A pH Sensitive Fluorescence Probe Based on Tricarbocyanine

A pH sensitive near-infrared (NIR) fluorescent probe, IR-DO3A, was synthesized. It was based on combination of tricarbocyanine that had typical NIR absorption and fluorescent emission spectrum, and DO3A (1,4,7,10-tetra azacyclododecane -1,4,7-triyl)triacetic acid) that had three side chain of carboxylic acids. Phenyl sulfonamide that was used to maintain the density of electronic cloud of tricarbocyanine backbone and improve the solubility of the whole molecule was selected to connect tricarbocyanine and DO3A by several substitution reactions under some certain conditions. The final product, IR-DO3A was confirmed by NMR and MS spectrums, and had a better solubility in polar solvent than tricarbocyanine. The optical properties of IR-DO3A in two mixture solvents were studied. It had maximum Uv-vis absorption at 750nm and 805nm, fluorescent emission at 808nm, all these wavelengths were in NIR range and there was a 58nm Stokes-Shift. Also its optical properties at different pH in water were studied and it was sensitive in pH 6~9, which related with the protonation and deprotonation processes of IR-DO3A at different pH. These pH sensitive and NIR-fluorescent properties may result in an application for revealing the pH changes of physiological diseases.     

Short-wave near-infrared spectroscopy analysis of major compounds in milk powder and wavelength assignment

In this study, short-wave near-infrared (NIR) spectroscopy at 800–1050 nm region was investigated for the analysis of main compounds in milk powder. Through quantitative analysis, the feasibility is further demonstrated for the simultaneous measurement of fat, proteins and carbohydrate in milk powder. Two models, partial least-squares and least-squares support vector machine, were compared and utilized for regression coefficients and loading weights. The affect of standard normal variate spectral pretreatment to model performance was evaluated. Based on the resulted coefficients and loading weights, interesting wavelength regions of nutrition in milk powder are screened and the assignment of all specific wavelengths is firstly proposed in the details associated with chemical base. Instead of the whole short-wave NIR spectral data, these assigned wavelengths which can be reliably exploited were used for the content determination. Compared with other spectroscopy technique, assigned short-wave NIR spectral wavelengths did a good work. Determination coefficients for prediction are 0.981, 0.984, and 0.982, respectively for three components. The proposed wavelength assignment in the short-wave NIR region could be used for the component contents determination of milk powder, and could be as a guidance to interpret the spectra of milk powder.     

Near-infrared Fluorescence Spectroscopy Detects Alzheimer's Disease In Vitro

The purpose of this study was to investigate whether near-infrared (NIR) fluorescence spectroscopy could be used to detect Alzheimer's disease (AD) by brain tissue autofluorescence. Unfixed temporal cortex specimens from AD cases and age-matched, non-AD controls were frozen at autopsy and then thawed just prior to spectral measurement. Spectra of intrinsic tissue fluorescence induced by 647 nm light were recorded from 650 to 850 nm. We used principal component analysis of the tissue spectra from 17 AD cases and 5 non-AD control cases in a calibration study to establish a diagnostic algorithm. Retrospectively applied to the calibration set, the algorithm correctly classified 23 of 24 specimens. In a prospective study of 19 specimens from 5 AD brains and 2 non-AD control brains, 3 of the 4 control specimens and all AD specimens were correctly diagnosed. Both the excitation light used and the measured brain tissue autofluorescence are at NIR wavelengths that can propagate through skull and overlying tissue. Therefore, our results demonstrate an optical spectroscopic technique that carries direct molecular level information about disease. This is the first step toward a clinical tool that has the potential to be applied to the noninvasive diagnosis of AD in living patients.     

