Analytical tools to pin down adulteration in spices

High-priced commodities such as spices and herbs are persistently susceptible to adulteration, which may have a detrimental impact on their overall quality. The most common cause of spice adulteration is an increased demand or shortage of supply caused by a number of reasons, including natural disasters. Recently, there has been an increasing trend in spice fraud that appears to be driven by economic motives in order to obtain higher profit margins. The adulteration of spices can be unintentional, due to carelessness or deliberate mixing with other substances to increase the weight or volume.

Spice adulteration is a highly dynamic fraudulent practice that exploits a wide range of adulterants, including inorganic, synthetic organic, and plant-derived components. Universal adulterants that are cheap and readily available, termed bulking substances or fillers, have been used to adulterate various products in several incidents. Bulk substances include starches, flour, bran, sawdust, grasses, and straw, for example. These substances may be both harmless and safe for ingestion or may pose a significant health threat to consumers. People with food allergies have experienced adverse reactions to the bulking material adulterant added to spices, which sometimes contains undeclared allergens. In many instances, synthetic dyes, such as Sudan dyes I–IV, a class of carcinogenic azo dyes that are not approved for human consumption, have been noted as adulterants and have triggered numerous recalls. In other cases, spices have been found to be adulterated with toxic heavy metals.

Adulteration of spices, whether economically motivated or not, may have serious implications on public health and industry. The increasing incidence of food adulteration and the occurrence of several massive product recalls over a short period of time have resulted in untoward consequences that affect producers, suppliers, retailers, and consumers. Keeping track of the quality and quantity of raw material is a challenge for manufacturers of spices. Spices are often harvested from small farms, without formal supply chains. In such conditions, it becomes difficult to trace the source of adulteration in spices.

Researchers have developed a variety of methods to detect food fraud and identify adulterants in a quick and economical method, in order to guarantee consumer protection and food safety. These techniques, including chemometric analyses combined with instrumental techniques, are used to generate accurate and consistent results relating to food fraud. Several classical and novel analytical techniques have been identified as being effective in combating the ongoing adulteration issues with spices.

The organoleptic method relies on unaided senses of sight, smell, or taste to perceive the colour, aroma, or flavour of spices. This sensory method provides an immediate assessment of the sample and is often used in conjunction with microscopy and macroscopy as well as other analytical techniques to ensure accuracy and reliability. Historically, microscopy has been used as a simple and rapid method for preliminary quality control of spices by identifying the tissues with standard histological characteristics. A plant or plant part (e.g., leaf, flower, fruit, seed, stem, bark, root, and rhizome) can be identified macroscopically by characterising its morphology. Because it is based on a variety of morphological characteristics, morphological identification is still considered accurate and reliable. Taxonomists can detect adulteration or contamination by examining the key anatomical aspects of botanicals. However, macroscopic studies are ineffective for authenticating ground or processed spices.

Spectroscopic techniques employ light to interact with matter and thus probe certain features of a sample to learn about its consistency or structure.

UV-spectrophotometry: A UV-Vis spectrophotometer is used to quantify food colourants, like turmeric and paprika. The colourants are added to spices to make them look more attractive. The quantification of colourants in spices can be used as an indicator of adulteration. The colourants can be quantified by the method of calibration curve with samples of coloured spices.

IR spectroscopy: Infra red spectroscopy, including FT-NIR and mid infrared Fourier transform, is simple, rapid, noninvasive, and environmentally friendly in detecting and quantifying adulterants in botanical ingredients including spices. Plant tissues and plant extracts can be measured for IR spectroscopy directly in order to gather characteristic absorption spectra. The spectral data provide information regarding the chemical structures of specific compounds (primary and secondary plant metabolites) that can be used to develop spectroscopic fingerprints. Chemometric methods are used to distinguish between different species and different chemotypes within the same species using IR spectroscopy. IR spectroscopic methods are also used in the herbal trade to monitor the processing stages, starting with the raw materials and finishing with the finished products.

Near-infrared (NIR) spectroscopy: NIR is a technique used extensively in the food and dairy industries. It is a low-cost alternative to HPLC, with comparable accuracy. NIR can be used to quantify major spices like black pepper, turmeric, and cinnamon. A few grams of ground spice can be used for NIR analysis. The spectra of ground spices are different than that of the same spices in their natural form. The ground spices can be classified as whole, broken, or powdered. A calibration curve can be generated for each type of ground spice. An additional advantage of using NIR spectroscopy is related to its low substance absorptivity that enables direct measurement of transmission through intact materials without any sample preparation.

Raman spectroscopy: Raman spectroscopy is thought to be more advantageous than NIR and Mid-IR because it is not influenced by water or inorganic materials. Hence, the measurements can be carried out directly on the packaging or through glass-walled containers. The significant disadvantage of Raman and other low-resolution IR spectroscopies is the requirement for a large sample data set and coordinated chemometric tools in order to validate the data generated by this method. In contrast to IR, Raman spectroscopy is less sensitive in identifying contaminated or adulterated samples at low concentrations. Nevertheless, it has been reported that both methods might provide supplementary data by using data analysis and determine sample authenticity.

Mass spectroscopy (MS): MS is one of the most widely utilised and trustworthy methodologies for detecting food adulterants. MS is used by quality assurance professionals for detecting food adulterants and impurities. Its sensitivity and selectivity make it a powerful confirmatory method. Mass spectrum-based analyses may be used for either targeted or non-targeted purposes. MS analysis is usually performed by a mass spectrometer that is connected to a separation machine like HPLC or UHPLC (GC for more volatile compounds). A small sample (1-2 g) of each spice can be used for HPLC and MS analysis. To interpret the acquired data, chemometric data analyses are utilised.

