Authentication of saffron using 60 MHz 1H NMR spectroscopy

60 MHz proton NMR spectroscopy was used to analyse extracts from saffron spice and a range of potential adulterants and mixtures. Using a simple extraction procedure, good quality spectra were obtained which contain peaks from the characteristic metabolites picrocrocin and crocins, fatty acids and kaempferol. The spectra of samples from trusted suppliers were used to train one-class classification models by SIMCA, nearest neighbour and isolation forest methods. Applying these to spectra of saffron samples purchased from the online marketplace, it was found that 7 out of 33 samples were highly anomalous. From comparison with the spectra of known mixtures and confirmatory spectral analysis using 600 MHz NMR, it is probable that these contain considerable amounts of undisclosed foreign matter

Quantitative authenticity testing of buffalo mozzarella via alpha(s1)-Casein using multiple reaction monitoring mass spectrometry

We address the detection and quantitation of bovine milk in 'buffalo' mozzarella cheese using multiple reaction monitoring (MRM) mass spectrometry (MS). Focussing on the abundant protein alpha(s1)-casein, present in both species but with 10 amino acid sequence differences, we extract a list of marker peptides specific to each species. 'Identical' peptides, exactly the same in both species, are used for relative quantitation of alpha(s1)-casein in each milk type, whereas 'similar' peptides, present in both species but differing typically by one amino acid, are used to demonstrate relative quantitation in binary cheese mixtures. In addition, we report a pilot survey of UK supermarket and restaurant products labelled as 'buffalo mozzarella', finding that 2/3 of restaurant meals and supermarket pizzas are either mislabelled or adulterated

Species Determination and Quantitation in Mixtures Using MRM Mass Spectrometry of Peptides Applied to Meat Authentication

We describe a simple protocol for identifying and quantifying the two components in binary mixtures of species possessing one or more similar proteins. Central to the method is the identification of 'corresponding proteins' in the species of interest, in other words proteins that are nominally the same but possess species-specific sequence differences. When subject to proteolysis, corresponding proteins will give rise to some peptides which are likewise similar but with species-specific variants. These are 'corresponding peptides'. Species-specific peptides can be used as markers for species determination, while pairs of corresponding peptides permit relative quantitation of two species in a mixture. The peptides are detected using multiple reaction monitoring (MRM) mass spectrometry, a highly specific technique that enables peptide-based species determination even in complex systems. In addition, the ratio of MRM peak areas deriving from corresponding peptides supports relative quantitation. Since corresponding proteins and peptides will, in the main, behave similarly in both processing and in experimental extraction and sample preparation, the relative quantitation should remain comparatively robust. In addition, this approach does not need the standards and calibrations required by absolute quantitation methods. The protocol is described in the context of red meats, which have convenient corresponding proteins in the form of their respective myoglobins. This application is relevant to food fraud detection: the method can detect 1% weight for weight of horse meat in beef. The corresponding protein, corresponding peptide (CPCP) relative quantitation using MRM peak area ratios gives good estimates of the weight for weight composition of a horse plus beef mixture

Low-field H-1 NMR spectroscopy for distinguishing between arabica and robusta ground roast coffees

This work reports a new screening protocol for addressing issues of coffee authenticity using low-field (60 MHz) bench-top H-1 NMR spectroscopy. Using a simple chloroform-based extraction, useful spectra were obtained from the lipophilic fraction of ground roast coffees. It was found that 16-O-methylcafestol (16-OMC, a recognized marker compound for robusta beans) gives rise to an isolated peak in the 60 MHz spectrum, which can be used as an indicator of the presence of robusta beans in the sample. A total of 81 extracts from authenticated coffees and mixtures were analysed, from which the detection limit of robusta in arabica was estimated to be between 10% and 20% w/w. Using the established protocol, a surveillance exercise was conducted of 27 retail samples of ground roast coffees which were labelled as "100% arabica". None were found to contain undeclared robusta content above the estimated detection limit. (C) 2016 Published by Elsevier Ltd

High-throughput screening of argan oil composition and authenticity using benchtop 1H NMR

We use 60-MHz benchtop nuclear magnetic resonance (NMR) to acquire(1)H spectra from argan oils of assured origin. We show that the low-field NMR spectrum of neat oil contains sufficient information to make estimates of compositional parameters and to inform on the presence of minor compounds. A screening method for quality and authenticity is presented based on nearest-neighbour outlier detection. A variety of oil types are used to challenge the method. In a survey of retail-purchased oils, several instances of fraud were found

16-O-methylcafestol is present in ground roast Arabica coffees: Implications for authenticity testing

