Introduction to Infrared and Raman-Based Biomedical Molecular Imaging and Comparison with Other Modalities

Molecular imaging has rapidly developed to answer the need of image contrast in medical diagnostic imaging to go beyond morphological information to include functional differences in imaged tissues at the cellular and molecular levels. Vibrational (infrared (IR) and Raman) imaging has rapidly emerged among the molecular imaging modalities available, due to its label-free combination of high spatial resolution with chemical specificity. This article presents the physical basis of vibrational spectroscopy and imaging, followed by illustration of their preclinical in vitro applications in body fluids and cells, ex vivo tissues and in vivo small animals and ending with a brief discussion of their clinical translation. After comparing the advantages and disadvantages of IR/Raman imaging with the other main modalities, such as magnetic resonance imaging (MRI), computed tomography (CT), positron emission tomography/single-photon emission-computed tomography (PET/SPECT), ultrasound (US) and photoacoustic imaging (PAI), the design of multimodal probes combining vibrational imaging with other modalities is discussed, illustrated by some preclinical proof-of-concept examples.     

Quantitative aspects of near-infrared Fourier transform Raman spectroscopy

Three fundamental behaviors of vibrational spectroscopy data manipulation routinely associated with Fourier transform infrared (FTIR) spectroscopy are evaluated for near-infrared (NIR) Fourier transform Raman spectroscopy. Spectral reproducibility, spectral subtraction and sensitivity are examined relative to the NIR FT-Raman experiment. Quantitative predictive ability is compared for identical sets of samples containing mixtures of the three xylene isomers. Partial least-squares analysis is used to compare predictive ability. IR performance is found to be better than Raman, though the potential for method development using NIR FT-Raman is shown to be quite promising.     

Raman mapping and FTIR imaging of lung tissue: congenital cystic adenomatoid malformation

Congenital cystic adenomatoid malformations (CCAMs) are benign masses of non-functional lung tissue developing from an overgrowth of the terminal bronchioles with subsequent suppressing of alveolar growth. For the first time CCAMs are studied by Raman mapping and Fourier transform infrared (FTIR) imaging. Both vibrational spectroscopic methods are able to analyze the biochemical composition of tissues and their pathological changes at the molecular level. Cryosections were prepared on calcium fluoride substrates from CCAMs and from normal lung tissue of two infant patients who underwent surgery. Raman maps were collected at a step size of 100 microm in order to assess the whole tissue section and at a smaller step size of 10 microm in order to resolve details in selected areas. FTIR images were collected in the macroscopic and microscopic modes. Data sets were segmented by cluster analysis and the mean spectra of each cluster were compared. At low lateral resolution a lower red blood cell content and higher lipid content were found in CCAMs than in normal lung tissue. At higher lateral resolution, accumulations of lipids and glycogen were identified in CCAMs. The lipid aggregates contain a high concentration of phosphatidylcholine. It is discussed how the combined application of Raman mapping and FTIR imaging might improve the differential diagnosis of lung malformations and how both modalities might be applied to other bioanalytical and biomedical problems in the future.     

The destruction-free analysis of polymers by fourier transform infrared photoacoustic and fourier transform Raman spectroscopy: A comparison

With the introduction of rapid–scanning Fourier transform infrared (FTIR) and recently Raman (FT–Raman) spectroscopy, vibrational spectroscopy has been launched into a new era of applications in polymer chemistry and physics. Thus, the increase in sensitivity provided by multiple scanning has led to the breakthrough of new, destruction–free sampling techniques, such as photoacoustic and Raman spectroscopy. This paper provides a comparison between data produced by FTIR photoacoustic and FT–Raman analysis of a range of polymers, and structural information available from both techniques is discussed.     

Vibrational spectroscopy differentiates between multipotent and pluripotent stem cells

Over the last few years, there has been an increased interest in the study of stem cells in biomedicine for therapeutic use and as a source for healing diseased or injured organs/tissues. More recently, vibrational spectroscopy has been applied to study stem cell differentiation. In this study, we have used both synchrotron based FTIR and Raman microspectroscopies to assess possible differences between human pluripotent (embryonic) and multipotent (adult mesenchymal) stem cells, and how O(2) concentration in cell culture could affect the spectral signatures of these cells. Our work shows that infrared spectroscopy of embryonic (pluripotent) and adult mesenchymal (multipotent) stem cells have different spectral signatures based on the amount of lipids in their cytoplasm (confirmed with cytological staining). Furthermore, O(2) concentration in cell culture causes changes in both the FTIR and Raman spectra of embryonic stem cells. These results show that embryonic stem cells might be more sensitive to O(2) concentration when compared to mesenchymal stem cells. While vibrational spectroscopy could therefore be of potential use in identifying different populations of stem cells further work is required to better understand these differences.     