Mass spectrometry is limited by the requirement for sample preparation, which requires the gas or liquid state to be present as ionisation sources. Furthermore, mass spectrometric equipment is costly and must be operated by highly skilled individuals.

LC-MS and GC-MS are two of the most frequently used mass spectrometric techniques in food analysis. GC-MS is an important technique for detecting food adulteration. It has been used largely to authenticate aromatic herbs and spices and to detect adulterants from their essential oils. However, some people believe that GC-MS methodology is unsuitable for examining adulterated spices and herbs, because it is designed to assess only the more volatile components found in essential oils or derivatised fatty acids and sterols. Ambient mass spectrometry has recently been described as an advanced method for detecting and quantifying spice adulterants. In ambient MS, samples are analysed in their original native states whether they are solid, liquid, or gaseous, and no sample preparation is required. Because of this advantage, ambient MS is preferred to other comparable technologies. Some popular techniques of ambient MS are atmospheric solid- analysis probe-MS (ASAP-MS) and direct analysis in real-time ionisation–time-of-flight–MS (DART- ToF-MS).

Nuclear magnetic resonance (NMR) spectroscopy: NMR spectroscopy is a prominent fingerprinting technique used for detecting food frauds. Unlike IR and Raman spectroscopy, NMR spectroscopy is one of the techniques that has the ability to record signals from all chemical components in a sample and detect unrelated substances that are not part of the authentic sample. The accurate quantification of several substances using one reference standard is possible using proton NMR (1H NMR) because it has the ability to distinguish between chemically similar compounds with minor structural variations. But the high cost, operational difficulty in upkeep of the instrument and need for specialised operators limit its popularity.

DNA sequencing is used to determine the genetic makeup of an organism. It is used to determine the identity of spices, their adulterants and even their geographical origin. As the cost of DNA sequencing has come down drastically in recent years, this technology is useful for manufacturers, to determine the identity of spices, and for regulatory bodies, to check the authenticity of spices.

DNA barcoding has become a popular method for identifying botanicals, differentiating closely connected plant species, and ensuring high quality. A few specific plant sequences, which are unique to a plant species, are compared to a standard reference material using these DNA-based strategies. The approach is able to distinguish between species, cultivars, and varieties, but not between organs or tissues of the same plant. DNA markers are not impacted by plant development stages, weather conditions, or location, unlike chemical markers, which can be used to verify a plant of any age or origin. Very recently Thermo Fisher Scientific has come out with a ready-to-use Thermo ScientificTM NGS Food Authenticity Workflow – a commercial DNA metabarcoding approach to identify food plant species in herbs and spices, not only when tested on pure samples, but also in mixtures down to 1% (w/w). It is easy to use, has high sensitivity and allows the detection of undeclared species in commercial samples.

On the other hand, DNA barcoding has some significant drawbacks as well. One of the biggest drawbacks is that some samples may not be able to be used for DNA extraction or amplification. When collecting plant tissue, DNA should be extracted as quickly as possible to avoid damage to the DNA during storage. Certain secondary metabolites, including polysaccharides, polyphenols, and terpenoid lactones, affect DNA extraction and may interfere with or inhibit PCR amplification. Furthermore, the DNA barcoding methodologies currently in use are not capable of providing reliable and accurate quantitative assessments of the relative abundance of plants in a mixture of botanical ingredients.

Conclusion

Adulteration in spices can have serious impacts on human health and the economy. To counteract the adulteration of spices and herbs, a variety of techniques across many different technologies have been developed. These techniques are expected to detect adulterants and measure their concentrations in a variety of matrixes, including spices, herbs, and plant materials. Chromatographic techniques such as HPLC, HPLC-MS, DART- ToF-MS, and GC-MS are currently used to detect adulterants and establish their concentrations. Spectroscopic techniques include FT-IR, NIR, Raman, and NMR spectroscopy. These visual and sensorial assessments of plant materials in addition to microscopy are still rapid and economical to find gross adulterations in spices and herbs. DNA-based technology has grown as an advantageous authentication device, primarily for recognising closely related species and cultivars. At the same time, several non-invasive methods are being tested across the globe for detecting adulteration in spices. Recently a GC-based automated chemical analysis system has been developed for identifying terpenes in spices and other botanicals by enhancing a proven bacterial identification system. In conclusion, spectroscopic techniques show a great efficiency to authenticate spices. But those involved in the food industry prefer portable devices due to their manageability and low cost.

Additional Reading

• Faith Ndlovu P, Samukelo Magwaza L, Zeray Tesfay S, Ramaesele Mphahlele R 2022. Destructive and rapid non-invasive methods used to detect adulteration of dried powdered horticultural products: A review. Food Res Int. 2022 Jul;157:111198. doi:10.1016/j.foodres.2022.111198

• Oliveira MM, Cruz-Tirado JP, Barbin DF. Nontargeted analytical methods as a powerful tool for the authentication of spices and herbs: A review. Compr Rev Food Sci Food Saf. 2019 May;18(3):670-689. doi:10.1111/1541-4337.12436

• Osman AG, Raman V, Haider S, Ali Z, Chittiboyina AG, Khan IA. Overview of analytical tools for the identification of adulterants in commonly traded herbs and spices. J AOAC Int. 2019 Mar 1;102(2):376-385. doi:10.5740/jaoacint.18-0389

• Xu Y, Zhang J, Wang Y 2022. Recent trends of multi-source and non-destructive information for quality authentication of herbs and spices. Food Chem. 2022 Aug 12;398:133939. doi:10.1016/j.foodchem.2022.133939

[Published in Spice India September 2022]