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.     

Principles and Limitations of Stable Isotopes in Differentiating Organic and Conventional Foodstuffs: 1. Plant Products

Among the lighter elements having two or more stable isotopes (H, C, N, O, S), δ¹⁵N appears to be the most promising isotopic marker to differentiate plant products from conventional and organic farms. Organic plant products vary within a range of δ¹⁵N values of +0.3 to +14.6%, while conventional plant products range from negative to positive values, i.e. ?4.0 to +8.7%. The main factors affecting δ¹⁵N signatures of plants are N fertilizers, biological N₂ fixation, plant organs and plant age. Correlations between mode of production and δ¹³C (except greenhouse tomatoes warmed with natural gas) or δ³⁴S signatures have not been established, and δ²H and δ¹⁸O are unsuitable markers due to the overriding effect of climate on the isotopic composition of plant-available water. Because there is potential overlap between the δ¹⁵N signatures of organic and conventionally produced plant products, δ¹⁵N has seldom been used successfully as the sole criterion for differentiation, but when combined with complementary analytical techniques and appropriate statistical tools, the probability of a correct identification increases. The use of organic fertilizers by conventional farmers or the marketing of organic produce as conventional due to market pressures are additional factors confounding correct identification. The robustness of using δ¹⁵N to differentiate mode of production will depend on the establishment of databases that have been verified for individual plant products.     

Influence of technological processing on apple aroma analysed by high resolution gas chromatography–mass spectrometry and on-line gas chromatography-combustion/pyrolysis-isotope ratio mass spectrometry

Extracts obtained by simultaneous distillation extraction (SDE) from industrial raw materials, namely single strength apple juices, and concentrates and aromas made thereof (each n = 31, from one production line; origin Poland, Germany, Turkey, Romania and China), as well as commercially available juices (n = 27), were analysed by standard controlled capillary gas chromatography–mass spectrometry (HRGC–MS). During the technological processing from juice to the aroma, no qualitative changes in the apple aroma profile were observed. Major constituents of the juices and aromas under study were found to be 1-hexanol (juice, 0.06–5.9 mg/l; aroma, 47–685 mg/l), 1-butanol (juice, 0.1–4.7 mg/l; aroma, 17–370 mg/l); E-2-hexenol (juice, 0.01–3.4 mg/l; aroma, 12–300 mg/l); E-2-hexenal (juice, 0–3.0 mg/l; aroma 0–470 mg/l), and butyl acetate (juice, 0–1.7 mg/l; aroma, 0–165 mg/l). By far the major component of the apple juice concentrates under study was furfural (2.4–56 mg/kg). The observed occurrence of 3-methyl-1-butanol (juice, 0.01–2.1 mg/l; aroma, 1.5–134 mg/l) and, in part, its acetate (juice, 0–0.3 mg/l; aroma, 0–3.3 mg/l), both known not to be genuine apple constituents, was obviously caused by fermentative effects in the course of industrial juice production. In addition, on-line capillary gas chromatography–isotope ratio mass spectrometry was used in the combustion (C) and the pyrolysis (P) modes (HRGC–C/P–IRMS) for the determination of δ¹³C_V-PDB and δ²H_V-SMOW values of selected apple flavour constituents to check potential isotope discrimination during distillative aroma production. As shown by means of the representative examples of E-2-hexenal, 1-hexanol and E-2-hexenol, their δ²H_V-SMOW values were slightly depleted. However, authenticity assessment by stable IRMS will not be influenced by this effect.     

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”.     

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.     

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.     

Δ desaturase protein expression and fatty acid composition of longissimus dorsi muscle in lambs fed green herbage or concentrate with or without added tannins

