Near-infrared Raman Spectroscopy Research Articles

基于NIR-SERS技术结合PCA-LDA统计分析肝癌术后恢复状况

基于NIR-SERS技术结合PCA-LDA统计分析肝癌术后恢复状况

Do formalin fixation and freeze‐thaw affect near‐infrared Raman spectroscopy of cartilaginous tissue? A preliminary ex vivo analysis of native human articular cartilage

Near‐infrared (NIR) Raman microprobe spectroscopy has been applied to the non‐invasive characterization of the biochemical structure of extracellular matrix in articular cartilage, a step forward along the path of in vivo diagnostic application of chondropathy. In most studies handling ex vivo cartilage specimens, formalin fixation or freeze‐thaw treatments have been applied in order to stabilize tissue and cell constituents prior to spectroscopic measurements. However, these pre‐processing manipulations might significantly affect certain target bands of the cartilage spectra, thus introducing biases in the characterizations, and potentially leading to data misinterpretation. In this study, we evaluated how formalin fixing and freeze‐thaw processes affect Raman spectra from human femur cartilage. Healthy cartilage specimens were fixed/stored either in a 10% neutral buffered formalin solution or in a deep freezer set at −80 °C. The results of this study show that formalin fixation significantly affects the NIR Raman spectra of cartilage specimens due to concurrent formalin absorption and water dehydration within both collagen and glycosaminoglycan macromolecules. Water dehydration was also confirmed in the amide I structure in the frozen‐thawed specimen, but to a much lesser extent. Furthermore, soaking the tissues in phosphate‐buffered saline solution minimized the storage‐induced Raman artifacts, but its immersion had limited effectiveness in formalin‐fixed specimens, predominantly due to an overlap of signals from the formalin liquid (i.e. emitting at 1046 and 1492 cm−1). Therefore, to provide a highly accurate biochemical evaluation of extracellular matrix using NIR Raman spectroscopy, freeze‐thaw processes are more suitable for ex vivo samples of human cartilage than formalin fixation. Copyright © 2015 John Wiley & Sons, Ltd.

Assessing the biochemical changes of tendons of rats in an experimental model of tenotomy under therapeutic ultrasound and LEDs (625 and 945nm) by near-infrared Raman spectroscopy.

Given the complexity of the tendon repair process, where the neoformed scar tissue tends to prevent or hamper the tendon from performing its normal functions, ultrasound (US) and light-emitting diode (LED) devices have been applied to modulate the tendon repair process. This study aimed to compare the anti-inflammatory and healing effects of therapeutic US and LED in an experimental model of tenotomy using histomorphometry and Raman spectroscopy. Forty Wistar rats (235 ± 8.3g) were separated into four groups: tenotomy (TC; control), tenotomy associated with ultrasound (US; 1MHz, 0.5W/cm(2)), tenotomy associated with LED at 625nm (LR; 4J/cm(2)), and tenotomy associated with LED at 945nm (LIR; 4J/cm(2)), in two experimental periods of 7 and 14days. Histomorphometry showed a decrease of inflammatory cells (macrophages and polymorphonuclear leukocytes) for the group treated with US compared to the TC group (ANOVA, p < 0.0001) and the LIR group compared to the TC group (p < 0.05) at the seventh day, indicating an anti-inflammatory effect. In relation to the LR and TC groups, there was no significant difference. Raman spectroscopy revealed that the LR group presented an increased synthesis of collagen I compared to the other treatments and the TC groups on the 14th day (p < 0.01). US therapy showed an adjuvant effect with anti-inflammatory action, and the LED therapy at 625nm showed an increase in collagen synthesis, contributing to the process of tendon healing.

