduction vs nontreatment control
standard deviation (%)
1.0 1.0 0
48 29 −9
8 6 21
The results are the mean of triplicate experiments.
left over on the pig skin inhibited Staphylococcus aureus better than the ZnO formula and placebo. This directly shows that ZnO deposited from a 1 solution possessed a much better antibacterial efficacy than regular ZnO. In summary, we illustrate the design, synthesis, and characterization of the first water-soluble cationic zinc lysine coordination compound 1 as a precursor for ZnO deposition. This novel coordination compound can be hydrolyzed to form ZnO when the solution is simply diluted with water. The in situ generation of ZnO takes place at neutral pH and under mild conditions, which is highly compatible with biomaterials. This method paves a new avenue to gently coat insoluble ZnO on the surfaces of biomaterials and other arbitrary 3D materials in a facile, efficient, and safe manner. Moreover, the reprecipitated zinc is more effective on bacterial inhibition than the parent compound ZnO. Further research will seek to understand the details of the deposition process and the mechanism of the antibacterial effect of a1 1-deposited ZnO film, as well as to develop 1 for consumer products applications.
Scheme 1. Proposed Mechanism of In Situ ZnO Formation by Hydrolysis of 1 on a Biomaterial Surface
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ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.inorgchem.6b01663. PXRD data and experimental details (PDF) Crystallographic data (CCDC 1022208) (CIF)
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first attaches to the negatively charged biomaterial surface by electrostatic attraction, (2) becaue of dilution, 1 is surrounded by large amounts of water molecules, which trigger the hydrolysis reaction, and (3) ZnO is formed in situ on the surface of biomaterials. The deposition of zinc on human skin has been demonstrated using an indicator test. Human fingers were simultaneously soaked in aqueous 1, a ZnCl2 solution, and a ZnO suspension under the same conditions for 30 s. The treated fingers were then rinsed under running water for 30 s. An indicator (Zincon) was
AUTHOR INFORMATION
Corresponding Author
*E-mail:
[email protected]. Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS The authors acknowledge Dr. Laurence Du-Thumm and Dr. Ravi Subramanyam for their support and many useful discussions. C
DOI: 10.1021/acs.inorgchem.6b01663 Inorg. Chem. XXXX, XXX, XXX−XXX
Communication
Inorganic Chemistry
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DOI: 10.1021/acs.inorgchem.6b01663 Inorg. Chem. XXXX, XXX, XXX−XXX
