Portable Shifted Excitation Raman Difference Spectroscopy (SERDS) using two excitation wavelengths around 785 nm is applied for selected applications in the field of agri-photonics. In the presence of daylight and laser-induced fluorescence, SERDS effectively separates Raman signals of green apple leaves and soil substances with more than 10-fold improved signal-to-background-noise ratios. Major ingredients of bovine milk are clearly detected and identified. A quantitative determination of the fat content in milk is performed and shows a limit-of-detection of 0.1 g / 100 mL. These results show a great potential of portable SERDS for real-world applications, e.g., for precision agriculture and food monitoring.
Diode laser based light sources (implemented monolithically or in a hybrid configuration) offer various functionalities to meet the requirements of specific applications. This includes tuning or switching between different wavelengths, modulating the optical output power, or implemented frequency conversion. Such light sources often contain multisection diode lasers or several active elements. Their operation requires multiple individually adjustable current sources, galvanically isolated current sources, and temperature control. A suitable optical interface should be available for a subsequent integration of the turnkey into the addressed application. In this contribution, a versatile turnkey system meeting the above-mentioned requirements will be presented. Ten p-type current sources, each with currents up to 750 mA, and four galvanically decoupled current sources are implemented. The ten individual sources enable switching frequencies up to 1 kHz and can be combined to provide currents up to 7.5 A. A temperature control unit capable to remove 10 W thermal load using a Peltier element completes the system, which contains an internal microcontroller, trigger in- and outputs, and an USB interface for the integration into various environments. Moreover, fiber coupling and free space optics to transfer the laser emission are offered. Turnkey systems containing in-house developed light sources at 488 nm or 785 nm were implemented into portable Raman spectroscopic measurement systems. To separate Raman signals from background disturbances, shifted excitation Raman difference spectroscopy (SERDS) was applied using dual-wavelength light sources. Systems addressing the measurement of carotenoids under clinical conditions and soil properties in the field will be presented.
Raman investigations are carried out to evaluate potential real-world application fields using an in-house developed portable shifted excitation Raman difference spectroscopy (SERDS) system at 785 nm. Pilot Raman measurements are performed in the presence of background light, laser-induced fluorescence, within optically turbid media and of weak Raman scatterers i.e., ambient air gases. In the presence of background light and laser-induce fluorescence, SERDS separates Raman signals of soil substances with a 15-fold improvement of the signal-to-background noise. Ingredients of bovine milk as the optically turbid test sample are clearly detected and identified. Ambient air gases are investigated even under daylight conditions. The results show the potential of the portable SERDS sensor systems for real-world applications,
Shifted excitation Raman difference spectroscopy (SERDS) has been successfully applied for on-site soil analysis. Here, a portable SERDS sensor system is developed for in-field investigations. A dual-wavelength diode laser emitting at 785 nm is integrated into an in-house realized turnkey laser system as the excitation source. For our experiments, an operating point for the two laser lines with a spectral distance of 10 cm-1 and 36 mW excitation power at the sample is selected. The sensor system allows a rapidly alternating operation between the two excitation wavelengths in the millisecond range and thus provides background-free SERDS spectra immediately after the measurement. This enables real-time evaluations, e.g., for quick on-site decisions. On-site soil analysis is carried out with the developed portable sensor system and SERDS extracts Raman signals of soil substances from background interferences with a 15-fold improvement of the signal-to-background-noise ratio. Besides others, closely neighbored Raman signals at 1084 cm-1 of calcite and at 1095 cm-1 of dolomite are identified and mixtures of both soil carbonates are successfully discriminated by using SERDS. The presented results demonstrate the capability of the portable SERDS sensor system for on-site soil analysis and furthermore show the potential for applications such as geological investigations.