Detecting alterations of glucose and lipid components in human serum by near-infrared Raman spectroscopy

Introduction Raman spectroscopy may become a tool for the analysis of glucose and triglycerides in human serum in real time. This study aimed to detect spectral differences in lipid and glucose components of human serum, thus evaluating the feasibility of Raman spectroscopy for diagnostic purposes. Methods A total of 44 samples of blood serum were collected from volunteers and submitted for clinical blood biochemical analysis. The concentrations of glucose, cholesterol, triglycerides, and low-density and high-density lipoproteins (LDL and HDL) were obtained using standard biochemical assays. Serum samples were placed in Eppendorf tubes (200 µL), kept cooled (5 °C) and analyzed with near-infrared Raman spectroscopy (830 nm, 250 mW, 50 s accumulation). The mean spectra of serum with normal or altered concentrations of each parameter were compared to determine which Raman bands were related to the differences between these two groups. Results Differences in peak intensities of altered sera compared to normal ones depended on the parameter under analysis: for glucose, peaks were related to glucose; for lipid compounds the main changes occurred in the peaks related to cholesterol, lipids (mainly triolein) and proteins. Principal Components Analysis discriminated altered glucose, cholesterol and triglycerides from the normal serum based on the differences in the concentration of these compounds. Conclusion Differences in the peak intensities of selected Raman bands could be seen in normal and altered blood serum samples, and may be employed as a means of diagnosis in clinical analysis.

Doping inhomogeneity and staging of ultra-thin graphite intercalation compound flakes probed by visible and near-infrared Raman spectroscopy**Project supported by the National Natural Science Foundation of China (Grant Nos. 11225421, 11474277, and 11434010).

When ultra-thin graphite intercalation compounds (GICs) are deposited on the SiO2/Si substrate, it is found that their colors are dependent on the thickness of GIC flakes. The sample colors of ultrathin GIC flakes can no longer provide qualitative information on the stage index. Here, multi-wavelength Raman spectroscopy is thus applied to study the doping inhomogeneity and staging of ultra-thin GICs by FeCl3 intercalation. The G band intensity of stage-1 GIC flakes is strongly enhanced by 532-nm laser excitation, while that of stage-2 and stage-3 flakes exhibits strong intensity enhancement for 785-nm laser excitation. The near-infrared lasers are suggested to probe the doping inhomogeneity and staging of ultra-thin GIC flakes.

Correlation between METAVIR scores and Raman spectroscopy in liver lesions induced by hepatitis C virus: a preliminary study.

The viral hepatitis C is one of the most important causes of chronic hepatic illness worldwide, affecting around 3 % of the world population. Raman spectroscopy has been employed to distinguish normal from hepatic lesions through differences in the spectral features related to the METAVIR score system. This preliminary study evaluated 11 patients with diagnoses of chronic hepatitis C who underwent hepatic biopsies; the biopsies were submitted to near-infrared Raman spectroscopy using a dispersive spectrometer (830-nm wavelength, 300-mW laser power, and 20-s exposure time). The METAVIR was further scored, and the spectra were submitted to principal component analysis (PCA). The results show a good correlation between the Raman spectroscopy features and the stage of hepatic inflammation and fibrosis. PCA showed that samples with a higher degree of fibrosis presented a higher amount of protein features (collagen), whereas samples with a higher degree of inflammation presented higher features of hemoglobin, in accordance to the expected evolution of chronic hepatitis. Quinone was found to be an important biomarker in early hepatic lesions with a spectral feature at 1595 cm(-1). This study demonstrates that Raman spectroscopy may become an important tool for diagnosing liver disease.