The aims of this study were to investigate the effect of feeding system and of supplementation of tannins (8.93% DM) on the relationship between intramuscular fat content, fatty acid composition and Δ⁹desaturase (Δ⁹d) protein expression in longissimus dorsi muscle of lamb. Twenty-eight Comisana lambs (age 45 days) were fed either vetch (Vicia sativa) or concentrate. The herbage diet was (i) lower in saturated fatty acids (especially in C16:0), C18:1 n−9 and in C18:2 n−6; (ii) higher in C16:1 and C18:3 n−3 when compared to concentrate. Within each feeding system the lambs were divided into two sub-groups, one of which received the diet without tannins supplementation, and the other was fed the diets supplemented with the tannins from Quebracho (Schinopsis lorentzii). The animals were slaughtered at age 105 days. The concentrate feeding system increased (p < 0.01) the total intramuscular fat content and the amount of SFA, MUFA and n−6 PUFA and decreased the level of n−3 PUFA (p = 0.05) when compared to the vetch-fed animals but did not affect Δ⁹ desaturase protein expression. There was no correlation between Δ⁹d protein expression and total intramuscular fatty acids, CLA and MUFA level. It was suggested that in ruminants, in contrast to monogastric animals, Δ⁹d expression does not play the key role in intramuscular fatty acids formation. Tannins supplementation resulted in higher (p < 0.05) muscle levels of transC18:1 and C18:2 n−6. It has also increased Δ⁹d expression in the case of herbage-based diet but not in the case of concentrate-based diet. The mechanism of tannins action on the enzyme expression needs to be elucidated.     

Isolation and evaluation of the antioxidant activity of phenolic constituents of the Garcinia brasiliensis epicarp

A new glycosylated biflavonone, morelloflavone-4′″-O-β-d-glycosyl, and the known compounds 1,3,6,7-tetrahydroxyxanthone, morelloflavone (fukugetin) and morelloflavone-7″-O-β-d-glycosyl (fukugeside) were isolated from the epicarp of Garcinia brasiliensis collected in Brazil. The structures of these compounds were established using ¹H and ¹³C NMR, COSY, gHMQC and gHMBC spectroscopy. The compounds exhibited antioxidant activity. The greatest potency was displayed by morelloflavone (2), with IC₅₀ = 49.5 mM against DPPH and absorbance of 0.583 at 400 μg/mL for the reduction of Fe³⁺. The weakest potency was displayed by 1,3,6,7-tetrahydroxyxanthone (1), with IC₅₀ = 148 mM against DPPH and absorbance of 0.194 at 400 μg/mL for the reduction of Fe³⁺.     

Enzymatic synthesis of primeverosides using transfer reaction by Trichoderma longibrachiatum xylanase

Enzymatic synthesis of two phenyl xylopyranosyl glucopyranosides, through transfer reaction by Trichoderma longibrachiatum endoxylanase, was achieved in the presence of n-hexane used as solvent, phenyl glucoside (10 mM) as acceptor and xylan (2 g/l) as donor. Kinetic study showed that only one compound, identified by ¹H and ¹³C NMR and heteronuclear 2D (¹H–¹³C) chemical shift correlation as phenyl primeveroside (phenyl 6-O-β-xylopyranosyl-1-β-d-glucopyranoside), was synthesized when the reaction time was beyond 1 h. Benzyl and hexyl primeverosides were obtained under the same conditions. When several phenyl glucoside concentrations, from 5 to 50 mM, were used with 2 g/l of xylan, a phenyl primeveroside isomer, identified as phenyl 4-O-β-xylopyranosyl-β-d-glucopyranoside, accumulated in the medium whereas the production of phenyl primeveroside decreased. Only phenyl primeveroside was produced when several xylan concentrations from 2 to 10 g/l were used with 10 mM of phenyl glucoside and its concentration in the reaction mixture increased with the increase of xylan concentration.     

Comparison of sterol composition between Tuber fermentation mycelia and natural fruiting bodies

Truffle, belonging to Tuber genera, is a nutritious and sterol-rich edible fungus, and sterol is a potential health beneficial compound. A comparison of Tuber sterol composition indicates that the total sterol contents in the fermentation mycelia (i.e., 10.5 mg g⁻¹) (n = 3) were approximately 3.2-5.6 times higher than that of the fruiting bodies (p < 0.05) with the addition of soybean flour into the basal fermentation media. Moreover, the phytosterol profile of fermentation mycelia could be significantly improved by adding soybean flour into the fermentation media. After the addition of soybean flour, stigmasterol and β-sitosterol appeared in the fermentation mycelia and the contents of total phytosterols (2279 μg g⁻¹) (n = 3), brassicasterol (943 μg g⁻¹) (n = 3), and campesterol (418 μg g⁻¹) (n = 3) were all increased significantly (p < 0.05). Moreover, the total contents of sterols and phytosterols in the fermentation mycelia cultured in the soybean media were much higher than those in the fruiting bodies (i.e., 1883-3240 and 479-1832 μg g⁻¹, respectively) (n = 3, p < 0.05). This work confirms the potentiality of Tuber fermentation mycelia as the alternative resource for its fruiting bodies from the viewpoint of sterols production.     