Screening of humic and fulvic acids in estuarine sediments by near-infrared spectrometry

Diffuse reflectance near-infrared spectroscopy (NIR) combined with partial least squares (PLS) data treatment has been employed for the rapid and nondestructive determination of sedimentary humic substances. Forty one samples of surface estuarine sediments, taken during distinct seasonal periods from different locations across Ria de Arousa (northwest of Spain), were scanned at wavelengths from 833 to 2,976 nm (12,000 to 3,360 cm⁻¹). Twenty four samples were randomly selected, from previous hierarchical cluster analysis of their NIR spectra, for the calibration set, and the 17 remaining samples were assigned to the validation set. NIR spectra of calibration samples were correlated to measured values of humic acids (HAs) and fulvic acids (FAs), which ranged from 1.53 to 28.17 mg/g and from 0.37 to 2.45 mg/g, respectively, using PLS regression and multiplicative scattering correction on the raw and first-derivative NIR spectra, respectively. Low root mean square error of prediction values of 4.3 mg HA/g sediment and 0.25 mg FA/g sediment were obtained. Good residual prediction deviation values of 1.16 and 1.2 were obtained for HA and FA, respectively, allowing the PLS models built to be considered as appropriate tools for screening purposes. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Inside Back Cover: A Near-Infrared Light-Activated Photocage Based on a Ruthenium Complex for Cancer Phototherapy (Angew. Chem. Int. Ed. 24/2023)

Conventional photocages only respond to short wavelength light, which is a significant obstacle to developing efficient phototherapy in vivo. The development of photocages activated by near-infrared (NIR) light at wavelengths from 700 to 950 nm is important for in vivo studies but remains challenging. Herein, we describe the synthesis of a photocage based on a ruthenium (Ru) complex with NIR light-triggered photocleavage reaction. The commercial anticancer drug, tetrahydrocurcumin (THC), was coordinated to the Ru^II center to create the Ru-based photocage that is readily responsive to NIR light at 760 nm. The photocage inherited the anticancer properties of THC. As a proof-of-concept, we further engineered a self-assembled photocage-based nanoparticle system with amphiphilic block copolymers. Upon exposure to NIR light at 760 nm, the Ru complex-based photocages were released from the polymeric nanoparticles and efficiently inhibited tumor proliferation in vivo.     

Determination of total nitrogen content, pH, density, refractive index, and brix in Thai fish sauces and their classification by near-infrared spectroscopy with searching combination moving window partial least squares

Near-infrared (NIR) transflectance spectra in the region of 1100-2500 nm were measured for 100 Thai fish sauces. Quantitative analyses of total nitrogen (TN) content, pH, refractive index, density and brix in the Thai fish sauces and their qualitative analyses were carried out by multivariate analyses with the aid of wavelength interval selection method named searching combination moving window partial least squares (SCMWPLS). The optimized informative region for TN selected by SCMWPLS was the region of 2264-2428 nm. A PLS calibration model, which used this region, yielded the lowest root mean square error of prediction (RMSEP) of 0.100% w/v for the PLS factor of 5. This prediction result is significantly better than those obtained by using the whole spectral region or informative regions selected by moving window partial least squares regression (MWPLSR). As for pH, density, refractive index and brix, the 1698-1722, and 2222-2258 nm regions, the 1358-1438 nm region, the 1774-1846, and 2078-2114 nm regions, and the 1322-1442, and 2000-2076 nm regions were selected by SCMWPLS as the optimized regions. The best prediction results were always obtained by use of the optimized regions selected by SCMWPLS. The lowest RMSEP for pH, density, refractive index and brix were 0.170, 0.007 g cm(-3), 0.0079 and 0.435 degrees Brix, respectively. Qualitative models were developed by using four supervised pattern recognitions, linear discriminant analysis (LDA), factor analysis-linear discriminant analysis (FA-LDA), soft independent modeling of class analog (SIMCA), and K neareat neighbors (KNN) for the optimized combination of informative regions of the NIR spectra of fish sauces to classify fish sauces into three groups based on TN. All the developed models can potentially classify the fish sauces with the correct classification rate of more than 82%, and the KNN classified model has the highest correct classification rate (95%). The present study has demonstrated that NIR spectroscopy combined with SCMWPLS is powerful for both the quantitative and qualitative analyses of Thai fish sauces.     

