Articles

Special Issues November 2018

Click the title above to open the Spectroscopy November 2018 Supplement, Advances in Portable and Handheld Spectroscopy, in an interactive PDF format.

Spectroscopy November 2018 Advances in Portable and Handheld Spectroscopy Supplement PDF

Content uniformity testing is a crucial task in pharmaceutical manufacturing, as it ensures that each product that reaches a consumer contains safe dosages of all components. Recently, transmission Raman spectroscopy has been explored as an alternative technique for content uniformity testing, due to its ability to rapidly obtain quantitative measurements representative of the whole volume of a tablet. In this case study, test tablets were created from five different blends of acetaminophen, mannitol, lactose, microcrystalline cellulose, and magnesium stearate. A portable Raman spectrometer connected to a high-efficiency transmission module was used to collect calibration spectra. The spectra were then used to create partial least squares (PLS) models for the acetaminophen and lactose components (ranging from 10-18% w/w and 10.79-28.66% w/w, respectively). Chemometric models can be used for prediction of acetaminophen and lactose conventrations in manufactured tablets to perform fast and accurate at-line content uniformity testing with portable Raman transmission spectroscopy. 

	Content uniformity (CU) testing is a crucial task in pharmaceutical manufacturing, as it ensures that each product that reaches consumers contains a safe dosage of the active pharmaceutical ingredient (API). Traditionally, high performance liquid chromatography (HPLC) is performed off-line in a quality control laboratory to monitor the dosage of finished tablets, due to its sensitivity and the ability to obtain a measurement from the entire volume of the tablet (1,2). However, HPLC analysis is a destructive technique that involves large amounts of solvents and consumables, and often requires highly skilled personnel to perform analyses. Depending on the workload of a QC lab, it may take several hours to complete analyses. Consequently, if a batch is found to be outside of specifications, the process defects leading to the deviation may not be found in a timely manner, potentially causing manufacturing delays for pharmaceutical drugs in heavy demand.

	Analytical methods for CU testing should ideally be fast, noninvasive, and with limited sample preparation. Recently, transmission near infrared (NIR) spectroscopy and transmission Raman spectroscopy have both been explored as alternative methods for nondestructive CU testing with no sample preparation (3-7). Although quick and nondestructive, transmission NIR spectroscopy suffers from poor chemical selectivity, and is sensitive to changes in the testing environment. Transmission Raman spectroscopy, combined with chemometric modeling, is quickly emerging as a valued technique for CU testing, due to its high chemical specificity, which is particularly useful when dealing with complex pharmaceutical formulations that contain multiple components.

	Conventionally, Raman spectrometers collect backscattered signal that is generated upon illumination of the sample surface with a laser. While conventional Raman spectroscopy is quite suitable for identity tests of incoming raw pharmaceutical materials, it does not provide an accurate quantitative measurement representative of the entire volume of a tablet, nor is it able to probe sufficiently beneath layers of coatings on finished tablets. Transmission Raman spectroscopy, however, is able to overcome the limitations of conventional Raman spectroscopy for content uniformity testing. Using transmission Raman spectroscopy, the sample is illuminated at one side, and the Raman signal is collected from the other side. The signal collected contains photons with information that is representative of the full volume of the tablet. Through the use of chemometric models, an accurate quantitative prediction of the content within the tablet can be achieved. This allows for not only reliable content uniformity testing, but can also assist formulation development and detection of pharmaceutical counterfeits.

	Although transmission Raman spectroscopy provides measurements representative of the full volume of a tablet, and is simple to operate with the use of predictive chemometric models, transmission Raman spectroscopy is generally a low- efficiency technique that requires long integration times to obtain sufficient Raman signal. In this case study, we demonstrate proof of concept for CU analysis using a portable Raman spectrometer with a high-efficiency transmission sampling attachment. A series of tablet formulations containing acetaminophen, mannitol, lactose, cellulose, and magnesium stearate were measured on a portable transmission Raman spectrometer. Chemometric models for at-line accurate prediction of acetaminophen and lactose were created using model building software. At-line high-throughput transmission Raman spectroscopy allows for increased efficiency of CU testing, and can quickly catch defects present in the tablet manufacturing process.

	Blended powders and pressed tablets were provided by courtesy of the Engineering Research Center for Structured Organic Particulate Systems of the Department of Chemical and Biochemical Engineering at Rutgers University (Piscataway, New Jersey). Formulations were blended with laboratory grade acetaminophen, mannitol, lactose, microcrystalline cellulose (MCC), and magnesium stearate. Five different blends were created. The concentration of acetaminophen in these blends ranged from 10-18% (w/w), and the concentration of lactose ranged from 10.79-28.66% (w/w). Blends were compressed to create round tablets approximately 6 mm in diameter and 4.5 mm thick using a Gamlen tablet press. Tablets were pressed with an approximate force of 392 kg. Each pill had a typical mass of ~140 mg. Ten tablets were manufactured from each blend. Table I lists the concentration of ingredients for all blends.