Evaluation of the performance of osmotic dehydration sheets on freshness parameters in cold-stored beef biceps femoris muscle

This study aimed to evaluate the effects of osmotic dehydration sheet (ODS) packaging on the quality parameters of beef biceps femoris muscle samples stored at 4 °C for 0, 1, 3 and 7 days. Quality indices such as Hunter color values (L^∗, a^∗ and b^∗, the percentage of metmyoglobin (Met-Mb%), K value (freshness index), and the contents of adenosine triphosphate (ATP)-related compounds (ARCs), thiobarbituric acid reactive substances (TBA-RS) and volatile basic nitrogen (VBN) were measured. ODS gave lower a^∗ and b^∗ values and lower Met-Mb% compared with control samples wrapped in polyvinylidene chloride film (PVDCF) (P < 0.01), but had no effect on L^∗ (P < 0.01). As a result, with higher levels of osmotic dehydration produced by the ODS, the percentage of weight loss and the total contents of ARCs and inosine monophosphate of the samples also increased (P < 0.05). The K values of ODS samples were also significantly lower than PVDCF-wrapped samples (P < 0.05). Low performance ODS wrapping reduced the TBA-RS values below those found with PVDCF and high performance ODS processing (P < 0.01). Moreover, the use of ODS had no effect on VBN values. Thus, although the bright red of beef samples changed to a dark purple color and the weights of samples decreased, the ODS approach has potential as a tool for decreasing the deterioration of other quality indices such as Met-Mb%, TBA-RS, ARCs, K values and the VBN content of cold-stored beef.     

Antioxidant and antiacetylcholinesterase activities of essential oils from Cymbopogon schoenanthus L. Spreng. Determination of chemical composition by GC–mass spectrometry and C NMR

Cymbopogon schoenanthus L. Spreng, is an aromatic herb consumed in salads and used to prepare traditional meat recipes in Tunisia. The chemical composition, antioxidant activities and acetylcholinesterase inhibitory properties of the essential oils from fresh leaves, dried leaves and roots collected from three different locations in southern Tunisia, were evaluated. Essential oils were analysed by GC–mass spectrometry and ¹³C NMR. The major components were limonene (10.5–27.3%), β-phellandrene (8.2–16.3%), δ-terpinene (4.3–21.2%) and α-terpineol (6.8–11.0%). Antioxidant activity was measured by DPPH assay. The results ranged from 36.0% to 73.8% (2 μl of essential oil per mL of test solution).     

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.     

Encapsulation of lipid by alginate beads reduces bio-accessibility: An in vivoC breath test and MRI study

This paper describes an in vivo study of the use of encapsulation to alter lipid bio-accessibility in the gastro-intestinal (GI) tract. The hypothesis that encapsulation would delay the accessibility of lipid in the GI tract was tested using subjects (N = 11) who consumed either 30 g of ¹³C-labelled sunflower oil encapsulated within 70 g alginate gel particles, or unencapsulated ‘free’ sunflower oil and alginate-only particles (control). Lipid accessibility was determined by the appearance of the ¹³C label in breath carbon-dioxide (CO₂), and the particles behaviour in the GI tract was investigated using non-invasive magnetic resonance imaging (MRI).     

Isotopic consequences of consumer food choice: Hydrogen and oxygen stable isotope ratios in foods from fast food restaurants versus supermarkets

We investigated geographic trends in the isotopic composition of the modern American diet, purchasing paired food items from fast food restaurants and supermarkets across the USA. We observed large ranges in δ²H and δ¹⁸O values, suggesting variation in the region-of-origin for beef, wheat, and potatoes. Mean restaurant meal δ²H and δ¹⁸O values (−114 and 22.6‰, respectively) were similar to supermarket values (−111 and 22.1‰, respectively). There were no correlations between restaurant beef and local tap water isotope values but significant correlations between supermarket beef and water (δ²H_beef = δ²H_water * 0.19−115‰ and δ¹⁸O_beef = δ¹⁸O_water * 0.17 + 14.8‰) suggesting regionality in the source of beef available to supermarket patrons. We observed no correlations between the stable isotopic composition of carbohydrates and local tap water. Understanding regional differences observed in some foods but not others will help refine parameters in models used to explore human movements in anthropological, archaeological, and forensic studies.     

