A Smart Wristband Integrated with an IoT-Based Alarming System for Real-Time Sweat Alcohol Monitoring.

Breathalyzer is a common approach to measuring blood alcohol concentration (BAC) levels of individuals suspected of drunk driving. Nevertheless, this device is relatively high-cost, inconvenient for people with limited breathing capacity, and risky for COVID-19 exposure. Here, we designed and developed a smart wristband integrating a real-time noninvasive sweat alcohol metal oxide (MOX) gas sensor with a Drunk Mate, an Internet of Thing (IoT)-based alarming system. A MOX sensor acquired transdermal alcohol concentration (TAC) which was converted to BAC and sent via the IoT network to the Blynk application platform on a smartphone, triggering alarming messages on LINE Notify. A user would receive an immediate alarming message when his BAC level reached an illegal alcohol concentration limit (BAC 50 mg%; TAC 0.70 mg/mL). The sensor readings showed a high linear correlation with TAC (R2 = 0.9815; limit of detection = 0.045 mg/mL) in the range of 0.10−1.05 mg/mL alcohol concentration in artificial sweat, achieving an accuracy of 94.66%. The sensor readings of ethanol in water were not statistically significantly different (p > 0.05) from the measurements in artificial sweat and other sweat-related solutions, suggesting that the device responded specifically to ethanol and was not affected by other electrolytes in the artificial sweat. Moreover, the device could continuously monitor TAC levels simulated in real-time in an artificial sweat testing system. With the integration of an IoT-based alarming system, the smart wristband developed from a commercial gas sensor presented here offers a promising low-cost MOX gas sensor monitoring technology for noninvasive and real-time sweat alcohol measurement and monitoring.

Rapid label-free detection of cholangiocarcinoma from human serum using Raman spectroscopy.

Cholangiocarcinoma (CCA) is highly prevalent in the northeastern region of Thailand. Current diagnostic methods for CCA are often expensive, time-consuming, and require medical professionals. Thus, there is a need for a simple and low-cost CCA screening method. This work developed a rapid label-free technique by Raman spectroscopy combined with the multivariate statistical methods of principal component analysis and linear discriminant analysis (PCA-LDA), aiming to analyze and classify between CCA (n = 30) and healthy (n = 30) serum specimens. The model's classification performance was validated using k-fold cross validation (k = 5). Serum levels of cholesterol (548, 700 cm-1), tryptophan (878 cm-1), and amide III (1248,1265 cm-1) were found to be statistically significantly higher in the CCA patients, whereas serum beta-carotene (1158, 1524 cm-1) levels were significantly lower. The peak heights of these identified Raman marker bands were input into an LDA model, achieving a cross-validated diagnostic sensitivity and specificity of 71.33% and 90.00% in distinguishing the CCA from healthy specimens. The PCA-LDA technique provided a higher cross-validated sensitivity and specificity of 86.67% and 96.67%. To conclude, this work demonstrated the feasibility of using Raman spectroscopy combined with PCA-LDA as a helpful tool for cholangiocarcinoma serum-based screening.

Rapid label-free analysis of Opisthorchis viverrini eggs in fecal specimens using confocal Raman spectroscopy.

Opisthorchis viverrini, a human liver fluke, is highly prevalent in Southeast Asia. Definitive diagnosis of infection is usually achieved parasitologically through the discovery of fluke eggs in feces. However, the eggs of O. viverrini are difficult to differentiate morphologically from those of other minute intestinal flukes in fecal samples, even for experienced technicians. The present study developed a label-free optical methodology for analysis of O. viverrini eggs using Raman spectroscopy. Raman features of O. viverrini eggs were reported that can be used as marker bands for the efficient analysis of O. viverrini eggs in fecal specimens. The methodology presented here allows for the rapid detection of O. viverrini egg infection and can be readily and practicably applied in any clinical setting, even those in which a trained parasitologist is not available.

Full depth measurement of tenofovir transport in rectal mucosa using confocal Raman spectroscopy and optical coherence tomography.

The prophylactic activity of antiretroviral drugs applied as microbicides against sexually transmitted HIV is dependent upon their concentrations in infectable host cells. Within mucosal sites of infection (e.g., vaginal and rectal mucosa), those cells exist primarily in the stromal layer of the tissue. Traditional pharmacokinetic studies of these drugs have been challenged by poor temporal and spatial specificity. Newer techniques to measure drug concentrations, involving Raman spectroscopy, have been limited by laser penetration depth into tissue. Utilizing confocal Raman spectroscopy (RS) in conjunction with optical coherence tomography (OCT), a new lateral imaging assay enabled concentration distributions to be imaged with spatial and temporal specificity throughout the full depth of a tissue specimen. The new methodology was applied in rectal tissue using a clinical rectal gel formulation of 1% tenofovir (TFV). Confocal RS revealed diffusion-like behavior of TFV through the tissue specimen, with significant partitioning of the drug at the interface between the stromal and adipose tissue layers. This has implications for drug delivery to infectable tissue sites. The new assay can be applied to rigorously analyze microbicide transport and delineate fundamental transport parameters of the drugs (released from a variety of delivery vehicles) throughout the mucosa, thus informing microbicide product design.

Label-Free Measurements of Tenofovir Diffusion Coefficients in a Microbicide Gel Using Raman Spectroscopy.