	Table I: Concentrations of ingredients in sample tablets (w/w %)

	The QTRam from B&W Tek with BWAnalyst software was used for all measurements. The QTRam uses a transmission Raman attachment for increased photon efficiency, with a laser spot size of ~4 mm. The system contains a 785 nm laser, with a maximum power of ~340 mW. For method development, method validation, and prediction analyses, all measurements used 100% of the maximum laser power, and a 20 second integration time. Measurements were performed on both sides of the tablets for an accurate representation of the full volume of the tablet. BWIQ software was used to create predictive chemometric models.

	Seven tablets from each blend were designated as method samples. Triplicate transmission Raman spectra were collected for each sample. Figure 1 shows representative transmission Raman spectra from a tablet containing 18% acetaminophen (Blend 1) and a tablet containing 28.66% lactose (Blend 4), compared to pure acetaminophen and lactose. Spectral regions that correspond to the signal from acetaminophen and the signal from lactose were isolated for chemometric modeling. The spectral regions used in the model are highlighted in blue and gray for lactose and acetaminophen, respectively. The spectral regions of 1190-1300 cm

 were isolated in the model spectra because their peaks correspond purely to acetaminophen. Spectral regions of 285-500 cm

 were isolated for lactose because their peaks contain contributions from lactose. Samples from all blends show contributions from both acetaminophen and lactose in these spectral regions. Peaks that were attributed to the other components of the blends were not used in the modeling.


	Figure 1: Transmission Raman spectra from two tablet formulation samples compared to acetaminophen and lactose. The blue shaded regions correspond to the areas for lactose modeling. The gray shaded regions correspond to the areas for acetaminophen modeling. Spectra are manually offset.

	The chemometric software randomly chose 70% of the data as calibration spectra, and designated the remaining 30% as validation spectra. A leverage outlier test found no statistical outliers. Several preprocessing steps were applied to the data in order to minimize background effects and baseline offsets. An adaptive, iteratively reweighted penalized least squares (air-PLS) background removal was used in order to mitigate the background effects from the excipients, and a multiplicative scatter correction (MSC) was used to offset effects from baseline differences. A first derivative (Savitzky-Golay, window 11, order 3) was also applied to the data. Figure 2 shows the method spectra after the applied preprocessing steps. A partial least squares (PLS) regression was chosen to model the data.


	Figure 2: Method spectra after air-PLS background correction, MSC, and Savitzky-Golay first derivative.

	Figure 3 shows the plots of the predicted vs. measured values for the acetaminophen and lactose PLS models. Blue samples represent calibration samples, and red samples represent validation samples. Four factors were used in the acetaminophen model, while five factors were used in the lactose model. The R

 coefficient of determination is a measure of how close the data are to the regression line. The R

 value of the calibration spectra for acetaminophen and lactose models were 0.9929 and 0.9915, respectively, indicating good linearity of the models. The root mean square error (RMSE) of the acetaminophen calibration curve was 0.233% and the RMSE of the lactose calibration curve is 0.531%.


	Figure 3: (a) Predicted vs measured plot of the calibration and validation samples for the acetaminophen PLS model. The wavenumber regions used in the model are from 1190-1300 cm

. Four factors are used in the model. (b) Predicted vs measured plot of the calibration and validation samples for the lactose PLS model. The wavenumber regions used in the model are from 285-500 cm

	Before the method can be released to production for CU analysis, method validation must be completed to make sure that the method is suitable for the tablet CU test. According to USP chapter <1125>, CU testing falls under the analytical category I test, which requires method validation on method specificity, linearity, accuracy, precision and range. For independent method validation, two samples were selected for each blend. Triplicate measurements of each sample were acquired on the spectrometer. The specificity validation was checked using the Mahalanobis distance and Q-residual limit.

	Linearity, accuracy, and precision were obtained through the built-in module for method validation. Table II shows the method validation results for the ten validation samples. Both acetaminophen and lactose models show good linearity, with R

 reported as 0.9889 for acetaminophen, and 0.9923 for lactose. Both models also show good precision, with the standard deviations for samples ranging from 0.02-0.17% and 0.13-0.36% for acetaminophen and lactose, respectively. The root mean square error of prediction (RMSEP) was calculated as 0.288% for the acetaminophen model, and 0.504% for the lactose model.


	Table II: Method validation results for acetaminophen and lactose PLS models

	The created chemometric models were used to predict the acetaminophen and lactose contents of new samples in the data analysis software. One tablet per blend was chosen as an independent sample to test the prediction of the acetaminophen and lactose content using the models created. Three spectra were collected for each sample. The prediction results for each tablet are listed in Table III.


	Table III: Prediction results for acetaminophen and lactose

	Portable high-efficiency transmission Raman spectroscopy is an effective tool for rapid, at-line content uniformity testing. We demonstrate through a matrix of tablets of varying compositions that portable transmission Raman is able to build methods that can accurately quantitate the acetaminophen and lactose components. As an alternative method of content uniformity testing, portable transmission Raman spectroscopy provides a simple and nondestructive analysis method, with no solvent consumption and no consumables at the point of measurement.

