Principles and limitations of stable isotopes in differentiating organic and conventional foodstuffs: 2. Animal products

In this review, we examine the variation in stable isotope signatures of the lighter elements (δ²H, δ¹³C, δ¹⁵N, δ¹⁸O, and δ³⁴S) of tissues and excreta of domesticated animals, the factors affecting the isotopic composition of animal tissues, and whether stable isotopes may be used to differentiate organic and conventional modes of animal husbandry. The main factors affecting the δ¹³C signatures of livestock are the C3/C4 composition of the diet, the relative digestibility of the diet components, metabolic turnover, tissue and compound specificity, growth rate, and animal age. δ¹⁵N signatures of sheep and cattle products have been related mainly to diet signatures, which are quite variable among farms and between years. Although few data exist, a minor influence in δ¹⁵N signatures of animal products was attributed to N losses at the farm level, whereas stocking rate showed divergent findings. Correlations between mode of production and δ²H and δ¹⁸O have not been established, and only in one case of an animal product was δ³⁴S a satisfactory marker for mode of production. While many data exist on diet–tissue isotopic discrimination values among domesticated animals, there is a paucity of data that allow a direct and statistically verifiable comparison of the differences in the isotopic signatures of organically and conventionally grown animal products. The few comparisons are confined to beef, milk, and egg yolk, with no data for swine or lamb products. δ¹³C appears to be the most promising isotopic marker to differentiate organic and conventional production systems when maize (C4) is present in the conventional animal diet. However, δ¹³C may be unsuitable under tropical conditions, where C4 grasses are abundant, and where grass-based husbandry is predominant in both conventional and organic systems. Presently, there is no universal analytical method that can be applied to differentiate organic and conventional animal products.     

Geographical origin of polished rice based on multiple element and stable isotope analyses

We determined carbon and nitrogen contents (C and N contents) and stable carbon, nitrogen, and oxygen isotopic compositions (δ¹³C, δ¹⁵N, and δ¹⁸O) of polished rice in order to develop a simple method to discriminate its geographical origin. As a first attempt, we examined a single cultivar, Koshihikari rice, from 14 different cultivation areas including Australia (n = 1), Japan (n = 12), and USA (n = 1). For all rice samples, C and N contents and the isotopic compositions are consistent with those of general plant materials, being 37.2–40.0% (C content), 0.8–1.4% (N content), −27.1 to −25.4% (δ¹³C), +0.4 to +9.0% (δ¹⁵N), and +18.8 to +22.9% (δ¹⁸O). However, its cultivated area is clearly distinguished by a pentagonal radar plot based on the elemental and isotopic compositions. Thus, the comparison of C and N contents and δ¹³C, δ¹⁵N, and δ¹⁸O values would potentially be useful for rapid and routine discrimination of geographical origin of the polished rice.     

Seasonal variations in chemical composition and stable isotopes of farmed and wild Brazilian freshwater fish

This study examined variations in the Fulton condition factor, chemical composition, and stable isotopes of carbon and nitrogen in the Brazilian freshwater fish cachara (Pseudoplatystoma fasciatum), comparing farmed and wild fish in different seasons. Values for energy, protein, moisture, and Fulton’s condition factor were higher for farmed than for wild fish in the rainy season, indicating better nutritional quality; however, these differences were not observed in the dry season. Likewise, we found significant enhancement of δ¹⁵N in farmed fish in the rainy season but not in the dry season, whereas enhancement of δ¹³C was observed in both seasons. The combined measurement of δ¹³C and δ¹⁵N provided traceability under all conditions. Our findings show that stable isotope analysis of C and N can be used to trace cachara origin, and that seasonal variations need to be considered when applying chemical and isotopic authentication of fish and fish products.     