氧化石墨烯水热还原前后的发光光谱

分别采用改进Hummers方法和水热还原法制备了氧化石墨烯(GO)和还原氧化石墨烯(RGO)。 GO和RGO经透射电子显微镜(TEM)、紫外-可见吸收光谱(UV-Vis)、红外光谱(IR)、荧光发射和激发光谱(PL、PLE)等技术手段进行了表征。 荧光发射光谱显示,氧化石墨烯(GO)在可见光的激发下可以得到波长在600~800 nm范围内的宽谱近红外荧光。 通过比较氧化石墨烯水热还原前后的光谱变化,发现氧化石墨烯近红外荧光起源于氧化石墨烯的表面含氧基团,如C=O、COOH。 近红外荧光穿透性好、对生物组织损坏小,非常适合于生物成像,预示着氧化石墨烯在生物成像方面的应用潜力。     

Monitoring of the Sol-Gel Synthesis of Organic-inorganic Hybrids by FTIR Transmission,FTIR/ATR,NIR and Raman Spectroscopy

Summary: The sol-gel synthesis of organic-inorganic hybrids based on triethoxysilane- terminated poly(ethylene oxide) and tetraethylorthosilicate was monitored in-situ using three spectroscopic methods (FTIR/ATR, Raman, NIR). These spectroscopic methods allow in-situ monitoring of the evolution of hybrid materials starting from the modification of the polymer and the early steps of hydrolysis up to the network formation. By application of ²⁹Si solid-state NMR spectroscopy the assignment and quantification of the Raman bands to different end groups and different cross-linking states was made. The sol-gel reaction was also followed by in-line NIR spectroscopy. A multivariate data analysis was accomplished to obtain a conversion-time curve. Furthermore, we investigated spin-coated films on wafers using FTIR transmission spectroscopy.     

FT-Raman,FT-infrared and NIR spectroscopic characterization of oxygen-delignified kraft pulp treated with hydrogen peroxide under acidic and alkaline conditions

The effects of bleaching treatment of oxygen-delignified softwood kraft pulp with hydrogen peroxide under acidic and alkaline conditions were studied using standard technological techniques and spectroscopic analytical methods: near-infrared (NIR), Fourier-transform infrared (FTIR) and Fourier-transform (FT) Raman spectroscopies. Among the three tested spectroscopic techniques, NIR analysis appeared to be the most appropriate in terms of possible technological applications. The use of NIR spectroscopy combined with multivariate data analysis allowed to create models for pulp bleaching monitoring based on CIE L*a*b* measurements. Near-infrared and FTIR spectroscopic studies allowed differentiating between the effects of the acidic and alkaline peroxide bleaching stages, but failed in relation to the delignification process. The most representative bands in the FTIR and FT-Raman spectra in terms of delignification and chromophore removal exhibited no correlation with standard technological measurement results.     

Near infrared spectroscopic imaging assessment of cartilage composition: Validation with mid infrared imaging spectroscopy

Disease or injury to articular cartilage results in loss of extracellular matrix components which can lead to the development of osteoarthritis (OA). To better understand the process of disease development, there is a need for evaluation of changes in cartilage composition without the requirement of extensive sample preparation. Near infrared (NIR) spectroscopy is a chemical investigative technique based on molecular vibrations that is increasingly used as an assessment tool for studying cartilage composition. However, the assignment of specific molecular vibrations to absorbance bands in the NIR spectrum of cartilage, which arise from overtones and combinations of primary absorbances in the mid infrared (MIR) spectral region, has been challenging. In contrast, MIR spectroscopic assessment of cartilage is well-established, with many studies validating the assignment of specific bands present in MIR spectra to specific molecular vibrations. In the current study, NIR imaging spectroscopic data were obtained for compositional analysis of tissues that served as an in vitro model of OA. MIR spectroscopic data obtained from the identical tissue regions were used as the gold-standard for collagen and proteoglycan (PG) content. MIR spectroscopy in transmittance mode typically requires a much shorter pathlength through the sample (≤10 microns thick) compared to NIR spectroscopy (millimeters). Thus, this study first addressed the linearity of small absorbance bands in the MIR region with increasing tissue thickness, suitable for obtaining a signal in both the MIR and NIR regions. It was found that the linearity of specific, small MIR absorbance bands attributable to the collagen and PG components of cartilage (at 1336 and 856 cm⁻¹, respectively) are maintained through a thickness of 60 μm, which was also suitable for NIR data collection. MIR and NIR spectral data were then collected from 60 μm thick samples of cartilage degraded with chondroitinase ABC as a model of OA. Partial least squares (PLS) regression using NIR spectra as input predicted the MIR-determined compositional parameters of PG/collagen within 6% of actual values. These results indicate that NIR spectral data can be used to assess molecular changes that occur with cartilage degradation, and further, the data provide a foundation for future clinical studies where NIR fiber optic probes can be used to assess the progression of cartilage degradation.     

