Subscriber access provided by UNIV OF NEW ENGLAND ARMIDALE
pH mapping on tooth surfaces for quantitative caries diagnosis using micro Ir/IrOx pH sensor Chindanai Ratanaporncharoen, Miyuki Tabata, Yuichi Kitasako, Masaomi Ikeda, Tatsuro Goda, Akira Matsumoto, Junji Tagami, and Yuji Miyahara Anal. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.analchem.8b00867 • Publication Date (Web): 07 Mar 2018 Downloaded from http://pubs.acs.org on March 7, 2018
Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.
is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.
Page 1 of 8 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Analytical Chemistry
pH mapping on tooth surfaces for quantitative caries diagnosis using micro Ir/IrOx pH sensor Chindanai Ratanaporncharoen†, Miyuki Tabata†, Yuichi Kitasako‡, Masaomi Ikeda‡, Tatsuro Goda†, Akira Matsumoto†, Junji Tagami‡ & Yuji Miyahara*,† †
Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Tokyo, Japan. Faculty of Dentistry, Tokyo Medical and Dental University, Tokyo, Japan. Phone: +81 3 5280 8095 FAX: +81 3 5280 8135 Email:
[email protected] ‡
ABSTRACT: A quantitative diagnostic method for dental caries would improve oral health, which directly affects the quality of life. Here we describe the preparation and application of Ir/IrOx pH sensors, which are used to measure the surface pH of dental caries. The pH level is used as an indicator to distinguish between active and arrested caries. After a dentist visually inspected and defined 18 extracted dentinal caries at various positions as active or arrested caries, the surface pH values of sound and caries areas were directly measured with an Ir/IrOx pH sensor with a diameter of 300 µm as a dental explorer. The average pH values of the sound root, the arrested caries, and active caries were 6.85, 6.07, and 5.30 respectively. The pH obtained with an Ir/IrOx sensor was highly correlated with the inspection results by the dentist, indicating that the types of caries were successfully categorized. This caries testing technique using a micro Ir/IrOx pH sensor provides an accurate quantitative caries evaluation and has potential in clinical diagnosis.
According to the World Health Organization (WHO), people around the world suffer from dental caries, which is also known as tooth decay, due to the lack of oral hygiene in daily life 1,2. Briefly, dental caries are mainly caused by the interaction of cariogenic bacteria and carbohydrates on the tooth surface. The bacteria produce organic acid as a byproduct, which interferes with the balance of the tooth restoration mechanism2-4. Early detection is necessary to maximize the quality of life and minimize tooth loss. In a clinical office, the contemporary method to detect caries is based on a visual inspection using a tool called a dental explorer5,6. The results of the visual method depend on professional experience. Although this skill is fundamental and necessary for every dentist, quantitative and reasonable estimates to diagnose carious lesions have yet to be established. Furthermore, improper use of the tool may cause fractures and infection7-9. Therefore, clinical demands for precise and quantitative diagnostic methods have been increasing from the viewpoint of available interventions for prevention and nonsurgical treatment of dental caries. The dentists have used several methods in combination with the visual inspection. Radiographic examination can detect mineral density loss as a gray-scale mapped image10-12. However, early caries and enamel lesions are undetectable due to the low sensitivity and selectivity of this method. A false-positive or a false-negative affects the treatment plan. To improve the accuracy of detection, several methods have been proposed based on pH13,14 and fluorescence12,15. Of these techniques, pH detection is directly related to the state of caries (i.e., bacterial activity16). Several kinds of electrodes and systems, such as an antimony electrode17,18, pH imaging19, and ion-sensitive field-effect transistor (ISFET)20-22, have been used in studies on pH measurements of dental caries. However, the antimony electrode ex-