	(3) N.W. Broad, R.D. Jee, A.C. Moffat and M.R. Smith, 

	(4) C. Peroza Meza, M. A. Santos, and R. J. Romañach, 

	(5) M.D. Hargreaves, N. A. Macleod, M.R. Smith, D. Andrews, S.V. Hammond, and P. Matousek,

	(6) D. Andrews, K. Geentjens, B. Igne, G. McGeorge, A. Owen, N. Pedge, J. Villaumié, and V. Woodward, 

	(7) M. Fransson, J. Johansson, A. Sparén and O. Svensson, 

Kristen Frano, John Maticchio, and Dawn Yang

 are with B&W Tek in Newark, DE. Direct correspondence to: 

Portable transmission Raman spectroscopy, combined with chemometric modeling, is quickly emerging as a valued technique for content-uniformity testing, given its high chemical specificity, which is particularly useful when dealing with complex pharmaceutical formulations that contain multiple components.

Portable, High-Efficiency Transmission Raman Spectroscopy for At-Line Content Uniformity Testing of Pharmaceutical Tablets

Recently, miniaturization of Raman, mid-infrared (MIR) and near infrared (NIR) spectrometers has made substantial progress, and marketing companies predict this segment of instrumentation will have a significant growth rate within the next few years. This increase will be based on a wider adoption of spectrometers for quality control by in-the-field testing and on-site measurements. Furthermore, based on high-volume manufacturability and significant reduction of costs, numerous companies target with (primarily NIR) instruments a non-expert user community for consumer applications. Especially from this last-mentioned development, a tremendous potential for everyday life can be expected ranging from food testing to detection of fraud and adulteration in a broad area of materials. Miniaturization, however, must not lead to compromises in measurement performance and the handheld instrumentation will only have a practical impact if Raman, MIR, and NIR spectra of comparable quality to laboratory spectrometers can be obtained. The present communication will shortly cover the building principles of state-of-the-art instruments, and will then discuss selected applications in order to demonstrate the advancement in this field of research.

	Miniaturization of vibrational spectrometers has started more than two decades ago, but only within the last decade real handheld Raman, MIR and NIR scanning spectrometers have become commercially available, and utilized for a broad range of analytical applications (1-6). While the weight of the majority of Raman and MIR spectrometers is still in the ~1 kg range, the miniaturization of NIR spectrometers has advanced down to the ~100 g level, and developments are under way to integrate them into mobile phones (7). Furthermore, Raman and MIR handheld spectrometers are still in the price range of several tens of thousands of US$, whereas miniaturized NIR systems have recently reached the < $1,000 USD level. In view of the high price level of Raman and MIR instruments in the near future, only the acquisition of NIR systems can be taken into consideration for private use, whereas handheld Raman and MIR spectrometers will be restricted to industrial, military, and homeland security applications, as well as public use by first responders, customs, or environmental institutions.

	In what follows, instrumental features of novel handheld Raman, MIR and NIR spectrometers will be discussed, and selected qualitative and quantitative case studies will be presented. Material quality control applications have been selected for Raman and MIR. Notwithstanding the indisputable value of handheld NIR for the same purpose, only everyday life examples for the private sector will be discussed here; this is to enhance the interest of a new non-expert user community.

	Presently, only two handheld Fourier transform infrared (FT-IR) spectrometers (based on Michelson interferometers), and that can be operated in the attenuated total reflection (ATR), external reflection, or diffuse reflection modes are available on the market. In contrast, a broad range of miniaturized Raman systems, with different excitation sources and specific operational features, is offered. Most of the handheld Raman spectrometers are operated with a 785 nm excitation source, with the inherent problem of disturbing fluorescence and a high-wavenumber cut-off at about 2200 cm

. The problem of fluorescence can be either tackled by 1064 nm excitation or by sequentially shifted excitation spectroscopy (8). Bruker also managed to extend the wavenumber range to 3200 cm

 by dual-wavelength excitation (9). A very powerful countermeasure to compensate the error source of a sharp focus in connection with a heterogeneous sample is the so-called orbital raster scan (ORS) (10) that is integrated in the Metrohm handheld Raman system. In 2015, Thermo Fisher Scientific merged their miniaturized FT-IR and Raman instruments into a new handheld FT-IR–Raman spectrometer, thereby launching vibrational spectroscopy into a new era of flexibility.

	Contrary to Raman and MIR spectroscopy, miniaturization has been much further driven for NIR spectrometers. The recent progress in miniaturization has taken advantage of new micro-technologies such as micro-electro-mechanical systems (MEMS), micro-opto-electro-mechanical systems (MOEMS), micro-mirror arrays, and linear variable filters (LVFs). These have led to a drastic reduction of spectrometer size (the weight of the spectrometers discussed in this communication varies between 100-200 g), while allowing good performance due to the high-precision implementation of important elements in the final device (11). High-volume manufacturability will further reduce costs, and thereby contribute towards a broader dissemination of such instruments for everyday-life applications. In what follows, the specific instrumental features of the four handheld NIR spectrometers used in our laboratory will be outlined.