Semi-quantitative analysis of indigo by surface enhanced resonance Raman spectroscopy (SERRS) using silver colloids

In this paper we report for the first time semi-quantitative analysis of indigo using surface enhanced Raman spectroscopy (SERS) and surface enhance resonance Raman spectroscopy (SERRS). Indigo, a dye widely used today in the textile industry, has been used, historically, both as a dye and as a pigment; the latter in both paintings and in printed material. The molecule is uncharged and largely insoluble in most solvents. The application of SERS/SERRS to the semi-quantitative analysis of indigo has been examined using aggregated citrate-reduced silver colloids with appropriate modifications to experimental protocols to both obtain and maximise SERRS signal intensities. Good linear correlations are observed for the dependence of the intensities of the SERRS band at 1151 cm(-1) using laser exciting wavelengths of 514.5 nm (R=0.9985) and 632.8 nm (R=0.9963) on the indigo concentration over the range 10(-7)-10(-5) and 10(-8)-10(-5) mol dm(-3), respectively. Band intensities were normalised against an internal standard (silver sol band at 243 cm(-1)). Resonance Raman spectra (RRS) of aqueous solutions of indigo could not be collected because of its low solubility and the presence of strong fluorescence. It was, however, possible to obtain RS and RRS spectra of the solid at each laser excitation wavelength. The limits of detection (L.O.D.) of indigo by SERS and SERRS using 514.5 and 632.8 nm were 9 ppm at both exciting wavelengths. Signal enhancement by SERS and SERRS was highly pH dependent due to the formation of singly protonated and possibly doubly protonated forms of the molecule at acidic pH. The SERS and SERRS data provide evidence to suggest that an excess of monolayer coverage of the dye at the surface of silver colloids is observed at concentrations greater than 7.85x10(-6) mol dm(-3) for each exciting wavelength. The data reported herein also strongly suggest the presence of multiple species of the indigo molecule.     

A multimodal spectrometer for Raman scattering and near-infrared absorption measurement

Raman and infrared (IR) spectroscopy are complementary spectroscopic techniques. However, measurement of Raman and IR spectra are commonly carried out on separate instruments. A dispersive system that enables both Raman spectroscopy and NIR spectroscopy was designed, built, and tested. The prototype system measures spectral ranges of 2600–300 cm⁻¹ and 752–987 nm for Raman and NIR channels, respectively. A wavelength accuracy better than 0.6 nm and spectral resolution better than 1 nm (14.4 cm⁻¹ for Raman channel) could be achieved with our configuration. The linearity of spectral response was better than 99.8%. The intensity stability of the instrument was found to be 0.7% and 0.4% for Raman and NIR channels, respectively. The performance of the instrument was evaluated using binary aqueous solutions of ethanol and ovalbumin. It was found that ethanol concentrations (2–10%) could be predicted with a root mean squared error of prediction (RMSEP) of 0.45% using Raman peak height at 882.2 cm⁻¹. Quantification of ovalbumin concentration (8–16 g/L) in aqueous solutions and in denatured states yielded RMSEP values of 1.05 g/L and 0.74 g/L, respectively. Using concentration as external perturbation in two-dimensional correlation spectroscopy (2DCOS), heterospectral correlation analysis revealed the relationship between NIR and Raman spectra.     

Qualitative and quantitative analysis of oxytetracycline by near-infrared spectroscopy

Near-infrared (NIR) spectroscopy, in combination with chemometrics, enable the analysis of raw materials without time-consuming sample preparation methods. The aim of our work was to estimate critical parameters in the analytical specification of oxytetracycline, and consequently the development of a method for quantification and qualification of these parameters by NIR spectroscopy. A Karl Fischer (K.F.) titration to determine the water content, a colorimetric assay method, and Fourier transform-infrared (FT-IR) spectroscopy to identify the oxytetracycline base, were used as reference methods, respectively. Multivariate calibration was performed on NIR spectral data using principal component analysis (PCA), partial least-squares (PLS 1) and principal component regression (PCR) chemometric methods. Multivariate calibration models for NIR spectroscopy have been developed. Using PCA and the Soft Independent Modelling of Class Analogy (SIMCA) approach, we established the cluster model for the determination of sample identity. PLS 1 and PCR regression methods were applied to develop the calibration models for the determination of water content and the assay of the oxytetracycline base. Comparing the PLS and PCR regression methods we found out that the PLS is better established by NIR, especially as the spectroscopic data (NIR spectra) are highly collinear and there are many wavelengths due to non-selective wavelengths. The calibration models for NIR spectroscopy are convenient alternatives to the colorimetric method and to the K.F. method, as well as to FT-IR spectroscopy, in the routine control of incoming material.     