Seasonal variation in the C,N and S stable isotope composition of retail organic and conventional Irish beef

The objective of this study was to investigate seasonal variation in the C, N and S stable isotope composition of retail organic and conventional Irish beef. A total of 242 beef samples (127 organic, 115 conventional) was collected in a one-year survey and analysed by isotope ratio mass spectrometry. The δ¹³C time series in conventional beef was significantly non-random, with a pronounced seasonal positive shift of >2‰ between December and June, whilst δ¹³C in organic beef was less variable and significantly more negative. In conventional beef, δ¹⁵N was remarkably invariant (remaining close to 7‰) throughout the year, while organic beef was more variable and also significantly lower in δ¹⁵N. The S isotope composition (δ³⁴S) exhibited a complex seasonal pattern in both types of beef. These results show that seasonal patterns can occur in the isotopic composition of beef, probably reflecting seasonality in animal feeding practices modulated by tissue turnover rates. Such seasonal variation needs to be considered in the isotopic authentication of beef and other animal-derived products.     

Influence of N doses and form on ¹⁵N natural abundance of pepper plants: considerations for using δ¹⁵N values as indicator of N source

BACKGROUND: The aim of this study was to ascertain the effect of the N form (NO₃^?, NH₄⁺ and organic N) and N concentration on plant isotopic fractionation and on the contribution of the different N sources to the plant N budget, in order to evaluate the feasibility of using plant δ¹⁵N values for discriminating between conventional and organic crops. To this end, different N concentrations (applied as NO₃^?), N forms (NO₃^? versus NH₄⁺), and increasing NO₃^? applications to an organic N‐based fertilization regime were studied. RESULTS: When using NO₃^? as N source, intra‐plant fractionation was significant and tended to increase when NO₃^? concentration increased in the root medium. However, negligible net isotopic fractionation was observed when comparing theoretical and measured plant δ¹⁵N values. On the other hand, when plants are fertilized with a mixture of NO₃^? and NH₄⁺, differences in uptake patterns for both salts could result in variation in plant δ¹⁵N regarding to the expected value. Finally, the application of NO₃^? to plants was detected when it was combined with high levels of organic N sources, from 99:1 organic:inorganic N ratio. CONCLUSION: Under certain conditions and following some considerations concerning sampling, δ¹⁵N values can be considered to be a potential tool to guarantee the authenticity of organic products. Copyright © 2011 Society of Chemical Industry     

氮肥种类对番茄δ~(15)N值的影响

在番茄生长过程中施加不同种类氮肥,并在固定时间取土壤、肥料、植株及果实样品,通过上述样品的氮稳定同位素比率特征研究氮肥种类对番茄δ15N值的影响。结果表明,氮肥种类对番茄果实δ15N值具有显著影响,施入化学氮肥的番茄果实中δ15N值会显著低于未施化学氮肥的番茄果实;相同氮肥条件下,时间因素对番茄δ15N值差异影响不显著;番茄植株(叶和茎)的δ15N值始终大于同株果实的δ15N值,与时间和氮肥种类无关;同时,植物样品的δ15N值一般高于土壤或肥料的δ15N值。植物δ15N值可以作为判断蔬菜在种植过程中是否施入化肥的指标,为区分有机和常规蔬菜提供理论和技术支持。     

Can δ15N in lettuce tissues reveal the use of synthetic nitrogen fertiliser in organic production?

BACKGROUND: The nitrogen isotopic fingerprint (δ¹⁵N) is reported to be a promising indicator for differentiating between organically and conventionally grown vegetables. However, the effect on plant δ¹⁵N of split nitrogen fertilisation, which could enable farmers to cover up the use of synthetic fertiliser, is not well studied. In this study the use of δ¹⁵N in lettuce as a potential marker for identifying the use of synthetic nitrogen fertiliser was tested on pot‐grown lettuce (Lactuca sativa L.) treated with synthetic and organic nitrogen fertilisers (single or split application). The effect of combined usage of synthetic and organic fertilisers on δ¹⁵N was also investigated. RESULTS: The δ¹⁵N values of whole plants treated with different fertilisers differed significantly when the fertiliser was applied in a single treatement. However, additional fertilisation (with isotopically the same or different fertiliser) did not cause a significant alteration of plant δ¹⁵N. CONCLUSION: The findings of the study suggest that the δ¹⁵N value of lettuce tissues could be used as a rough marker to reveal the history of nitrogen fertilisation, but only in the case of single fertiliser application. However, if the difference in δ¹⁵N between the applied synthetic and organic nitrogen fertilisers was > 9.1‰, the detection of split and combined usage of the fertilisers would have greater discriminatory power. Copyright © 2010 Society of Chemical Industry     