Shifted excitation Raman difference spectroscopy (SERDS) is a powerful tool for the investigation of fluorescent samples such as biological materials. In case of rapidly changing emission backgrounds the efficiency of SERDS can however be limited as alternating detection of spectra excited at the two shifted laser wavelengths is usually restricted to sampling rates of less than 10 Hz. To overcome this issue, a novel optical lock-in detection approach enabling rapid SERDS operation in the kilohertz range using a custom 830-nm dual-wavelength diode laser and a specialized CCD enabling charge shifting on the CCD chip is presented. As an example of fluorescent and heterogeneous natural specimens, six mineral samples were selected and moved irregularly during spectral acquisition. Compared to conventional CCD read-out (operated at 5.4 Hz) the fast charge-shifting read-out performed at 1,000 Hz demonstrated superior reproducibility between repeat spectra. Using partial least squares-discriminant analysis an improved classification performance of the charge-shifting mode (sensitivity: 99 %, specificity: 94 %) over conventional read-out (sensitivity: 90 %, specificity: 92 %) was achieved. Translating the charge-shifting concept to sub-surface analysis using spatially offset Raman spectroscopy (SORS) enabled also the successful detection of charge-shifting SERDS-SORS spectra from a polytetrafluoroethylene layer concealed behind a 0.25 mm thick opaque heterogeneous layer. Chargeshifting SERDS-SORS results demonstrate two-fold improvement in signal-to-background-noise-ratio and match reference spectra much more closely. The charge-shifting approach shows large potential when rapidly changing background interference due to sample heterogeneity, dynamically evolving systems and ambient light variations presents a major challenges, e.g. in biological and biomedical applications.
Food safety and quality is of worldwide concern and for meat the authentication of different species is a frequent issue with many implications including economic, religious, ethical, and health issues. Common analytical methods for meat species authentication are mostly labor-intensive, time-consuming and expensive. Optical techniques are a promising alternative enabling rapid and non-invasive in-situ analysis. This study extends our previous investigations to the analysis of frozen-thawed meat and meat juice using Shifted Excitation Raman Difference Spectroscopy (SERDS) applying two miniaturized SERDS probes operating at 783 nm (110 mW optical power, 0.5 nm spectral shift) and 671 nm (40 mW optical power, 0.7 nm spectral shift) that are fiber-optically coupled to compact spectrometers. Specimens comprise pork, beef, chicken and turkey and for each species 12 fresh meat slices were frozen at -18 °C for 7 days. After thawing each slice was measured at 15 different spots while for the meat juice 5 drops originating from each slice were analyzed recording 10 spectra with integration times of 10 seconds each. Partial least squares discriminant analysis models using 4 latent variables showed a clear distinction between individual species for meat (Sensitivity < 94 %, Specificity < 92 %) and meat juice (Sensitivity < 97 %, Specificity < 98 %). The classification is based on variations in myoglobin content and complex differences in protein Raman bands. The results underline the large potential of SERDS for rapid and non-invasive in-situ meat authentication paving the way for future applications at selected points along the process chain.
Food authentication and the detection of adulterated products are recent major issues in the food industry as these topics are of global importance for quality control and food safety. To effectively address this challenge requires fast, reliable and non-destructive analytical techniques. Shifted Excitation Raman Difference Spectroscopy (SERDS) is well suited for identification purposes as it combines the chemically specific information obtained by Raman spectroscopy with the ability for efficient fluorescence rejection. The two slightly shifted excitation wavelengths necessary for SERDS are realized by specially designed microsystem diode lasers. At 671 nm the laser (optical power: 50 mW, spectral shift: 0.7 nm) is based on an external cavity configuration whereas an emission at 783 nm (optical power: 110 mW, spectral shift: 0.5 nm) is achieved by a distributed feedback laser. To investigate the feasibility of SERDS for rapid and nondestructive authentication purposes four types of cheese and three different cheese analogues were selected. Each sample was probed at 8 different positions using integration times of 3-10 seconds and 10 spectra were recorded at each spot. Principal components analysis was applied to the SERDS spectra revealing variations in fat and protein signals as primary distinction criterion between cheese and cheese analogues for both excitation wavelengths. Furthermore, to some extent, minor compositional differences could be identified to discriminate between individual species of cheese and cheese analogues. These findings highlight the potential of SERDS for rapid food authentication potentially paving the way for future applications of portable SERDS systems for non-invasive in situ analysis.