Discrimination of non‐melanoma skin lesions from non‐tumor human skin tissues in vivo using Raman spectroscopy and multivariate statistics

Raman spectroscopy was used to discriminate human non-melanoma skin lesions from non-tumor tissues in vivo. This work proposed the discrimination between non-melanoma (basal cell carcinoma, BCC; squamous cell carcinoma, SCC) and pre-cancerous lesions (actinic keratosis, AK) from benign lesions and normal (non-tumor group, NT) tissues, using near-infrared Raman spectroscopy with a Raman probe. Prior to surgery, the spectra of suspicious lesions were obtained in situ. The spectra of adjacent, clinically normal skin were also obtained. Lesions were resectioned and submitted for histopathology. The Raman spectra were measured using a Raman spectrometer (830 nm). Two types of discrimination models were developed to distinguish the different histopathological groups. The principal components analysis discriminant analysis (PCA/DA) and the partial least squares discriminant analysis (PLS/DA) were based on Euclidean, quadratic and Mahalanobis distances. PCA and PLS spectral vectors showed spectral features of skin constituents, such as lipids (between 1,250 cm(-1) and 1,300 cm(-1) and at 1,450 cm(-1)) and proteins (between 870 cm(-1) and 940 cm(-1), 1,240 cm(-1) and 1,271 cm(-1), and at 1,000 cm(-1) and 1,450 cm(-1)). Despite the small spectral differences between malignant lesions and benign tissues, the algorithms discriminated the spectra of non-melanoma skin and pre-cancerous lesions from benign and normal tissues, with an overall accuracy of 82.8% and 91.9%, respectively. PCA and PLS could discriminate Raman spectra of skin tissues, opening the way for an in vivo optical diagnosis.

Identification of different forms of cocaine and substances used in adulteration using near-infrared Raman spectroscopy and infrared absorption spectroscopy.

Identification of cocaine and subsequent quantification immediately after seizure are problems for the police in developing countries such as Brazil. This work proposes a comparison between the Raman and FT-IR techniques as methods to identify cocaine, the adulterants used to increase volume, and possible degradation products in samples seized by the police. Near-infrared Raman spectra (785 nm excitation, 10 sec exposure time) and FT-IR-ATR spectra were obtained from different samples of street cocaine and some substances commonly used as adulterants. Freebase powder, hydrochloride powder, and crack rock can be distinguished by both Raman and FT-IR spectroscopies, revealing differences in their chemical structure. Most of the samples showed characteristic peaks of degradation products such as benzoylecgonine and benzoic acid, and some presented evidence of adulteration with aluminum sulfate and sodium carbonate. Raman spectroscopy is better than FT-IR for identifying benzoic acid and inorganic adulterants in cocaine.

Identification and characterization of colorectal cancer using Raman spectroscopy and feature selection techniques.

This study aims to detect colorectal cancer with near-infrared Raman spectroscopy and feature selection techniques. A total of 306 Raman spectra of colorectal cancer tissues and normal tissues are acquired from 44 colorectal cancer patients. Five diagnostically important Raman bands in the regions of 815-830, 935-945, 1131-1141, 1447-1457 and 1665-1675 cm(-1) related to proteins, nucleic acids and lipids of tissues are identified with the ant colony optimization (ACO) and support vector machine (SVM). The diagnostic models built with the identified Raman bands provide a diagnostic accuracy of 93.2% for identifying colorectal cancer from normal Raman spectroscopy. The study demonstrates that the Raman spectroscopy associated with ACO-SVM diagnostic algorithms has great potential to characterize and diagnose colorectal cancer.

XRD and Raman spectroscopy studies of (Bi1–xLaxFeO3)0.5(PbTiO3)0.5 solid solution

(Bi1–xLaxFeO3)0.5(PbTiO3)0.5 solid solution obtained by mechanochemical synthesis was characterized by X-ray diffraction (XRD), atomic force microscopy and Fourier transform near-infrared Raman spectroscopy. The single-phase solid solution with La content between x = 0 and 0.5 was prepared. The solid solution exhibits a tetragonal symmetry P4mm for 0 ≤ x ≤ 0.2. It was found that Raman spectra undergo a significant change for the solid solution with La content between x = 0.16 and 0.20. It may indicate the existence of a morphotropic boundary in this range of the La content. Detailed XRD study on (Bi0.5La0.5FeO3)0.5(PbTiO3)0.5 pointed to a rhombohedral symmetry R3c.