Near-infrared spectroscopy of polymers

Although near–infrared (NIR) spectroscopy has been used over many decades for the – primarily quantitative – analysis of polymers containing OH–, NH– and CH–functionalities (e.g. determination of OH–number, water content and residual carbon–carbon double–bonds), it has never been established as a wide–spread analytical and physical tool comparable to other spectroscopic techniques. In the late seventies, however, two new developments have initiated a renaissance of NIR spectroscopy in analytical chemistry. On the one hand, chemo–metric data evaluation techniques have – in combination with diffuse reflection measurements – opened up the possibility of non–destructive, rational multicomponent analysis and identity control of solid polymers with varying morphologies. On the other hand, the introduction of optical light fibres has contributed to an enormous expansion of conventional NIR spectroscopy in terms of remote control. Thus, specific fibre–optic probes allow a separation of the spectrometer and the location of sample measurement over several hundred meters and tremendously alleviate the analysis of toxic and hazardous materials including process and reaction control. Recently, further progress is made by the development of new, rapid-scan NIR monochromator systems without mechanically moved parts, such as acousto–optic tunable filters. Last, but not least, it should be mentioned, that – although not treated here – the new approach of Fourier–Transform Raman spectroscopy with Nd–YAG laser excitation at 1064 nm is principally also a near–infrared technique.     

Application of fluorescence correlation spectroscopy to the measurement of local temperature in solutions under optical trapping condition

Fluorescence correlation spectroscopy (FCS) was applied to the quantitative evaluation of the local heating in small domains <1 microm in solutions under the laser trapping condition in the presence of a near-infrared (NIR) laser beam at 1064 nm. On the basis of the translational diffusion coefficient of fluorescent molecules obtained by FCS, the relationship between temperature rise and the incident NIR laser power, DeltaT/DeltaP, were determined to be 62 +/- 6, 49 +/- 7, and 23 +/- 1 deg K/W in ethylene glycol, ethanol, and water, respectively, while no remarkable temperature increase was observed for deuterated water. The value of DeltaT/DeltaP linearly increased as a function of alpha/lambda (alpha is the extinction coefficient of solvent at the wavelength and lambda is the thermal conductivity of the medium). The validity and the applicability of the present method for the measurement of the local temperature increase were discussed by comparing the present results with previous ones by other various methods.     

Study on the resonance light-scattering spectrum of anionic dye xylenol orange-cetyltrimethylammonium-nucleic acids system and determination of nucleic acids at nanogram levels

The interaction of xylenol orange (XO) and nucleic acids in the presence of cetyltrimethylammonium bromide (CTMAB) in aqueous solution has been studied by a resonance light-scattering (RLS) technique with a common spectrofluorometer. In hexamethylenetetramine (HMTA) buffer (pH7.30), XO and nucleic acids react with cetyltrimethylammonium bromide to form large particles of three-component complex, which results in strong enhanced RLS signals characterized by three peaks at 295.9, 335.5 and 542 nm, Mechanistic studies showed that the enhanced RLS stems from the aggregation of XO on DNA through the bridged and synergistic effect of CTMAB. With the enhanced RLS signals at the three wavelengths, the enhanced RLS intensity is proportional to the concentration of nucleic acids in an appropriate range. The lowest limit of determination was 5.31 ng ml(-1), three synthetic samples of yDNA were analyzed satisfactorily.     

An enrichment device of silica-based monolithic material and its application to determine micro-carbaryl by NIRS

Silica-based monolithic column material was synthesized and an enrichment device was fabricated with the material by assembling the material inside a glass column.The enrichment device was applied for the determination of micro-carbaryl with near-infrared spectroscopy(NIRS).The aqueous solutions of carbaryl passed through the device and the carbaryl was enriched on the surface of the material where diffuse reflection NIR spectra were measured.These procedures of enrichment and measurement ensured to conc...     