Interaction between nitrogen and sulfur by foliar application and its effects on flour bread‐making quality

BACKGROUND: Optimization of nitrogen (N) and sulfur (S) nutrition of wheat influences the bread‐making quality of grain. This study was conducted to determine whether the application of N and S in winter wheat at anthesis stage during two field trials could influence the absorption and distribution of N and S in grain and flour bread‐making quality RESULTS: The incorporation of ¹⁵N and ³⁴S tracers in various plant parts (leaves, stems, spikes) was determined by isotopic ratio mass spectrometry coupled with an elemental analyser (EA‐IRMS). The effects of foliar fertilization on percentage of ¹⁵N and ³⁴S recovery in various plant parts, grain yield, flour protein content, and dough properties were investigated after a combination of treatments: N (urea), S (micronized elemental S) and NS (both urea and micronized elemental S). Nitrogen (70–80%) and S (35–55%) were quickly assimilated during the days following application. Sixty days after application, 42–68% N and 6–12% S were recovered in the grain. When N and S fertilizers were applied simultaneously, N and S recoveries (68% and 12%), N/S ratios (11–14), flour protein content (115 g kg^?1) and dough strength (310 × 10^?4 J), swelling (21.8 mmH₂O) and extensibility (96.4 mm) were increased. CONCLUSION: A synergistic effect between the foliar applied N and S fertilizers appears to increase N and S assimilation in grain and may improve bread‐making qualities. Copyright © 2007 Society of Chemical Industry     

Passage of stable isotope-labeled grass silage fiber and fiber-bound protein through the gastrointestinal tract of dairy cows

Fractional passage rates are required to predict nutrient absorption in ruminants but data on nutrient-specific passage kinetics are largely lacking. With the use of the stable isotope ratio (δ) as an internal marker, we assessed passage kinetics of fiber and fiber-bound nitrogen (N) of intrinsically labeled grass silage from fecal and omasal excretion patterns of δ¹³C and δ¹⁵N. In a 6 × 6 Latin square, lactating dairy cows received grass silages [455 g/kg of total diet dry matter (DM) ] in a 2 × 3 factorial arrangement from ryegrass swards fertilized at low (45 kg of N/ha) or high (90 kg of N/ha) levels of N and harvested at 3 maturity stages. Feed intake (16.7 ± 0.48 kg of DM/d; mean ± standard error of the mean) and milk yield (26.7 ± 0.92 kg/d) increased at the high level of N fertilization and at decreasing maturity. Nutrient digestibility decreased with increasing plant maturity, particularly at the high level of N fertilization, essentially reflecting dietary treatment effects on the nutritional composition of the grass silage. Fractional rumen passage rates (K₁) were highest and total mean retention time in the gastrointestinal tract (TMRT) was lowest when based on the external marker chromium mordanted fiber (Cr-NDF; 0.047/h and 38.0 h, respectively). Fecal δ¹³C in the acid detergent fiber fraction (¹³CADF) provided the lowest K₁ (0.023/h) and the highest TMRT (61.1 h) and highest peak concentration time (PCT; 24.3 h) among markers. In comparison, fecal fiber-bound N (¹⁵NADF) had a considerably higher K₁ (0.032/h) and lower TMRT (46.4 h) than ¹³CADF. Total N (measured with ¹⁵NDM) had a comparable K₁ (0.034/h) to that of ¹⁵NADF but provided the highest fractional passage rates from the proximal colon-cecum (K₂; 0.37/h) and lowest PCT (17.4 h) among markers. A literature review indicated unclear effects of grass silage maturity on K₁ and unknown effects of N fertilization on K₁. Our study indicated no effect of advancing maturity on fecal K₁ and a trend for K₁ to increase with the high level of N fertilization. Parameter K₂ increased, whereas PCT and TMRT generally decreased with the high level of N fertilization. Omasal digesta sampling largely confirmed results based on fecal sampling. Results indicate that the use of δ¹³C and δ¹⁵N can describe fiber-specific passage kinetics of forage.     