Raman, surface-enhanced Raman, and density functional theory characterization of (diphenylphosphoryl)(pyridin-2-, -3-, and -4-yl)methanol.

This work presents near-infrared Raman spectroscopy (FT-RS) and surface-enhanced Raman scattering (SERS) studies of three pyridine-α-hydroxymethyl biphenyl phosphine oxide isomers: (diphenylphosphoryl)(pyridin-2-yl)methanol (α-Py), (diphenylphosphoryl)(pyridin-3-yl)methanol (β-Py), and (diphenylphosphoryl)(pyridin-4-yl)methanol (γ-Py) adsorbed onto colloidal and roughened in oxidation-reduction cycles silver surfaces. The molecular geometries in the equilibrium state and vibrational frequencies were calculated by density functional theory (DFT) at the B3LYP 6-311G(df,p) level of theory. The results imply that the most stable structure of the investigated molecules is a dimer created by two intermolecular hydrogen bonds between the H atom of the α-hydroxyl group (in up (HOU) or down (HOD) stereo bonds position) and the O atom of tertiary phosphine oxide (═O) of the two monomers. Comparison the FT-RS spectra with the respective SERS spectra allowed us to predict the orientation of the hydroxyphosphonate derivatives of pyridine that depends upon both the position of the substituent relative to the ring N atom (in α-, β-, and γ-position, respectively) and the type of silver substrate.

Nondestructive discrimination between normal and hematological malignancy cell lines using near-infrared Raman spectroscopy and multivariate analysis

An accurate understanding of biomolecular changes in living cells associated with malignant transformation is of paramount importance in early cancer detection. The aim of this study was to apply near-infrared Raman spectroscopy (RS) for differentiating cancer from normal cells. High-quality Raman spectra in the range of 450–1800 cm−1 can be obtained from 31 normal and 64 hematological malignancy cells including 43 CA46 and 21 U266 cells. There were significant differences in Raman spectra between normal and cancer groups, which suggests special changes in the percentage of biomolecules including lipid, nucleic acids and proteins in different cell lines. A diagnostic accuracy of 100% can be achieved by principal components analysis (PCA) combined with linear discriminant analysis (LDA) for classification between cancer and normal cell lines. This exploratory study demonstrates the potential application of the RS technique combined with PCA–LDA as a clinical cell-based biosensor for the noninvasive cancer detection and screening at the molecular level.

Detection of Urea Adulteration in Milk Using Near-Infrared Raman Spectroscopy

We report results of a spectroscopic study to assess the applicability of near-infrared Raman spectroscopy for quantitative determination of urea adulteration in milk without any preprocessing requirement. Different batches of milk adulteration were prepared with each batch containing milk samples added with urea at various concentrations ranging from 10 to 1,000 mg/dl. The Raman spectra were measured from these urea-mixed milk samples with a Raman spectroscopy setup that incorporates a 785-nm diode laser for Raman excitation. An algorithm based on partial least square (PLS) regression was developed to quantitatively predict the concentration of urea based on the measured sets of Raman spectra of the urea-mixed milk samples. The results showed that Raman spectroscopy in combination with the PLS-based chemometric algorithm could detect urea mixed in milk samples with an accuracy of >90 %.