Characterization and analysis of mycobacteria and Gram-negative bacteria and co-culture mixtures by Raman microspectroscopy, FTIR, and atomic force microscopy

The molecular composition of mycobacteria and Gram-negative bacteria cell walls is structurally different. In this work, Raman microspectroscopy was applied to discriminate mycobacteria and Gram-negative bacteria by assessing specific characteristic spectral features. Analysis of Raman spectra indicated that mycobacteria and Gram-negative bacteria exhibit different spectral patterns under our experimental conditions due to their different biochemical components. Fourier transform infrared (FTIR) spectroscopy, as a supplementary vibrational spectroscopy, was also applied to analyze the biochemical composition of the representative bacterial strains. As for co-cultured bacterial mixtures, the distribution of individual cell types was obtained by quantitative analysis of Raman and FTIR spectral images and the spectral contribution from each cell type was distinguished by direct classical least squares analysis. Coupled atomic force microscopy (AFM) and Raman microspectroscopy realized simultaneous measurements of topography and spectral images for the same sampled surface. This work demonstrated the feasibility of utilizing a combined Raman microspectroscopy, FTIR, and AFM techniques to effectively characterize spectroscopic fingerprints from bacterial Gram types and mixtures.

Biodiesel classification by base stock type (vegetable oil) using near infrared spectroscopy data

The use of biofuels, such as bioethanol or biodiesel, has rapidly increased in the last few years. Near infrared (near-IR, NIR, or NIRS) spectroscopy (>4000 cm⁻¹) has previously been reported as a cheap and fast alternative for biodiesel quality control when compared with infrared, Raman, or nuclear magnetic resonance (NMR) methods; in addition, NIR can easily be done in real time (on-line). In this proof-of-principle paper, we attempt to find a correlation between the near infrared spectrum of a biodiesel sample and its base stock. This correlation is used to classify fuel samples into 10 groups according to their origin (vegetable oil): sunflower, coconut, palm, soy/soya, cottonseed, castor, Jatropha, etc. Principal component analysis (PCA) is used for outlier detection and dimensionality reduction of the NIR spectral data. Four different multivariate data analysis techniques are used to solve the classification problem, including regularized discriminant analysis (RDA), partial least squares method/projection on latent structures (PLS-DA), K-nearest neighbors (KNN) technique, and support vector machines (SVMs). Classifying biodiesel by feedstock (base stock) type can be successfully solved with modern machine learning techniques and NIR spectroscopy data. KNN and SVM methods were found to be highly effective for biodiesel classification by feedstock oil type. A classification error (E) of less than 5% can be reached using an SVM-based approach. If computational time is an important consideration, the KNN technique (E = 6.2%) can be recommended for practical (industrial) implementation. Comparison with gasoline and motor oil data shows the relative simplicity of this methodology for biodiesel classification.     

Intra-operative optical diagnostics with vibrational spectroscopy

Established methods for characterization of tissue and diagnostics, for example histochemistry, magnetic resonance imaging (MRI), X-ray tomography, or positron emission tomography (PET), are mostly not suitable for intra-operative use. However, there is a clear need for an intra-operative diagnostics especially to identify the borderline between normal and tumor tissue. Currently, vibrational spectroscopy techniques (both Raman and infrared) complement the standard methods for tissue diagnostics. Vibrational spectroscopy has the potential for intra-operative use, because it can provide a biochemically based profile of tissue in real time and without requiring additional contrast agents, which may perturb the tissue under investigation. In addition, no electric potential needs to be applied, and the measurements are not affected by electromagnetic fields. Currently, promising approaches include Raman fiber techniques and nonlinear Raman spectroscopy. Infrared spectroscopy is also being used to examine freshly resected tissue ex vivo in the operating theater. The immense volume of information contained in Raman and infrared spectra requires multivariate analysis to extract relevant information to distinguish different types of tissue. The promise and limitations of vibrational spectroscopy methods as intra-operative tools are surveyed in this review.     