Geographical origin of imported and domestic teas (Camellia sinensis) from Turkey as determined by stable isotope signatures

In this study, stable isotope signatures (δ¹³C, δ¹⁵N, and δD) of both tea leaves and tea infusions were investigated to identify the geographical origin of Turkish domestic and imported tea samples. Sixteen domestic tea samples collected from different locations in the Black Sea Region, which produces almost 100% of tea in Turkey, and 11 imported tea samples (Kenya, India, Sri Lanka, Indonesia, and China) purchased from importers were studied. δ¹³C, δ¹⁵N, and δD in the samples were determined using isotope ratio mass spectrometry (IR-MS). δ¹³C in the samples ranged from ?29.18 ± 0.01 to ?25.7 ± 0.2, while δ¹⁵N ranged between 1.1 ± 0.2 and 5.2 ± 0.8. However, δD in the samples were found to be in the range from 56.5 ± 0.3 to 72 ± 1. The classifications of the tea samples into domestic and imported tea samples were achieved with 100% accuracy using multivariate statistical analyses (principal component analysis, PCA, and hierarchical cluster analysis, HCA). In conclusion, the domestic tea samples had a distinctive isotopic fingerprint and the isotopic ratios used in the study can be significant predictors in determination of the geographical source of Turkish tea.     

The combination of stable isotope abundance ratios of H,C, N and S with Sr/Sr for geographical origin assignment of orange juices

Within the EU-project “Pure Juice” established stable isotope methods (δ²H, δ¹³C, δ¹⁵N) have been applied and improved in order to determine and verify the geographical origin of orange juices. In addition, new approaches employing analyses of δ³⁴S and ⁸⁷Sr/⁸⁶Sr have been developed and tested. Approximately 150 authentic orange juice samples from several regions in North- and South-America, Africa and Europe have been analysed. A discrimination of orange producing regions, based on the results which ultimately depend on geographical, climatic and lithological differences was successfully performed. Furthermore, we demonstrate that blending of single strength juice by adding concentrate can be revealed by comparing ⁸⁷Sr/⁸⁶Sr of soluble and insoluble components of the juices. We conclude that regional assignment of orange juice samples is most successful when single parameters are combined in a “multi-element approach”.     

Evaluation of markers for the traceability of potato tubers grown in an organic versus conventional regime

Identification of parameters discriminating organic and conventional produce should help prevent misconduct and could provide a firm basis for comparative assessment of the two types of produce. In this report, we compare for selected markers organically versus conventionally grown tubers in four separate field trials. Within each field trial, organic and conventional tubers were subjected to the same pedoclimatic conditions as they were grown in adjacent plots. In all sites and in both cultivars tested, irrespective of environment, organic tubers exhibited a significant enrichment in ¹⁵N‰ when compared to their conventional counterparts. Also, the sum of all samples from the four sites (organic versus conventional) showed higher δ¹⁵N‰ (7.17 ± 48% versus 3.36 ± 38%; means ± CV%; n = 39; P < 0.001); furthermore, setting as threshold value for assignment in the organic group the lowest δ¹⁵N observed for organic samples (i.e. + 4.3‰), only 15% of conventional tubers were misclassified. On the other hand, further parameters selected on the basis of reports of alterations as a consequence of agricultural technique—ascorbic acid, protein content and dry matter—did not show consistent trends of variation throughout the four field trials. We conclude that, as opposed to other relevant parameters, ¹⁵N enrichment appears as a promising discriminative marker. Copyright © 2007 Society of Chemical Industry     