Activation and deactivation of vibronic channels in intact phycocyanin rods

We investigated the excitation modes of the light-harvesting protein phycocyanin (PC) from Thermosynechococcus vulcanus in the crystalline state using UV and near-infrared Raman spectroscopy. The spectra revealed the absence of a hydrogen out-of-plane wagging (HOOP) mode in the PC trimer, which suggests that the HOOP mode is activated in the intact PC rod, while it is not active in the PC trimer. Furthermore, in the PC trimer an intense mode at 984 cm(-1) is assigned to the C-C stretching vibration while the mode at 454 cm(-1) is likely due to ethyl group torsion. In contrast, in the similar chromophore phytochromobilin the C5,10,15-D wag mode at 622 cm(-1) does not come from a downshift of the HOOP. Additionally, the absence of modes between 1200 and 1300 cm(-1) rules out functional monomerization. A correlation between phycocyanobilin (PCB) and phycoerythrobilin (PEB) suggests that the PCB cofactors of the PC trimer appear in a conformation similar to that of PEB. The conformation of the PC rod is consistent with that of the allophycocyanin (APC) trimer, and thus excitonic flow is facilitated between these two independent light-harvesting compounds. This excitonic flow from the PC rod to APC appears to be modulated by the vibration channels during HOOP wagging, C = C stretching, and the N-H rocking in-plan vibration.

Near-infrared Raman spectroscopy for assessing biochemical changes of cervical tissue associated with precarcinogenic transformation.

Raman spectroscopy measures the inelastically scattered light from tissue that is capable of identifying native tissue biochemical constituents and their changes associated with disease transformation. This study aims to characterize the Raman spectroscopic properties of cervical tissue associated with the multi-stage progression of cervical precarcinogenic sequence. A rapid-acquisition fiber-optic near-infrared (NIR) Raman diagnostic system was employed for tissue Raman spectral measurements at 785 nm excitation. A total of 68 Raman spectra (23 benign, 29 low-grade squamous intraepithelial lesions (LSIL) and 16 high grade squamous intraepithelial lesions (HSIL)) were measured from 25 cervical tissue biopsy specimens, as confirmed by colposcopy-histopathology. The semi-quantitative biochemical modeling based on the major biochemicals (i.e., DNA, proteins (histone, collagen), lipid (triolein) and carbohydrates (glycogen)) in cervical tissue uncovers the stepwise accumulation of biomolecular changes associated with progressive cervical precarcinogenesis. Multi-class partial least squares-discriminant analysis (PLS-DA) together with leave-one tissue site-out, cross-validation yielded the diagnostic sensitivities of 95.7%, 82.8% and 81.3%; specificities of 100.0%, 92.3% and 88.5%,for discrimination among benign, LSIL and HSIL cervical tissues, respectively. This work suggests that the Raman spectral biomarkers have identified the potential to be used for monitoring the multi-stage cervical precarcinogenesis, forming the foundation of applying NIR Raman spectroscopy for the early diagnosis of cervical precancer in vivo at the molecular level.

Study on Fresh Breast Tissues by Near-infrared Raman Spectroscopy

A portable Raman spectrometer was used for distinguishing the characteristics of normal,malignant and benign fresh breast biopsy samples.Based on spectral profiles,the presence of lipids(1230,1268,1301,1440,1743 cm-1) is indicated in normal tissue.And proteins(amide I,and amide Ⅲ,1246,1271,1315,1364 cm-1) are found in benign and malignant tissues.Between benign and malignant,nucleic acids(1340 cm-1) are found to be good discrimination parameters.In the process of data analysis,the model was set up by Gaussian Process with the intensity of the feature,and obtained the specificity(0.94),sensibility(0.95) and Matthews correlation coefficient(MCC,0.86).This study shows the significance in diagnosing breast disease,and contributes fundamentally to further application on clinic.

Discrimination of premalignant lesions and cancer tissues from normal gastric tissues using Raman spectroscopy