	Based on the type of detector, handheld NIR spectrometers can be classified in the two categories of array-detector and single-detector instruments (11). One of the first commercial handheld NIR spectrometers (VIAVI MicroNIR 1700 [formerly JDSU], Santa Rosa, CA) has an array detector covering the wavelength range from 908-1676 nm, and uses a monochromator based on an LVF. It has so far been used for a multiplicity of applications, ranging from authentication of seafood and determination of food nutrients, the analysis of hydrocarbon contaminants in soil, and authentication and quantitative determination of pharmaceutical drugs (12-15). However, compared to an array detector, the price for a single detector is much lower, and, to further reduce the hardware costs, new developments focus on systems with single detectors. For example, the DLP NIRscan Nano EVM (Dallas, TX, USA), is based on Texas Instruments´s digital micro-mirror device (DMD), in combination with a grating and a single-element detector, covering the wavelength range from 900-1701 nm. Very recently, a MEMS-based FT-NIR instrument, that contains a single-chip Michelson interferometer with a monolithic opto-electro-mechanical structure, has been introduced by Si-Ware Systems (Cairo, Egypt). Contrary to most of the other handheld spectrometers, this instrument can scan FT-NIR spectra over the extended range from 1298 nm to 2606 nm. Finally, Spectral Engines (Helsinki, Finland) has developed miniaturized NIR spectrometers that are based on a tunable Fabry-Perot interferometer. In order to cover the NIR wavelength region 1350-2450 nm, however, four spectrometers are required.

	The schematic principles of the different monochromator designs of the described NIR spectrometers are summarized in Figure 1.


	Figure 1: The optical schemes of handheld NIR spectrometers, based on different monochromator principles: (a) VIAVI MicroNIR 1700, linear variable filter; (b) DLP NIRscan Nano EVM with Texas Instruments digital micromirror device (DMD); (c) Si-Ware Systems, MEMS-based FT-NIR spectrometer; (d) Spectral Engines NIR spectrometer with tunable Fabry-Perot interferometer).

	To demonstrate the potential of handheld vibrational spectrometers, only a few, selected application examples will be presented here. Note that Raman spectroscopy has also proved valuable for the characterization of inorganic materials. The rapid noninvasive determination of the different crystalline forms of TiO

 (rutile and anatase), or the discrimination of a genuine diamond from an imitation, are just typical examples.

	In a feasibility study, the change of the ionic structure of sulfuric acid as a function of concentration has been demonstrated by Raman spectra recorded with a handheld instrument (Rigaku Raman Technologies). The spectra measured consisting of 96% (concentrated), 80%, and 20% sulfuric acid are shown in Figure 2. The differences in ionization are an important issue regarding non toxicity of diluted sulfuric acid for cleaning operations of aluminum surfaces.


	Figure 2: Raman spectra of sulfuric acid at different concentration levels, measured with a handheld instrument.

	The application of handheld FT-IR spectroscopy in the ATR mode is recommended primarily for organic materials (preferentially liquids) that provide good contact with the diamond (or ZnSe) reflection element. Despite its rather difficult sample handling, an interesting practical application is the quality control of bitumen. For road construction work, bitumen is modified with different polymers in order to adjust its rheological and adhesion properties (polymer modified bitumen, PmB). The rapid quantitative analysis of this admixture is an important quality control issue, and the question that arises is whether handheld FT-IR–ATR spectroscopy is applicable for this purpose, and could, at the end of the day, be used for "on the road" analysis. In an extensive feasibility study, FT-IR–ATR spectra of bitumen samples, modified by different admixtures of polypropylene maleic anhydride (PPMA), were measured with the handheld Thermo Fisher Scientific FT-IR spectrometer (accumulating 60 scans at 4 cm

 spectral resolution). Based on these spectra with specific wavenumber selection (3250-2500 cm

), and standard normal variate (SNV) transformation (Figure 3a), a cross-validated partial least squares (PLS) calibration model was developed (Figure 3b). Test samples that were not contained in the calibration model were successfully predicted regarding their PPMA content.

	Due to the advanced level of miniaturization, by far the most flexible technique for on site and in-the-field measurements is NIR spectroscopy. Although this technique can also be applied for a broad range of industrial material quality control applications (15-18), the present communication is targeted at practical everyday life applications in order to attract the attention of a prospective non-expert user community. In these days, fraud and adulteration are so widely spread that the progress in miniaturization and the increasing affordability of handheld NIR spectrometers makes them an attractive tool to efficiently fight the aforementioned evils. The following application examples shall demonstrate the potential of handheld NIR spectroscopy, not only for this purpose, but also for exemplary everyday life applications.


	Figure 3: Calibration spectra of (a) polymer-modified (PPMA) bitumen samples, and (b) cross-validated PLS calibration of the polymer admixture developed thereof. 