Development of FRET-based dual-excitation ratiometric fluorescent pH probes and their photocaged derivatives

Dual-excitation ratiometric fluorescent probes allow the measurement of fluorescence intensities at two excitation wavelengths, which should provide a built-in correction for environmental effects. However, most of the small-molecule dual-excitation ratiometric probes that have been reported thus far have shown rather limited separation between the excitation wavelengths (20-70 nm) and/or a very small molar absorption coefficient at one of the excitation wavelengths. These shortcomings can lead to cross-excitation and thus to errors in the measurement of fluorescence intensities and ratios. Herein, we report a FRET-based molecular strategy for the construction of small-molecule dual-excitation ratiometric probes in which the donor and acceptor excitation bands exhibit large separations between the excitation wavelengths and comparable excitation intensities, which is highly desirable for determining the fluorescence intensities and signal ratios with high accuracy. Based on this strategy, we created a coumarin-rhodamine FRET platform that was then employed to develop the first class of FRET-based dual-excitation ratiometric pH probes that have two well-resolved excitation bands (excitation separations>160 nm) and comparable excitation intensities. In addition, these pH probes may be considered as in a kind of "secured ratioing mode". As a further application of these pH probes, the dual-excitation ratiometric pH probes were transformed into the first examples of photocaged dual-excitation ratiometric pH probes to improve the spatiotemporal resolution. It is expected that the modular nature of our FRET-based molecular strategy should render it applicable to other small-molecule dual-dye energy-transfer systems based on diverse fluorescent dyes for the development of a wide range of dual-excitation ratiometric probes with outstanding spectral features, including large separations between the excitation wavelengths and comparable excitation intensities.     

In vivo application of intestinal pH measurement using 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF) fluorescence imaging

Measurement of gastrointestinal intramucosal pH (pHim) has been recognized as an important factor in the detection of hypoxia-induced dysfunctions. However, current pH measurement techniques are limited in terms of time and spatial resolutions. A major advance in accurate pH measurement was the development of the ratiometric fluorescent indicator dye, 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF). This study aimed to set up and validate a fluorescence imaging technique to measure in vivo the intramucosal pH (pHim) of the intestine. The intestine was inserted into an optical chamber placed under a microscope. Animals were injected intravenously with the pH-sensitive fluorescent dye BCECF. Fluorescence was visualized by illuminating the intestine alternately at 490 and 470 nm. The emitted fluorescence was directed to an intensified camera. The ratio of emitted fluorescence at excitation wavelengths of 490 and 470 nm was measured, corrected and converted to pHim by constructing a calibration curve. The pHim controls were performed with a pH microelectrode and were correlated with venous blood gas sampling. Results show that pHim is determined with an accuracy of +/- 0.07 pH units and a response time of 1 min. In conclusion pHim mapping of rat intestine can be obtained by fluorescence imaging using BCECF. This technology could be easily adapted for endoscopic pH measurements.     

Spectroscopic studies of solutes in aqueous solution

Absorption and fluorescence characteristics of aqueous solutions of salts, sugars, and amino acids were studied using UV-vis spectroscopy and spectrofluorometry. Motivation stemmed from unanticipated absorption spectral and fluorescence features of the "exclusion zone" seen adjacent to various hydrophilic surfaces. Those features implied a structure distinct from that of bulk water (Adv. Colloid Interface Sci. 2006, 127, 19). Absorption peaks at approximately 270 nm similar to those observed in the exclusion zone were seen in solutions of the following substances: salts, Nafion 117 solution/film, l-lysine, d-alanine, d-glucose and sucrose. To determine the fate of the absorbed energy, we studied the fluorescence properties of these solutions. The salts showed fluorescence emission around 480-490 nm under different excitation wavelengths. The fluorescence intensity of LiCl was higher than NaCl, which was in turn higher than KCl-the same ordering as the absorption intensities. Fluorescence of Nafion 117 solution/film, l-lysine, d-alanine, d-glucose and sucrose were observed as well, with multiple excitation wavelengths. Hence, at least some of the absorbed energy is released as fluorescence. The results show features closely similar to those observed in the exclusion zone, implying that the aqueous region around the solutes resembles the aqueous zone adjacent to hydrophilic surfaces. Both may be more extensively ordered than previously thought.     