KEYWORDS: Bone, Raman spectroscopy, Signal detection, In vivo imaging, Raman scattering, Scattering, Spectroscopy, Diseases and disorders, Collagen, Minerals, Optical properties
Bone diseases and disorders are a growing challenge in aging populations; so effective diagnostic and therapeutic solutions are now essential to manage the demands of healthcare sectors effectively. Spatially offset Raman spectroscopy (SORS) allows for chemically specific sub-surface probing and has a great potential to become an in vivo tool for early non-invasive detection of bone conditions. Bone is a complex hierarchical material and the volume probed by SORS is dependent on its optical properties. Understanding and taking into account the variations in diffuse scattering properties of light in various bone types is essential for the effective development and optimization of SORS as a diagnostic in vivo tool for characterizing bone disease. This study presents SORS investigations at 830 nm excitation on two specific types of bone with differing mineralization levels. Thin slices of bone from horse metacarpal cortex (0.6 mm thick) and whale bulla (1.0 mm thick) were cut and stacked on top of each other (4-7 layers with a total thickness of 4.1 mm). To investigate the depth origin of the detected Raman signal inside the bone a 0.38 mm thin Teflon slice was used as test sample and inserted in between the layers of stacked bone slices. For both types of bone it could be demonstrated that chemically specific Raman signatures different from those of normal bone can be retrieved through 3.8-4.0 mm of overlying bone material with a spatial offset of 7-8 mm. The determined penetration depths can be correlated with the mechanical and optical properties of the specimens. The findings of this study increase our understanding of SORS analysis of bone and thus have impact for medical diagnostic applications e.g. enabling the non-invasive detection of spectral changes caused by degeneration, infection or cancer deep inside the bone matrix.
Raman Spectroscopy has become an important technique for assessing the composition of excised sections of bone, and is currently being developed as an in vivo tool for transcutaneous detection of bone disease using spatially offset Raman spectroscopy (SORS). The sampling volume of the Raman technique (and thus the amount of bone material interrogated by SORS) depends on the nature of the photon scattering in the probed tissue. Bone is a complex hierarchical material and to date little is known regarding its diffuse scattering properties which are important for the development and optimization of SORS as a diagnostic tool for characterizing bone disease in vivo. SORS measurements at 830 nm excitation wavelength are carried out on stratified samples to determine the depth from which the Raman signal originates within bone tissue. The measurements are made using a 0.38 mm thin Teflon slice, to give a pronounced and defined spectral signature, inserted in between layers of stacked 0.60 mm thin equine bone slices. Comparing the stack of bone slices with and without underlying bone section below the Teflon slice illustrated that thin sections of bone can lose appreciable number of photons through the unilluminated back surface. The results show that larger SORS offsets lead to progressively larger penetration depth into the sample; different Raman spectral signatures could be retrieved through up to 3.9 mm of overlying bone material with a 7 mm offset. These findings have direct impact on potential diagnostic medical applications; for instance on the detection of bone tumors or areas of infected bone.
Weak Raman bands are often covered by pronounced background signals due to fluorescence or Rayleigh scattering.
Several techniques to separate Raman lines from the background are known. In this paper, diode laser based light
sources will be presented suitable for shifted excitation Raman difference spectroscopy (SERDS). The two
wavelengths are realized by varying the injection current, by addressing two micro-integrated ECLs or by
temperature tuning.
Due to the freedom of choice in the wavelengths using diode lasers, the emission wavelength can be selected with
respect to the addressed application (e.g. the required penetration depth) or the plasmonic resonances of the
substrates for surface enhanced Raman spectroscopy. Devices were developed for the wavelengths 488 nm, 671 nm,
and 785 nm. The two emission wavelengths each were selected to have a spectral distance of 10 cm-1 according to
the typical width of Raman lines of solid or liquid samples. Output powers between 20 mW for the shorter
wavelength devices and 200 mW for the red emitting lasers were achieved at electrical power consumptions below
1 W. With a footprint of only 25 x 25 mm2 including all collimation and filter elements, these devices are well suited
for portable applications. The diode lasers were implemented into Raman measurement systems. The SERDS signal-to-background ratio was improved by several orders of magnitude.