Measurement of stable isotope abundances in milk and milk ingredients — a possible tool for origin assignment and quality control

Relative carbon and nitrogen stable isotope abundances in total milk reflect the isotopic composition of the diet fed to the dairy cows; this diet and its δ-values depend on geographical and climatic factors. Milk from regions dominated by grassland typically shows relatively negative δ¹³C-values, while in regions dominated by crop cultivation the δ¹³C-values are more positive. The δ¹⁵N-values are influenced by factors such as soil conditions, the intensity of agricultural use and the climate. Casein in authentic milk samples is enriched in both ¹³C and ¹⁵N as compared to total milk, while the whey fraction is slightly enriched in ¹³C and depleted of ¹⁵N. The isotopic content of milk, casein and whey from one location have been measured throughout a period of longer than 1 year; variations are usually not greater than 1‰. In milk water, the ¹⁸O content was increased by between 2 and 6‰ as compared to ground water.     

Fertilizer characterization: isotopic data (N, S, O, C, and Sr)

A detailed isotopic characterization (delta15N(Ntotal), delta15N(NO3), delta18O(NO3), delta34S(SO4), delta18O(SO4), (delta13C(Ctotal), and 87Sr/86Sr) of 27 commercial fertilizers used in Spain is presented in this paper. Results together with a compilation of fertilizer isotopic published data are used for two purposes: (i) to identify the origin of the primary constituents and raw materials used in fertilizer manufacture and relate these data with their heavy metals and rare earth elements (REE) contents; (ii) to compare the fertilizer isotopic signatures with natural values and other anthropogenic pollutants and evaluate the usefulness of multi-isotopic analyses to trace fertilizer contaminations in future study cases. Isotope data permit us to know, in most cases, the origin of the primary constituents of fertilizers, and the 87Sr/86Sr ratio distinguishes the origin of the phosphate content--phosphorites or carbonatites--which in turn implies a qualitatively defined and potentially contaminant presence of REE and heavy metals in fertilizers. Delta15N, delta34S, and 87Sr/86Sr have already been used to trace fertilizer contaminations. Their utility can be improved by the coupled use of delta15N(NO3)-delta18O(NO3) and delta34S(SO4)-delta18O(SO4) to evaluate the fractionation processes that can affect contaminants. Moreover, multi-isotopic analyses, using heavy isotopes, allow us to see beyond the fractionation effects to the fertilizer stable isotope signatures and a better distinction from other anthropogenic contaminants.     

Potential isotopic and chemical markers for characterising organic fruits

Several isotopic (¹³C/¹²C, ¹⁵N/¹⁴N, ¹⁸O/¹⁶O, ²H/¹H, ³⁴S/³²S) and chemical–physical parameters (pH, fruit weight, juice yield, titratable acidity, total soluble solids, skin resistance, flesh firmness, colorimetric characteristics, weight loss after harvesting, antioxidant activity, earliness index, total nitrogen, ascorbic acid, synephrine, anthocyanins and polyphenols, citric acid, malic acid, sucrose, glucose and fructose content) were investigated as potential markers of organically cultivated oranges, clementines, strawberries and peaches produced in Italy between 2006 and 2008, in experimental fields and in certified farms. The ratio ¹⁵N/¹⁴N, ascorbic acid and total soluble solids were shown to be the most significant variables for distinguishing between organically and conventionally cultivated fruits. It was not possible to define general threshold limits typical of organic fruits because these parameters are influenced also by fruit specie, cultivar, year and site of production. Combining isotopic and chemical markers a good discrimination between organic and conventional fruits of different species was achieved.     