High-field and low-field proton NMR spectroscopy were used to analyse lipophilic extracts from ground roast coffees. Using a sample preparation method that produced concentrated extracts, a small marker peak at 3.16 ppm was observed in 30 Arabica coffees of assured origin. This signal has previously been believed absent from Arabicas, and has been used as a marker for detecting adulteration with robusta. Via 2D 600 MHz NMR and LC-MS, 16-O-methylcafestol and 16-O-methylkahweol were detected for the first time in Arabica roast coffee and shown to be responsible for the marker peak. Using low-field NMR, robusta in Arabica could be detected at levels of the order of 1-2% w/w. A surveillance study of retail purchased "100% Arabica" coffees found that 6 out of 60 samples displayed the 3.16 ppm marker signal to a degree commensurate with adulteration at levels of 3-30% w/w

Mitigating instrument effects in 60 MHz 1H NMR spectroscopy for authenticity screening of edible oils

Low field (60 MHz) 1H NMR spectroscopy was used to analyse a large (n = 410) collection of edible oils, including olive and argan, in an authenticity screening scenario. Experimental work was carried out on multiple spectrometers at two different laboratories, aiming to explore multivariate model stability and transfer between instruments. Three modelling methods were employed: Partial Least Squares Discriminant Analysis, Random Forests, and a One Class Classification approach. Clear inter-instrument differences were observed between replicated data collections, sufficient to compromise effective transfer of models based on raw data between instruments. As mitigations to this issue, various data pre-treatments were investigated: Piecewise Direct Standardisation, Standard Normal Variates, and Rank Transformation. Datasets comprised both phase corrected and magnitude spectra, and it was found that that the latter spectral form may offer some advantages in the context of pattern recognition and classification modelling, particularly when used in combination with the Rank Transformation pre-treatment

Venus trapped, Mars transits: Cu and Fe redox chemistry, cellular topography, and in situ ligand binding in terrestrial isopod hepatopancreas

Woodlice efficiently sequester copper (Cu) in ‘cuprosomes' within hepatopancreatic ‘S' cells. Binuclear ‘B’ cells in the hepatopancreas form iron (Fe) deposits; these cells apparently undergo an apocrine secretory diurnal cycle linked to nocturnal feeding. Synchrotron-based µ-focus X-ray spectroscopy undertaken on thin sections was used to characterize the ligands binding Cu and Fe in S and B cells of Oniscus asellus (Isopoda). Main findings were: (i) morphometry confirmed a diurnal B-cell apocrine cycle; (ii) X-ray fluorescence (XRF) mapping indicated that Cu was co-distributed with sulfur (mainly in S cells), and Fe was co-distributed with phosphate (mainly in B cells); (iii) XRF mapping revealed an intimate morphological relationship between the basal regions of adjacent S and B cells; (iv) molecular modelling and Fourier transform analyses indicated that Cu in the reduced Cu+ state is mainly coordinated to thiol-rich ligands (Cu–S bond length 2.3 Å) in both cell types, while Fe in the oxidized Fe3+ state is predominantly oxygen coordinated (estimated Fe–O bond length of approx. 2 Å), with an outer shell of Fe scatterers at approximately 3.05 Å; and (v) no significant differences occur in Cu or Fe speciation at key nodes in the apocrine cycle. Findings imply that S and B cells form integrated unit-pairs; a functional role for secretions from these cellular units in the digestion of recalcitrant dietary components is hypothesized

Loss of ACTN3 gene function alters mouse muscle metabolism and shows evidence of positive selection in humans

More than a billion humans worldwide are predicted to be completely deficient in the fast skeletal muscle fiber protein α-actinin-3 owing to homozygosity for a premature stop codon polymorphism, R577X, in the ACTN3 gene. The R577X polymorphism is associ

Born in Bradford’s Age of Wonder cohort : protocol for adolescent data collection

Background Adolescence and transition into adulthood are periods shaping life-long mental health, cardiometabolic risk, and inequalities. However, they are poorly studied and understood. By extending and expanding the Born in Bradford (BiB) cohort study through this period using innovative, co-produced approaches to collect and analyse data, we aim to understand better the interplay of factors that influence health and wellbeing, and inform/evaluate interventions to improve them and reduce inequalities. Protocol BiB Age of Wonder (AoW) is a large, whole city cohort that will capture the contemporary lived experience amongst multi-ethnic adolescents progressing into young adulthood. We will collect repeated data from existing BiB participants and their peers (N~30,000 adolescents). The protocol for the first phase of the quantitative methods, involving survey measurements and health assessments in mainstream secondary schools is described here. We describe the co-production behind these methods, and lessons learned from the first year of data collection