	In China, the stem of the Dendrobium orchid is processed into a fengdou that is considered not only a convenient dosage form of a valuable health-care food, but also of a Chinese traditional medicine (TCM) with efficacy in liver protection, treatment of pharyngitis, and many other diseases (19-20). Because fengdous processed from 

 (DOK) not only have high medicinal value, but are also in short supply, and, consequently, are very expensive. Therefore, it would be desirable to discriminate them from fengdous based on 

(DDP) with lower efficacy and correspondingly much lower (1/4-1/5) price. However, this is not possible by visual inspection only (Figure 4a) Thus, this case is an example of high public interest. The handheld DLP NIRscan Nano EVM spectrometer, in combination with a PLSDA (partial least squares discriminant analysis) evaluation method, was used to rapidly identify fengdous processed either from DOK or DDP. For this purpose, a total of 468 fengdou samples based on DOK (288) and DDP (180), were collected from Luosiwan (Yunnan, China), and the calibration and prediction set were randomly distributed at a ratio of 2:1. NIR spectra were collected for each sample with the DLP NIRScan Nano EVM spectrometer by accumulating 32 scans in approximately seven seconds. After pretreatment by first derivative, SNV, and mean centering, the identification model was developed by PLSDA, and the competitive adaptive reweighted sampling (CARS) wavelength optimization algorithm was used to filter out the wavelengths with high information content to improve the prediction accuracy of the model (Figure 4). The results showed that, for the calibration, cross validation and prediction sets, the accuracy is 93.9%, 89.6%, and 84.1%, respectively. This approach clearly demonstrates that the DLP NIRScanNano EVM spectrometer, combined with CARS-PLSDA evaluation, can be successfully used for the rapid discrimination of fengdous produced from DOK and DDP (Figure 4).


	Figure 4: Identification of the species of dendrobium with a handheld NIR spectrometer: (a) samples of DOK and DDP, (b) the variables selected by CARS, and (c) prediction of DOK by calibration model.

	The list of adulterations and their successful detection by the use of handheld NIR spectroscopy is not limited to pharmaceutical formulations (21). More exotic cases such as the offer for sale of silk carpets (imitated by carpets knotted from mercerized cotton) (22), amber gem stones (counterfeited by colored poly(methylmethacrylate)(PMMA) with an artificially embedded insect), or ivory carvings (made from regular bone) frequently occur in tourist centers all over the world.

	As a typical household example, the use of handheld NIR spectroscopy is demonstrated with reference to the identification of an unknown contamination in a cotton textile. In Figure 5 (top), the diffuse-reflection NIR spectra (measured with a Si-Ware Systems FT-NIR spectrometer) from both a clean and a blood-contaminated cotton textile are shown. Absorbance subtraction of these spectra (Figure 5, bottom) can readily highlight the protein specific N-H overtone and amide I and amide II combination bands in the 6600 cm

 wavenumber regions (gray shaded areas), respectively. This information will then allow selection of the specific parameters of the washing machine for the perfect removal of this stain.


	Figure 5: Identification of a textile contamination by handheld NIR spectroscopy: Isolation of blood-specific absorption bands (gray shaded areas) by absorbance subtraction of the FT-NIR spectra of a blood-contaminated and a clean cotton textile. 

	Finally, an application example from food testing is shortly outlined. Over the last few years, the health awareness of people has grown strongly, and the control of nutritional parameters of everyday diet is just one aspect of this issue. At this point, however, a word of caution is necessary. Contrary to the approach of direct-to-consumer companies that video-advertise their "food scanners" with "cloud evaluation of big data" and (in our opinion) exaggerated claims, predictions of nutritional parameters with an acceptable accuracy can only be achieved when calibrations based on reliable spectral data are developed.

	As a starting project in this field, the analysis of the nutritional parameters of pasta sauce mixtures has been chosen. For this purpose, more than 700 plates containing five different combinations of a certain pasta sauce blend (ranging from 0% to 100% sauce addition) were investigated. Each pasta sauce mixture was prepared "ready to eat" on a plate, and NIR spectra were recorded in five different positions (to compensate for inevitable heterogeneities) with a handheld NIR spectrometer (Viavi MicroNIR 1700). Prior to the development of the calibration models, the five spectra were averaged and pretreated by extended multiplicative scatter correction (EMSC) (Figure 6a), and, based on the known nutritional parameters of the utilized pastas and sauce ingredients, calibration models were developed for fat, energy, protein, carbohydrates, sugar, and fiber (Figure 6b–d).


	Figure 6: NIR spectra (after EMSC pretreatment) measured with a handheld spectrometer of 700+ plates with pasta and sauce mixtures for calibration development of nutritional parameters: (a) averaged spectra of five replicate measurements, (b) actual vs. predicted plot for fat, (c) actual vs. predicted plot for energy, and (d), actual vs. predicted plot for fiber. The corresponding calibration results for protein, carbohydrates, and sugar are not shown here.

	Contrary to what some of the above mentioned direct-to-consumer companies claim, acceptable calibrations could, at best, be achieved for fat (Figure 6b), energy (Figure 6c), protein, and, with limitations, carbohydrates. Due to the similar structural characteristics of carbohydrates, sugar, and fiber (Figure 6d), no reliable calibrations were obtained for the last two parameters.