Optical sensors based on Raman spectroscopy are suitable for a rapid identification and quantification of pollutants such
as Polycyclic Aromatic Hydrocarbons (PAHs). Additionally, Surface enhanced Raman spectroscopy (SERS) has gained
increasing attention as a powerful technique for in-situ monitoring of these substances in seawater to achieve limits of
detection (LODs) in the sub-nmol/l range.
A low-cost method based on electroless plating solution of chloroauric acid (HAuCl4) and hydrogen peroxide (H2O2)
was developed in our group to construct a gold island film as SERS substrate to achieve a well reproducible, high
sensitive and seawater resistant SERS sensor. The substrates show good resistance against seawater determined by long-term
stability tests carried out over 12 weeks of storage of the substrates in artificial seawater. The investigations show
that the substrates still have about 50 % of their initial activity after 4 weeks of storage and about 15 % after two months.
This type of substrate is reproducible with variability in the SERS intensities of about 8 %.
Shifted excitation Raman difference spectroscopy (SERDS) was applied by using a microsystem diode laser emitting at
784.3 nm and 784.8 nm to remove the fluorescence interference and to improve the Raman signals. This combination of
SERS and SERDS yields a limit of detection of 1 nmol/l for pyrene which was selected as representative PAH. These
quantitative results show that the designed SERS substrates are suitable for the in-situ monitoring of PAHs in the marine
environment.
Raman spectroscopy is a well established analytical method with applications in many areas, e.g. analysis of biological
samples. To overcome the problem of an undesired fluorescence background masking the Raman signals we present a
multi-spectral approach using shifted excitation Raman difference spectroscopy (SERDS). For our investigations we
applied microsystem diode lasers which realize two slightly shifted excitation wavelengths required to perform SERDS
at 488 nm, 671 nm, and 785 nm.
The emission at 488 nm with an optical power of up to 30 mW and a spectral shift of 0.3 nm (12 cm-1) is realized by
frequency doubling of a 976 nm distributed feedback (DFB) diode laser. The 671 nm laser diode contains two separate
laser cavities (spectral shift: 0.7 nm (13 cm-1)) each incorporating a volume Bragg grating as frequency selective
element. In that case, optical powers up to 50 mW can be obtained. For investigations at 785 nm we used a DFB laser
with a maximum optical power of 110 mW and a spectral shift of 0.5 nm (7 cm-1).
Meat, fat tissue, connective tissue and bones from pork and beef were used as test samples to demonstrate the effective
background removal using SERDS. For all three wavelengths integration times of only 5 - 10 seconds were necessary
showing the possibility of SERDS for rapid sample identification. A comparison with conventional Raman spectra is
given pointing out the improvement of spectral quality. The applicability of SERDS for other analytical applications, e.g.
medical diagnosis will be discussed.
The identification of food products and the detection of adulteration are of global interest for food safety and quality
control. We present a non-invasive in-situ approach for the differentiation of meat from selected animal species using
microsystem diode laser based shifted excitation Raman difference spectroscopy (SERDS) at 671 nm and 785 nm. In that
way, the fingerprint Raman spectra can be used for identification without a disturbing fluorescence background masking
Raman signals often occurring in the investigation of biological samples.
Two miniaturized SERDS measurement heads including the diode laser and all optical elements are fiber-optically
coupled to compact laboratory spectrometers. To realize two slightly shifted excitation wavelengths necessary for
SERDS the 671 nm laser (spectral shift: 0.7 nm, optical power: 50 mW) comprises two separate laser cavities each with
a volume Bragg grating for frequency selection whereas the 785 nm light source (spectral shift: 0.5 nm, optical power:
110 mW) is a distributed feedback laser.