hibits toxicity toward an organism17,23. Additionally, the pH imaging system cannot be applied to the patients directly due to the large image acquisition device. In a previous study, Murakami et al. demonstrated the pH analysis on active and arrested caries using ISFET and a pH imaging-microscope20. The pH data were clearly related to the state of the caries. Based on microfabrication technology, the sensing area of the ISFET was only 0.015 mm long and 0.75 mm wide, but the overall ISFET device, including the support and the cover of the ISFET sensor was large20. Consequently, employing ISFET in a clinical application was not practical. In contrast, the properties of iridium oxide (IrO x) are acknowledged to overcome these issues. IrO x may realize an ideal pH sensor compatible with clinical applications24. In this paper, we propose a pH mapping on tooth surfaces using a micro pH sensor suitable for a caries diagnostic system without risk of X-ray exposure. The Ir/IrOx is mechanically strong and chemically inert. Furthermore, even at high temperatures such as sterilization temperature, properties of IrOx can be maintained. In order to contact sensing area with the tooth surface, we develop the needle-type micro pH sensor of which the tip can be controlled in the order of 100 μm. Making use of these advantages, Ir/IrO x micro pH sensors were prepared and characterized, and applied to measure the pH of extracted dental caries. The results were compared to a dentist’s inspection according to the international standard25. Experimental section Fabrication of Ir/IrOx pH sensor. The Ir wires were oxidized according to the literature24 with a minor change. Briefly, an iridium wire (5-mm long and 300-µm in diameter; 99.9% purity, The Nilaco Corp., Japan) and a gold wire (15-mm long and 0.6-mm in diameter; 99.9% purity, The Nilaco Corp., Japan)
ACS Paragon Plus Environment
Analytical Chemistry 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
were welded with a piezo gas burner. The welded wires were cleaned twice using ultrasonication in acetone followed by ethanol and milli-Q water (ultrapure water type 1, Merck Millipore Corporation, USA). Then the wires were dried in air before oxidization. The cleaned wires were placed on a gold sheet in an alumina crucible. The wires were covered with Li2CO3 powder (anhydrous powder +99.9% purity, Wako Pure Chemical Industries, Japan) and oxidized at 870°C for 5 hours in air with an electric furnace. After cooling to room temperature, the oxidized wires were rinsed with Milli-Q water to dissolve the remaining crystallized Li2CO3 and stirred with 0.2 M HCl. This process was repeated several times, depending on the amount of the leftover Li2CO3. Afterwards, the pH sensors were rinsed with Milli-Q water and treated at 120°C overnight. Finally, the sensors were completely coated with a silicone adhesive (Onecomponent RTV, Shin-Etsu Chemical Co., Ltd) except for the sensing area. The length of the exposed IrOx tip was around 0.5 mm. The gold tip was connected to the data acquisition instrument. Characterization of Ir/IrOx pH sensor. The pH response of the sensor was evaluated in standard pH solutions from pH 3 to pH 9 (pH standard solution set 101-S, Horiba, Japan) against a reference electrode (Ag/AgCl) with agarose gel as a salt bridge. The Ir/IrOx wires were analyzed with SEM (SU8240, Hitachi High-Technologies Corporation, Japan) and energy dispersive X-ray spectrometry (EDX) [EDAX (Octane Super), AMETEK Co., Japan]. Dental caries samples. Dental clinics kindly donated 18 extracted human tooth samples (Supplementary table S-1). All experiments using extracted human teeth were conducted with the approval of the Research Ethics Committee of Tokyo Medical and Dental University (approval number: 725). The caries tooth samples with moderate to severe dentinal caries on the various sites were used. Prior to the experiments, the samples were stored frozen at –30ºC without any solution. The caries samples were classified by a dentist into two groups: active caries and arrested caries. Criteria such as a soft surface and yellow or light brown pigmentation were set for active caries, while a hard surface and dark brown or black pigmentation indicated arrested caries2,4. Some samples contained both active caries and arrested caries from different origins. pH measurements of caries surfaces. The potentiometric responses of the pH sensor were measured with a Keithley 6514 system electrometer and recorded with the LabVIEW data logging application (Figure S1). At the beginning of each experiment, the Ir/IrOx pH sensor was calibrated in standard solutions of pH 9, pH 4, and pH 7, and normal saline (0.9% NaCl, Otsuka Pharmaceutical Factory, Japan) for 10 minutes each. We used newly opened saline solution to minimize the effect of carbon dioxide absorption in every experiment, while the saline solutions were kept in sealed bottles. The caries samples were rinsed with Milli-Q water, dried, and fixed on the impression material (Imprinsis putty, Tokuyama Dental Corp., Japan). The surfaces of the samples were covered with a small amount of saline (