	Generally, handheld instruments have launched vibrational spectroscopy into a new era of in-the-field and on-site analysis. State-of-the-art handheld Raman, MIR and NIR spectrometers demonstrate good performance for a multiplicity of case studies. However, miniaturization of NIR spectrometers has reached a much higher level compared to Raman and MIR spectrometers. Furthermore, high volume manufacturability has considerably reduced costs for handheld NIR spectrometers, and will, hopefully, contribute towards a broader dissemination of such instruments for everyday life applications by a non-expert user community.

	The authors gratefully acknowledge instrumental support by Si-Ware Systems (Egypt), VIAVI (USA), and Spectral Engines (Finland). Furthermore, experimental and organizational help by Frank Pfeifer (University of Duisburg-Essen, Germany) and Farzad Ehsani (Vitruvi LLC, USA) are acknowledged.

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	(3) A. Guillemain, K. Degardin, and Y. Roggo, 

	(4) J.M. Soriano-Disla, L.J. Janik, and M.J. McLaughlin, 

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	(9) J.B. Cooper, S. Marshall, R. Jones, M. Abdelkader, and K.L. Wise, 

https://www.news-medical.net/whitepaper/20151214/Orbital-Raster-Scan-Technique-Helps-Improve-Performance-of-Mobile-Raman-Systems-for-Pharmaceutical-Analysis.aspx

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Zhonghua Renmin Gongheguo wei sheng bu yao dian wei yuan hui (Chemical Industry Press, Beijing, 2000), pp. 85-87.

	(21) M. Alcalà, M. Blanco, D. Moyano, N. Broad, N. O'Brien, D. Friedrich, F. Pfeifer, and H.W. Siesler, 

 is with the School of Biotechnology at the Jiangsu University of Science and Technology in Zhenjiang, China. 

 is with the Department of Chemistry at the University of Duisburg-Essen in Essen, Germany. Direct correspondence to: 

Handheld Raman, mid-infrared and near infrared spectrometers have launched vibrational spectroscopy into a new era of in-the-field and on-site analysis, as the application examples discussed here demonstrate. We assess the technological developments that have led to this progress.

Handheld Raman, Mid-Infrared and Near Infrared Spectrometers: State-of-the-Art Instrumentation and Useful Applications

Raman spectroscopy is an optical spectroscopic method that enables characteristic identification of chemical substances. Spatially offset Raman spectroscopy (SORS), a novel variant of Raman spectroscopy, enables new capabilities in the rapid identification of materials concealed by a wide variety of nonmetallic sealed containers, such as colored and opaque plastics, paper, cardstock, sacks, fabric, and glass, which are often challenging for conventional backscattered Raman spectroscopy. Handheld spectroscopy based on SORS has been developed for detection of several materials and barrier types, including explosives, chemical agents, precursors, and hazardous narcotics (such as fentanyl). SORS has the potential to improve the safety, efficiency, and critical decision-making in incident management and response. The operator is able to obtain a positive identification of a potentially hazardous material without opening or disturbing the container-to gain access to take a sample-thus improving safety. The technique is fast and simple, therefore protective suit and breathing gear time is used more efficiently when operating in a "hot zone".

	There is a continuing evolution of new threats and chemical hazards throughout the world. Therefore, a method that can positively identify these threats, while being user friendly, highly specific, fast, and safe, is required. Raman spectroscopy is an accurate, specific, and rapid technique for identification, which has led to an exponential increase in portable and handheld Raman spectrometers. At present, there are over 30 vendors supplying portable Raman equipment to a range of sectors, such as first responders, hazmat operatives, military, explosive ordnance disposal, police, investigation teams, and customs. Differentiation between competing systems is generally limited to laser excitation wavelength (longer wavelengths, less interference from fluorescence), matching algorithms, software interface, and library content.

	Conventional Raman has many advantages, one being the ability to identify contents within a clear glass vial or thin plastic bag. However, an opaque plastic or colored glass container results in a large fluorescent background, and can attenuate the beam. Spatially offset Raman spectroscopy (SORS) is a method developed at the Rutherford Appleton Laboratory, UK (1) that provides the ability to identify the contents of a truly opaque or complex container, without opening it to gain access or to take an aliquot of the sample (Figure 2). This capability provides several benefits, including safety (not disturbing the object or releasing materials into the environment), efficiency (time spent in personal protective equipment [PPE] is minimized), decision making (mission-critical data are obtained earlier in an operation), and the preservation of evidence and intelligence.


	Figure 1: Schematic representation of SORS geometry showing the "zero offset" laser excitation relative to the detector, and the "offset" laser excitation, separated by a distance ΔS. Laser excitation generates scattered photons which have diffused through the container and contents. There is a change in Raman contribution from the container and its contents depending on offset measurement.