Quantitative analysis of diphenhydramine hydrochloride in pharmaceutical wafers using near infrared and Raman spectroscopy

Wafers with varying concentrations of diphenhydramine hydrochloride (DPH-HCl) as active pharmaceutical ingredient (API) were prepared and their near infrared (NIR) and Raman spectra recorded. The purpose of this study was to compare the suitability of these two vibrational spectroscopic techniques for the quantification of DPH-HCl in pharmaceutical wafers. Partial least squares (PLS1) calibration models with different data pretreatments were tested. Both NIR and Raman spectroscopy proved to be suitable to predict DPH-HCl contents at lower concentration ranges. At higher concentrations, interference by crystallization processes was observed. For investigating the general applicability of the quantification methods, two commercially available products were examined.     

Fourier transform raman spectroscopy using a bench-top fourier transform infrared spectrometer

The use of a bench top FTIR spectrometer for near infrared Fourier transform Raman spectroscopy is demonstrated. The use of near infrared excitation results in fluorescence free Raman spectra allowing previously difficult samples to be measured.     

具有聚集诱导发光性质的近红外荧光染料

聚集诱导发光(AIE)现象的发现为解决传统有机荧光分子在高浓度和聚集形态下存在的荧光猝灭问题提供了最佳方案,并实现了在光电器件、化学传感、生物成像和靶向治疗等众多领域的广泛应用.随着对AIE 发光机理研究的不断深入,AIE 分子体系得到了极大的扩展.其中,一类具有给体-受体结构的AIE分子能够显著降低分子能隙,使发光分...     

The use of near infrared (NIR) spectroscopy to study specific interactions in polymer blends

The purpose of this brief communication is to demonstrate how near infrared (NIR) spectroscopy can be used to study specific interactions is miscible polymer blends. We will first demonstrate, using model systems and blends of polycaprolactone (PCL) and phenoxy [poly(hydroxypropylether of bisphenol-A)], that information obtained from the OH stretch overtone absorption in the NIR is consistent with the information obtained from the OH stretch fundamental in the mid-IR region. We will then illustrate the advantages of NIR analysis of specific interaction with spectra obtained, over a range of temperatures, from thick (2 mm) films of blends of Nylon-6 and the zinc salt of sulfonated polystyrene (ZnSPS). © 1994 John Wiley & Sons, Inc.     

Fluorescence Imaging In Vivo at Wavelengths beyond 1500 nm

Compared to imaging in the visible and near‐infrared regions below 900 nm, imaging in the second near‐infrared window (NIR‐II, 1000–1700 nm) is a promising method for deep‐tissue high‐resolution optical imaging in vivo mainly owing to the reduced scattering of photons traversing through biological tissues. Herein, semiconducting single‐walled carbon nanotubes with large diameters were used for in vivo fluorescence imaging in the long‐wavelength NIR region (1500–1700 nm, NIR‐IIb). With this imaging agent, 3–4 μm wide capillary blood vessels at a depth of about 3 mm could be resolved. Meanwhile, the blood‐flow speeds in multiple individual vessels could be mapped simultaneously. Furthermore, NIR‐IIb tumor imaging of a live mouse was explored. NIR‐IIb imaging can be generalized to a wide range of fluorophores emitting at up to 1700 nm for high‐performance in vivo optical imaging.     

Optical Spectroscopy and Theoretical Studies in Calixarene Chemistry

This review summarizes different applications of optical spectroscopic methods in calixarene chemistry including UV/Vis spectrometry, vibrational spectroscopic techniques (FTIR and Raman spectroscopy), luminescence spectroscopy (including fluorescence and phosphorescence), ellipsometry and various optical microscopic methods. Moreover, the results of theoretical studies (AM1, PM3, DFT, ab initio, etc.) are summarized based on selected papers in the field of conformational studies, thermodynamics and complexation features. About 300 references are processed systematically from the results reported mainly in the recent years with emphasis on the potential of practical application of these molecules.     

Analysis by Raman and infrared spectroscopy combined with theoretical studies on the identification of plasticizer in PVC films

Raman and Fourier Transform infrared spectroscopy were used to study the vibrational spectra in commercial polyvinyl chloride (PVC) flexible films. In order to have insights of the PVC and the plasticizer structures, density functional theory calculations were conducted via B3LYP hybrid functional. The infrared and Raman calculations took into account geometry optimization of the PVC film, the plasticizer DEHA and the convolution of both. The convoluted spectra were then compared with the experimental data. Finally, the thermal stability of the PVC films was checked through one-hour thermal treatment of the samples comprising temperatures ranges from 50 to 200 °C.