For our investigations we chose the most consumed meat types in the US and Europe, i.e. chicken and turkey as white
meat as well as pork and beef as red meat species. The applied optical powers were sufficient to detect meat Raman
spectra with integration times of 10 seconds pointing out the ability for a rapid discrimination of meat samples. Principal
components analysis was applied to the SERDS spectra to reveal spectral differences between the animals suitable for
their identification. The results will be discussed with respect to specific characteristics of the analyzed meat species.
Due to its analytical ability and sensitivity to molecular vibrations, Raman spectroscopy provides valuable information of
the secondary structure of proteins. Moreover, polarized Raman spectroscopy is shown to be a useful instrument to
investigate the structural changes resulting from the aging and spoilage process of meat.
In this work, polarized Raman spectra were measured on oriented cuts of pork and turkey. Fresh meat slices were stored
at 5 °C and measured for a consecutive time period of 10 days. A 671 nm microsystem diode laser was used as excitation
light source. The laser power at the sample was 50 mW and the integration time of each Raman spectrum was set to
5 seconds. Measurements were performed with a laser beam orientation perpendicular to the long axis of the muscle
fibers. In that arrangement, the fibers were aligned either parallel or perpendicular to the polarization direction of the
laser source.
By using the statistical method of principal components analysis (PCA), a clear separation of the meat samples can be
found for fresh meat according to the orientation (parallel or perpendicular) using the first two principal components.
During the storage period, this separation subsequently vanishes due to the aging process and due to an increase of the
microbial spoilage of the meat surface. For the latter effect, a time-dependent distinction of the Raman spectra is
presented as well. Furthermore, specific changes of conformation-sensitive Raman bands were recognized, notably a
decrease of the intensities of α-helical protein conformation.
In-situ monitoring of pollutant chemicals in sea-water is of worldwide interest. For that purpose, fast response sensors
based on Raman spectroscopy are suitable for a rapid identification and quantification of these substances. Surface-enhanced
Raman scattering (SERS) was applied to achieve the high sensitivity necessary for trace detection. In the
project SENSEnet, funded by the European Commission, a SERS sensor based on calixarene-functionalized silver
nanoparticles embedded in a sol-gel matrix was developed and adapted for the in-situ detection of polycyclic aromatic
hydrocarbons (PAHs).
The laboratory set-up contains a microsystem Raman diode laser with two slightly different emission wavelengths (670.8
nm and 671.3 nm) suitable also for shifted excitation Raman difference spectroscopy (SERDS). The output power at
each of both wavelengths is up to 200 mW. For the detection of the SERS spectra integration times of typically 1 - 10
seconds were chosen. The SERS substrate is located inside a flow-through cell which provides continuous flow
conditions of the analyte. The spectra were recorded using a laboratory spectrograph with a back-illuminated deep
depletion CCD-detector.
We present scanning electron microscope images of the developed calixarene-functionalized Ag colloid based SERS
substrates as well as results for the SERS adsorption properties of major PAHs (pyrene, fluoranthene, and anthracene) in
artificial sea-water and their limits of detection (e. g. 0.1 nM for pyrene). The suitability of the presented device as an in-situ
SERS sensor for application on a mooring or buoy will be discussed.
Based on a miniaturized optical bench with attached 671 nm microsystem diode laser we present a portable Raman
system for the rapid in-situ characterization of meat spoilage. It consists of a handheld sensor head (dimensions: 210 x
240 x 60 mm3) for Raman signal excitation and collection including the Raman optical bench, a laser driver, and a
battery pack. The backscattered Raman radiation from the sample is analyzed by means of a custom-designed miniature
spectrometer (dimensions: 200 x 190 x 70 mm3) with a resolution of 8 cm-1 which is fiber-optically coupled to the sensor
head. A netbook is used to control the detector and for data recording.