The feasibility of early detection of gastric cancer using near-infrared (NIR) Raman spectroscopy (RS) by distinguishing premalignant lesions (adenomatous polyp, n=27) and cancer tissues (adenocarcinoma, n=33) from normal gastric tissues (n=45) is evaluated. Significant differences in Raman spectra are observed among the normal, adenomatous polyp, and adenocarcinoma gastric tissues at 936, 1003, 1032, 1174, 1208, 1323, 1335, 1450, and 1655 cm(-1). Diverse statistical methods are employed to develop effective diagnostic algorithms for classifying the Raman spectra of different types of ex vivo gastric tissues, including principal component analysis (PCA), linear discriminant analysis (LDA), and naive Bayesian classifier (NBC) techniques. Compared with PCA-LDA algorithms, PCA-NBC techniques together with leave-one-out, cross-validation method provide better discriminative results of normal, adenomatous polyp, and adenocarcinoma gastric tissues, resulting in superior sensitivities of 96.3%, 96.9%, and 96.9%, and specificities of 93%, 100%, and 95.2%, respectively. Therefore, NIR RS associated with multivariate statistical algorithms has the potential for early diagnosis of gastric premalignant lesions and cancer tissues in molecular level.

Raman spectroscopy investigation of load-assisted microstructural alterations in human knee cartilage: Preliminary study into diagnostic potential for osteoarthritis

A preliminary investigation into the diagnostic potential of Raman spectroscopy for assessing pathological articular cartilage was conducted. Five arthritic human tibial cartilages retrieved after total knee arthroplasty were examined using near-infrared (NIR) Raman spectroscopy excited with 647.1nm lines of a Kr-ion laser. A “healthy” cartilage obtained from cadaver donor was also examined as a control sample. Degradation severity was first visually classified into five grades (Grade 0–VI) on the surface of both medial and lateral zones in each tibial plateau, according to the Collins scale. Raman spectra were then collected from selected zones with different damage severity. A systematic increase in relative intensity ratio between the Raman bands located at 1241 and 1269cm−1 (amide III doublet) was observed with increasing degradation grades, which could be related to structural changes under loading in type II collagen. This finding suggests that the present spectroscopic approach might be useful for recognizing and quantitatively assessing the degree of osteoarthritis (OA) in its early manifestation stage.

Rapid and in Vivo Quantification of Cellular Lipids in Chlorella vulgaris Using Near-Infrared Raman Spectrometry

A rapid and noninvasive quantification method for cellular lipids in Chlorella vulgaris is demonstrated in this study. This method applied near-infrared Raman spectroscopy to monitor the change of signal intensities at 1440 cm(-1) and 2845-3107 cm(-1) along the nitrogen depletion period, and calibration curves relating signal intensity and cellular lipid abundance were established. The calibration curves show that signal intensity at 2845-3107 cm(-1) and cellular lipid abundance were highly correlated. When the calibration curve was applied on the lipid quantification of two unknown samples, the differences between lipid abundances estimated by the calibration curve and measured by gas chromatography were less than 2 wt %. Carotenoids produced a strong and broad peak near 1440 cm(-1), and it weakened the correlation between signal intensity and lipid abundance. The consistency of detection and effects of cellular contents and water on the Raman spectrogram of Chlorella vulgaris were also addressed. The sample pretreatment only involved centrifugation, and the time required for lipid quantification was shortened to less than 1.5 h. The rapid detection has great potential in high-throughput screening of microalgae and also provides valuable information for monitoring the quality of microalgae culture and determining parameters for the mass production of biodiesel from microalgae.

Towards an on-line characterization of liquid bovine milk molecular content using Raman spectroscopy methods

Non-resonant near-infrared Raman spectroscopy supported by statistical analysis techniques was used to determine the molecular content of raw milk in flow. We created first a spectral library of Raman lines of the pivotal biomolecules included in milk in the pure state: fat globules, proteins (whey and caseins), lactose, and progesterone. Using a fluid chamber with windows coated with an anti-biocoating layer, we acquired the Raman spectra of milk from cows over a one-year period covering all seasons. Combining these spectra with the spectral library and a multivariate statistical analysis technique, Partial Least Square regression, we elaborated the PLS-regression model. This model permitted us to quantify the concentration of fat, proteins, lactose, and progesterone in unidentified samples with a root mean square error of cross validation of 0.37%, 0.14%, 0.13%, 4ngmL−1, respectively. This method paves the way for the on-line characterization of raw milk in the real conditions of its production.