Confocal Raman spectroscopy was implemented in a new label-free technique to quantify molecular diffusion coefficients within gels. A leading anti-HIV drug, tenofovir, was analyzed in a clinical microbicide gel. The gel was tested undiluted, and in 10%-50% wt/wt dilutions with vaginal fluid simulant to capture the range of conditions likely occurring in vivo. The concentration distributions of tenofovir in gel over time and space were measured and input to a mathematical diffusion model to deduce diffusion coefficients. These were 3.16 ± 0.11 × 10-6 cm2/s in undiluted gel, and increased by 11%-46% depending on the extent of dilution. Results were interpreted with respect to traditional release rate measurements in devices such as Franz cells. This comparison highlighted an advantage of our assay in that it characterizes the diffusive barrier within the gel material itself; in contrast, release rate in the traditional assay is affected by external conditions, such as drug partitioning at the gel/liquid sink interface. This new assay is relevant to diffusion in polymeric hydrogels over pharmacologically relevant length scales, for example, those characteristic of topical drug delivery. Resulting transport parameters are salient measures of drug delivery potential, and serve as inputs to computational models of drug delivery performance.

Label-free analysis of tenofovir delivery to vaginal tissue using co-registered confocal Raman spectroscopy and optical coherence tomography.

Vaginally applied microbicide products offer a female-controlled strategy for preventing sexual transmission of HIV. Microbicide transport processes are central to their functioning, and there is a clear need for a better understanding of them. To contribute to that end, we developed an assay to analyze mass transport rates of microbicide molecules within the epithelial and stromal layers of polarized vaginal mucosal tissue during contact with a gel vehicle. The assay utilizes a new diffusion chamber mounted in a custom instrument that combines confocal Raman spectroscopy and optical coherence tomography. This measures depth-resolved microbicide concentration distributions within epithelium and stroma. Data for a tenofovir gel were fitted with a compartmental diffusion model to obtain fundamental transport properties: the molecular diffusion and partition coefficients in different compartments. Diffusion coefficients in epithelium and stroma were computed to be 6.10 ± 2.12 x 10-8 and 4.52 ± 1.86 x 10-7 cm2/sec, respectively. The partition coefficients between epithelium and gel and between stroma and epithelium were found to be 0.53 ± 0.15 and 1.17 ± 0.16, respectively. These drug transport parameters are salient in governing the drug delivery performance of different drug and gel vehicle systems. They can be used to contrast drugs and vehicles during product design, development and screening. They are critical inputs to deterministic transport models that predict the gels' pharmacokinetic performance, which can guide improved design of products and optimization of their dosing regimens.

Co-localized confocal Raman spectroscopy and optical coherence tomography (CRS-OCT) for depth-resolved analyte detection in tissue.

We report the development of a combined confocal Raman spectroscopy (CRS) and optical coherence tomography (OCT) instrument (CRS-OCT) capable of measuring analytes in targeted biological tissues with sub-100-micron spatial resolution. The OCT subsystem was used to measure depth-resolved tissue morphology and guide the acquisition of chemically-specific Raman spectra. To demonstrate its utility, the instrument was used to accurately measure depth-resolved, physiologically-relevant concentrations of Tenofovir, a microbicide drug used to prevent the sexual transmission of HIV, in ex vivo tissue samples.

Quantitative analysis of microbicide concentrations in fluids, gels and tissues using confocal Raman spectroscopy.

Topical vaginal anti-HIV microbicides are an important focus in female-based strategies to prevent the sexual transmission of HIV. Understanding microbicide pharmacokinetics is essential to development, characterization and implementation of efficacious microbicide drug delivery formulations. Current methods to measure drug concentrations in tissue (e.g., LC-MS/MS, liquid chromatography coupled with tandem mass spectrometry) are highly sensitive, but destructive and complex. This project explored the use of confocal Raman spectroscopy to detect microbicide drugs and to measure their local concentrations in fluids, drug delivery gels, and tissues. We evaluated three candidate microbicide drugs: tenofovir, Dapivirine and IQP-0528. Measurements were performed in freshly excised porcine buccal tissue specimens, gel vehicles and fluids using two Horiba Raman microscopes, one of which is confocal. Characteristic spectral peak calibrations for each drug were obtained using serial dilutions in the three matrices. These specific Raman bands demonstrated strong linear concentration dependences in the matrices and were characterized with respect to their unique vibrational signatures. At least one specific Raman feature was identified for each drug as a marker band for detection in tissue. Sensitivity of detection was evaluated in the three matrices. A specific peak was also identified for tenofovir diphosphate, the anti-HIV bioactive product of tenofovir after phosphorylation in host cells. Z-scans of drug concentrations vs. depth in excised tissue specimens, incubated under layers of tenofovir solution in a Transwell assay, showed decreasing concentration with depth from the surface into the tissue. Time-dependent concentration profiles were obtained from tissue samples incubated in the Transwell assay, for times ranging 30 minutes - 6 hours. Calibrations and measurements from tissue permeation studies for tenofovir showed good correlation with gold standard LC-MS/MS data. These results demonstrate that confocal Raman spectroscopy holds promise as a tool for practical, minimally invasive, label-free measurement of microbicide drug concentrations in fluids, gels and tissues.