Comparison of mineral concentrations in commercially grown organic and conventional crops – Tomatoes (Lycopersicon esculentum) and lettuces (Lactuca sativa)

Trace element concentrations and stable nitrogen isotope data (δ¹⁵N‰) from tomatoes (Lycopersicon esculentum) and lettuces (Lactuca sativa) were subjected to multivariate analysis in an attempt to distinguish between conventional and organic cultivation. This approach improved the correct classification of tomato samples but appears to have had a limited effect on lettuces. Our findings support the growing body of evidence which suggests that systematic differences in the concentrations of certain elements such as manganese, calcium, copper, and zinc may occur between crops cultivated under organic and conventional regimes possibly due to the presence of elevated levels of arbuscular mycorrhizal fungi (AMF) in soils cultivated organically. We assert that such differences in elemental composition may be useful as ‘indicators of authenticity’. However, we recognise the limitation that this approach may be restricted to horticultural crops where there are significant differences in agricultural practice such as conventional-hydroponic versus soil-grown organic tomatoes.     

Reliability of stable isotope values from meat juice for the determination of the meat origin

For the determination of the origin of foods, analysis of the oxygen and hydrogen isotope composition of the food water often is of great importance, because of the differences in the isotope signature of water between different regions. However, it was reported previously that the changes in δ¹⁸O during storage of meat are of such magnitude that existing variations between regions and countries are obscured. We have investigated the δ²H and δ¹⁸O changes in meat juice after slaughtering during storage under real cold storage conditions. Our results disagree with a previous publication as we did not observe heavy isotope enrichment of meat juice during industrial cold storages. Additionally we investigated possible changes in δ¹⁸O of the meat juice during the roasting process.     

No tillage affects the phosphorus status,isotopic composition and crop yield of Phaseolus vulgaris in a rain‐fed farming system

BACKGROUND: Conservation tillage promotes the accretion of soil organic matter and often leads to improved soil fertility and moisture availability. However, few studies have looked at the physiological response of crop plants to different tillage practices. It was therefore hypothesised that measuring the nutrient concentrations and stable isotope composition (δ¹³C, δ¹⁸O, δ¹⁵N) of shoots could help evaluate the physiological response of common bean (Phaseolus vulgaris L.) to different tillage treatments (no tillage (NT) and mouldboard ploughing (MP)) in a rain‐fed farming system in northern Mexico. RESULTS: NT significantly enhanced shoot phosphorus concentration in bean plants. Tillage exerted a negative effect on the extent of root colonisation (%) by arbuscular mycorrhizal fungi (AMF). Lower shoot δ¹⁸O but unchanged δ¹³C values in plants from the NT system suggest enhanced stomatal conductance but also enhanced photosynthetic rate, which overall resulted in unchanged water use efficiency. Bean plants in the NT system showed lower shoot δ¹⁵N values, which suggests that a larger proportion of total plant nitrogen was obtained through atmospheric nitrogen fixation in this treatment. CONCLUSION: Greater diversity of AMF soil communities and heavier colonisation of roots by AMF in the NT compared with the MP system appeared to contribute to improved crop nutrition, water relations and yield in this rain‐fed agroecosystem. Copyright © 2010 Society of Chemical Industry     

Classification of geographical origins and prediction of δ13C and δ15N values of lamb meat by near infrared reflectance spectroscopy

Near infrared spectroscopy (NIRS) combined with chemometric analysis was investigated for its potential to classify the geographical origin and predict δ(13)C and δ(15)N values of lamb meat samples (n=99) from three pastoral regions and two agricultural regions of China. Principal component analysis (PCA), discriminant partial least squares analysis (D-PLS), linear discriminant analysis (LDA) and partial least squares regression (PLSR) were used for data analysis. D-PLS and LDA correctly classified 100% of the both pastoral and agricultural region samples, and gave a total correct classification of 88.9% and 75% to the five individual region samples, respectively. The best PLSR calibration models for predicting δ(13)C and δ(15)N of lamb meat were obtained with the determination coefficient (R(2)) 0.76 and 0.87, respectively. These results show that NIRS combined with chemometrics can be used as a rapid and effective method to discriminate the geographical origin and estimate the δ(13)C and δ(15)N of lamb meat.