	Raman spectroscopy relies on the inelastic (Raman) scattering of monochromatic light. Laser light excites molecular vibrations, resulting in a change in the energy of the scattered light shifted in wavelength from the excitation source. Subsequently, this produces a highly unique and characteristic spectrum of the molecules being analyzed, enabling Raman spectroscopy to be used to positively identify specific molecules of interest. The principle of SORS is based on taking multiple Raman measurements. First, a "zero offset" measurement is made; the detector and laser excitation is positioned in the same configuration as a conventional backscattered Raman system. This measurement typically consists of the container surface. Second, one or more measurements are then made with either the detector or the laser offset by a distance of ΔS (shown in Figure 1). This novel configuration allows for the detection of photons that have diffused further into the container. Therefore, the spectra obtained will consist of both the container and its contents. As the offset is increased, the container surface component of the signal depletes while the contents signal is increased. Acquiring multiple measurements results in varying signal contributions from both the container and its contents, making it possible to identify and separate the signal into its respective components. The process is fully automated, and does not require any additional intervention of the operator or prior knowledge of the barrier.


	Figure 2: (Top): SORS extends through barrier giving the capability to work with a wide range of nonmetallic containers. (Bottom): Conventional Raman systems require line-of-sight and thus work with clear plastic bags and vials, and some translucent packaging.

	A handheld SORS system (Resolve) was supplied by Agilent Technologies LDA UK limited, formerly Cobalt Light Systems (Oxford, UK), and used without any modification.

	A major advantage of SORS technology is the ability to detect and positively identify the contents of a container through the barrier. In many parts of the world, improvised explosive devices (IEDs) are constructed from readily available materials, meaning explosive materials being transported may look like common benign household containers. A good example of this is yellow palm oil containers (YPOC), commonly used in many parts of Africa and Asia, used to make fuel-oxidizer type IEDs. The yellow, or sometimes blue, containers are typically made from molded high-density polyethylene (HDPE). This material attenuates the beam, adds a fluorescent background, and produces a strong Raman signal. Conventional Raman spectroscopy would not be able to scan effectively through these barriers. In the example shown in Figure 3, SORS successfully identified the mixture of ammonium nitrate and sugar from within the container, through the barrier wall, in < 2 min. Even more difficult is the ability to distinguish the "benign" version of this container; that is a YPOC which contains unprocessed (bright orange colored) palm oil. Difficulty arises from the fact that the contents are fluorescent, and the spectrum of many oil-like substances is very similar to HDPE making the processing challenge greater.


	Figure 3: Example scans of West African yellow palm-oil containers with ammonium nitrate and palm oil contents, respectively. Items tagged as priorities in the software are displayed as red. A warning message (such as, "explosive precursor") is configurable by an authorized user.

	SORS can also operate with a direct line of sight to a sample, such as exposed white powder or liquid spillage with a "surface scan" mode. Despite conventional Raman being a very effective technique, in practice, its use is often limited by operational procedures. The unique optical geometry used in this manifestation of SORS means the laser spot is not delivered coaxially with the collection path. This, in turn, results in a much larger laser spot, and, consequently, the power density of the laser on the sample is reduced (by orders of magnitude) compared to the tightly focused beam typical of a conventional Raman system. This is highly advantageous when scanning for sensitive explosives. Together with shorter exposure times (from high optical efficiency), there is a significantly reduced risk of detonation or deflagration, which improves user safety when working with sensitive explosives. Figure 4 shows an example of Raman spectra from nine primary and sensitive explosives. Sensitive explosives (approximately 2-5 g in each case) were scanned directly in point-and-shoot mode, or contained in a glass vial. Maximum laser power was used in each case (475 mW), and a timer delay was used to allow the operator to retreat to a safe distance.


	Figure 4: Right: processed SORS result and material identification. Left: example of silver fulminate scans. Scans are normalized in all cases.

	The setup in Figure 4 was chosen to increase safety by not containing the sample, and to maximize laser power by not putting an attenuating barrier between the system and the explosive. Total measurement times in these modes were < 1 min at 100% laser power, and the scan timer delay function was used. Experiments were repeated 10 times and on several batches of the same material, with 100% detection and correct identification achieved. The high signal-to-noise ratio means that these sensitive materials can be accurately identified from the libraries of many thousands of chemicals.

	Silver fulminate is a white or off-white powder which is highly sensitive to percussion or energy input and is known to detonate instantly when scanned using conventional Raman. Using SORS, the material could be identified in all scans with 100% accuracy and no deflagration or detonation. Care and caution should always be used when scanning sensitive explosive materials.

Narcotics and New Psychoactive Substances

	The rise in the prevalence of hazardous narcotics and new psychoactive substances (NPS) presents a major challenge to law enforcement professionals. These materials vary in composition and color from pure white powders to raw and unrefined colored and brown samples. Many narcotics, and in particular NPS, are hazardous by inhalation, ingestion, or eye or skin contact. There are several worldwide reports that describe accidental exposure to officers, resulting in serious medical consequences. An example is fentanyl; this opioid is reportedly 10-1000 times more potent than heroin, with fatal doses comparable to a few grains of sugar. This material presents a high risk in its pure form (as it is smuggled or transported). Given the high potency of these substances, the risk remains after they are cut down into the multitude of street forms and products. This means there is a risk of exposure in situations where full protective equipment is not worn. Therefore, a technique that allows materials to be scanned through a variety of wrapping and packaging minimizes the risk of exposure to the operator.