Selected cuts from pork (musculus longissimus dorsi and ham) stored refrigerated at 5 °C were investigated in timedependent
measurement series up to three weeks to assess the suitability of the system for the rapid detection of meat
spoilage. Using a laser power of 100 mW at the sample meat spectra can be obtained with typical integration times of 5 -
10 seconds.
The complex spectra were analyzed by the multivariate statistical tool PCA (principal components analysis) to determine
the spectral changes occurring during the storage period. Additionally, the Raman data were correlated with reference
analyses performed in parallel. In that way, a distinction between fresh and spoiled meat can be found in the time slot of
7 - 8 days after slaughter. The applicability of the system for the rapid spoilage detection of meat and other food products
will be discussed.
Experimental results in shifted excitation resonance Raman difference spectroscopy (SERRDS) at 488 nm will be
presented. A novel compact diode laser system was used as excitation light source. The device is based on a distributed
feedback (DFB) diode laser as a pump light source and a nonlinear frequency doubling using a periodically poled lithium
niobate (PPLN) waveguide crystal. All elements including micro-optics are fixed on a micro-optical bench with a
footprint of 25 mm × 5 mm. An easy temperature management of the DFB laser and the crystal was used for wavelength
tuning. The second harmonic generation (SHG) provides an additional suppression of the spontaneous emission. Raman
spectra of polystyrene demonstrate that no laser bandpass filter is needed for the Raman experiments. Resonance-Raman
spectra of the restricted food colorant Tartrazine (FD&C Yellow 5, E 102) in distilled water excited at 488 nm
demonstrate the suitability of this light source for SERRDS. A limit of detection (LOD) of 0.4 μmol·l-1 of E102 enables
SERRDS at 488 nm for trace detection in e.g. food safety control as an appropriate contactless spectroscopic technique.
A hand-held Raman sensor head was developed for the in-situ characterization of meat quality. As light source, a
microsystem based external cavity diode laser module (ECDL) emitting at 671 nm was integrated in the sensor head and
attached to a miniaturized optical bench which contains lens optics for excitation and signal collection as well as a
Raman filter stage for Rayleigh rejection. The signal is transported with an optical fiber to the detection unit which was
in the initial phase a laboratory spectrometer with CCD detector.
All elements of the ECDL are aligned on a micro optical bench with 13 x 4 mm2 footprint. The wavelength stability is
provided by a reflection Bragg grating and the laser has an optical power of up to 200 mW. However, for the Raman
measurements of meat only 35 mW are needed to obtain Raman spectra within 1 - 5 seconds. Short measuring times are
essential for the hand-held device.
The laser and the sensor head are characterized in terms of stability and performance for in-situ Raman investigations.
The function is demonstrated in a series of measurements with raw and packaged pork meat as samples. The suitability
of the Raman sensor head for the quality control of meat and other products will be discussed.
Due to the narrow linewidth signals and its fingerprinting nature, Raman spectra provide information about the
molecular structure and composition of the samples. In this paper, the applicability of Raman spectroscopy is shown for
the in-situ characterization of the aging of meat. Miniaturized diode lasers are utilized as light sources with excitation
wavelengths of 671 nm and 785 nm with a view to the development of a portable field device for meat. As test sample,
musculus longissimus dorsi from pork was taken. The chops were stored refrigerated at 5 °C and Raman spectra were
measured daily from slaughter up to three weeks.
Throughout the entire period of one month, the Raman spectra preserve the basic spectral features identifying the
samples as meat. More specific, the spectra exhibit gradual changes of the Raman signals and they show a time-dependent
modification of the background signal which arises from a laser-induced fluorescence (LIF). To analyze the
time-correlation of the complex spectra, multivariate statistical methods are employed. By means of principal components
analysis (PCA) a distinction of spectra is found on the time scale between day 8 and 10. This corresponds to the
transition from ripened meat to meat at and beyond the limit of inedibility. After ca. 10 days of storage at 5 °C the
microbial load is overwhelming and LIF increases.
The results of the Raman measurements depending on the storage time of meat are discussed in the context of reference
analyses which have been performed in parallel.
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