Controlled interparticle spacing for surface-modified gold nanoparticle aggregates

Aggregation of gold nanoparticles of increasing size has been studied as a consequence of adsorption of 2-aminothiophenol (ATP) on gold nanoparticle surfaces. The capping property of ATP in the acidic pH range has been accounted from UV-vis absorption spectroscopy and surface-enhanced Raman scattering (SERS) studies. The effect of nanoparticle size (8-55 nm) on the nature of aggregation as well as the variation in the optical response due to variable degree of interparticle coupling effects among the gold particles have been critically examined. Various techniques such as transmission electron microscopy, X-ray diffraction, zeta-potential, and average particle size measurement were undertaken to characterize the nanoparticle aggregates. The aggregate size, interparticle distances, and absorption band wavelengths were found to be highly dependent on the pH of the medium and the concentration of the capping agent, ATP. The acquired SERS spectra of ATP relate the interparticle spacing. It has been observed that the SERS signal intensities are different for different sized gold nanoparticles.     

Optimization of a new mobile phase to know the complex and real polyphenolic composition: towards a total phenolic index using high-performance liquid chromatography

An HPLC method is reported for the separation and quantification of five major polyphenolic groups found in fruits and related products: single ring phenolic acids (hydroxybenzoic acid and hydroxycinnamic acid derivatives), flavan-3-ols, flavonols, anthocyanins, and dihydrochalcones. A binary mobile phase consisting of 6% acetic acid in 2 mM sodium acetate aqueous solution (v/v, final pH 2.55) (solvent A) and acetonitrile (solvent B) was used. The use of sodium acetate was new and key to the near baseline separation of 25 phenolics commonly found in fruits. A photodiode array detector was used and data were collected at four wavelengths (280, 320, 360, and 520 nm). This method was sensitive and gave good separation of polyphenolics in apple, cherry, strawberry, blackberry, grape, apple juice, and a processing by-product. The improved separation has led to better understanding of the polyphenolic profiles of these fruits. Individual as well as total phenolic content was obtained, and the latter was close to and correlated well with that obtained by the Folin-Ciocalteu method (FC). The HPLC data can be used as a total phenolic index (TPI) for quantification of fruit phenolics, which is advantageous over the FC because it has more information on individual compounds.     

Effect of temperature variation on the visible and near infrared spectra of wine and the consequences on the partial least square calibrations developed to measure chemical composition

Many studies have reported the use of near infrared (NIR) spectroscopy to characterize wines or to predict wine chemical composition. However, little is known about the effect of variation in temperature on the NIR spectrum of wine and the subsequent effect on the performance of calibrations used to measure chemical composition. Several parameters influence the spectra of organic molecules in the NIR region, with temperature being one of the most important factors affecting the vibration intensity and frequency of molecular bonds. Wine is a complex mixture of chemical components (e.g. water, sugars, organic acids, and ethanol), and a simple ethanol and water model solution cannot be used to study the possible effects of temperature variations in the NIR spectrum of wine. Ten red and 10 white wines were scanned in triplicate at six different temperatures (25 °C, 30 °C, 35 °C, 40 °C, 45 °C and 50 °C) in the visible (vis) and NIR regions (400-2500 nm) in a monochromator instrument in transmission mode (1 mm path length). Principal component analysis (PCA) and partial least squares (PLS) regression models were developed using full cross validation (leave-one-out). These models were used to interpret the spectra and to develop calibrations for alcohol, sugars (glucose + fructose) and pH at different temperatures. The results showed that differences in the spectra around 970 nm and 1400 nm, related to OH bonding were observed for both varieties. Additionally an effect of temperature on the vis region of red wine spectra was observed. The standard error of cross validation (SECV) achieved for the PLS calibration models tended to inverse as the temperature increased. The practical implication of this study it is recommended that the temperature of scanning for wine analysis using a 1 mm path length cuvette should be between 30 °C and 35 °C.