	Combining SORS, high quality data with a library that is continuously updated provides a powerful tool in detection of NPS. Many narcotics, especially those that are unrefined or cut with excipients, are fluorescent. Using a 830 nm laser excitation wavelength significantly reduces fluorescence emission in comparison to a commonly used 785 nm laser excitation. Excitation at 830 nm also produces significantly more Raman scattering than a 1064 nm system (λ

 law), and allows the use of more sensitive detectors. Handheld SORS can identify narcotics and new psychoactive substances concealed within typical packaging, such as white and brown paper (fluorescent), plastic wrapping (Raman background), and green, brown, blue, or clear thick glass bottles (fluorescence).

	The ability of handheld SORS to identify hazardous narcotics and NPS concealed behind barriers such as post packs, brown paper wrapping, cardstock, opaque polyethylene, opaque polypropylene, opaque PVC containers, and rubber or nitrile rubber has been tested at expert laboratories in the United Kingdom, United States, and China.

	The SORS process can still operate successfully through multiple layers of differing composition without any prior knowledge. However, in cases such as these, the user must ensure that the nose cone is placed directly opposite and pressed towards the material to be scanned. The material needs to be within a maximum distance of ~5 mm of the nose cone for the SORS optics to function correctly. In cases where this distance is too great, the system will often report a plastic or no match.

	SORS works very well with narcotics in concentrations from 100% down to ~10% w/w, such as those typical of smuggling operations and the raw material used in street product manufacturing. However, the concentrations of active component in some street samples is very low. This is especially true of fentanyl samples. In some instances, only trace (a few grains) of sample are readily available. This is not enough material to obtain a high quality Raman signal; therefore the sample must be concentrated, or the Raman scattering has to be enhanced or intensified. One method which enhances the Raman effect is surface enhanced Raman scattering (SERS). Metallic surfaces that have one dimension in the range of 1-100 nm exhibit interesting optical properties, which the corresponding bulk material does not. In the presence of metallic nanoparticles, typically gold or silver, a significant increase in sensitivity from this optical effect is achieved, an enhancement factor of 10

 has been reported, indicating that single molecule detection is possible (2). Not only do metallic nanoparticles enhance the Raman effect, the nanoparticle also quenches the competing fluorescence process (unrefined narcotics are typically fluorescent).

	Using handheld SORS in connection with a trace detection kit also helps the user. For 2-3 mg of a mixture of 90:10 heroin to fentanyl (mixed with excipient this is typical of a street sample) was added to such a kit, and analysis was performed using 100% laser power (Figure 5). The presence of both fentanyl citrate and heroin was successfully identified. This amount of fentanyl material mixed with heroin would be extremely hard to detect in these quantities using conventional Raman. This trace kit test enables increased sensitivity of materials, hence detection of materials at lower concentrations. This is a significant step towards preventing and detecting chemical hazards and threats.


	Figure 5: Results using handheld SORS showing the positive identification of fentanyl citrate and heroin found in a 2-3 mg sample.

	A portable SORS system has been developed and is successfully deployed and utilized by military, first responders, customs, and law enforcement operators. Detection could be achieved in almost 100% of cases where the sample was concealed by a thick, opaque, or colored barrier with maximum scan times < 2 min. The examples presented represent a step change in the capability of portable Raman spectroscopy as these samples were previously inaccessible to the technique. A portable SORS system enables better information to be obtained earlier in an operation, and better decisions made without disturbing potentially hazardous materials (such as explosives or fentanyls). Hazardous materials and narcotics can be scanned from within the container, reducing the risk of accidental exposure to the operator.

	(1) C. Eliasson, N.A. Macleod, and P. Matousek, 

	(2) R Stokes, A Macaskill, P.J. Lundahl, W.E. Smith, K Faulds, and D Graham, 

Robert J Stokes, Ana Blanco, Rachel McGee,

 are with Agilent Technologies LDA UK Limited, in Abingdon, United Kingdom. Direct correspondence to: 

Portable spatially offset Raman spectroscopy (SORS) enables rapid identification of materials concealed by a wide variety of nonmetallic sealed containers. This technique has been successfully used by the military, first responders, and customs and law enforcement operators in the detection of explosives, chemical agents, precursors and hazardous narcotics, thus enabling the acquisition of better information without disturbing the materials in question.

Special Issues-11-02-2018

Spectroscopy November 2018 Advances in Portable and Handheld Spectroscopy Supplement PDF

Portable, High-Efficiency Transmission Raman Spectroscopy for At-Line Content Uniformity Testing of Pharmaceutical Tablets

Handheld Raman, Mid-Infrared and Near Infrared Spectrometers: State-of-the-Art Instrumentation and Useful Applications

Portable Spatially Offset Raman Spectroscopy for Rapid Hazardous Materials Detection Within Sealed Containers