C NMR and electrospray ionization mass spectrometric study of sucrose aqueous solutions at high pH: NMR measurement of sucrose dissociation constant

The ¹³C NMR technique is used for the measurement of the first dissociation constant of sucrose (HL) in highly alkaline solutions. In 1.0 M NaCl/NaOH medium and for 25 °C, the concentration dissociation constant (pK₁) was 13.1 ± 0.3; and, for 60 °C, pK₁ = 12.30 ± 0.05. The β-d-fructofuranosyl ring was found to be responsible for dissociation. The NMR data reveal no clear evidence of the second dissociation step below pH 14, either at 25 °C or at 60 °C. In the solutions with 4–10 mol dm⁻³ NaOH content the ¹³C NMR technique indicated the chemical shift changes, treated as the second dissociation step of sucrose and a sodium complex formation. A very rough estimation, for variable ionic strength, gives the value: pK₂ ∼ 15.8 ± 0.8. The anionic species L⁻ and NaH₋₁L⁻ have been registered by electrospray ionization time-of-flight mass spectrometry (ESI-ToF MS) for 0.01 M sucrose solutions with initial pH 13.     

Zinc fortification of whole rice grain through parboiling process

The present study evaluated the effectiveness of zinc (Zn) fortification in a parboiling process for improving Zn density in parboiled-polished rice and its potential bioavailability in the human diet. Fortification of Zn in whole paddy rice grain with 50–400 mg Zn/kg paddy rice, during parboiling, increased Zn concentrations in polished-parboiled rice from 1.3 to 4.5 times those in unfortified parboiled rice. The added Zn rapidly penetrated into parboiled rice grains in the initial soaking process before saturation. There was an exponential correlation between Zn concentrations in unpolished (r = 0.63) (p < 0.01) and polished rice (r = 0.30) (p < 0.05) and soaking time. Zinc concentrations in unpolished rice were linearly correlated with Zn concentration in the polished rice (r = 0.60) (p < 0.01). Moreover, more than half of the added Zn is retained after a simulated washing process before cooking, ranging from 64–100%. In the Zn-fortified parboiled rice, 57–100% of Zn in polished rice grain was soluble in dilute acid, which was indicative of a high potential Zn bioavailability for human intake. The results suggest that parboiled rice has great potential for Zn fortification.     

Speciation of organic and inorganic selenium in selenium-enriched rice by graphite furnace atomic absorption spectrometry after cloud point extraction

A new method was developed for the determination of organic and inorganic selenium in selenium-enriched rice by graphite furnace atomic absorption spectrometry detection after cloud point extraction. Effective separation of organic and inorganic selenium in selenium-enriched rice was achieved by sequentially extracting with water and cyclohexane. Under the optimised conditions, the limit of detection (LOD) was 0.08 μg L⁻¹, the relative standard deviation (RSD) was 2.1% (c = 10.0 μg L⁻¹, n = 11), and the enrichment factor for selenium was 82. Recoveries of inorganic selenium in the selenium-enriched rice samples were between 90.3% and 106.0%. The proposed method was successfully applied for the determination of organic and inorganic selenium as well as total selenium in selenium-enriched rice.     

Contents of biologically active polyamines in chicken meat,liver, heart and skin after slaughter and their changes during meat storage and cooking

Dietary polyamines, putrescine, spermidine (SPD) and spermine (SPM), participate in an array of important physiological roles, including tumour growth. Thus, reliable information on polyamine content in foods has been needed. We therefore determined polyamine contents in chilled chicken meat and giblets (n = 20) and skin (n = 10) 24 h after slaughter. The polyamines were determined, after extraction with perchloric acid, as dansyl derivatives, using an HPLC method. Mean SPD values were 4.8, 10.2, 11.4, 48.7 and 12.1 mg kg⁻¹ and SPM values were 36.8, 38.0, 24.3, 133 and 82.7 mg kg⁻¹ in breast, thigh, skin, liver and heart, respectively. Significant statistical correlations between SPD and SPM contents were observed in breast, thigh, skin and liver, whereas correlations were insignificant in heart. An increase of SPD and SPM was apparent in breasts and thighs stored at −18 °C for 6 months; however, it was significant only for SPM in thighs. The losses of both SPD and SPM were statistically insignificant during storage of aerobically packaged breasts up to 9 days at +2 °C. A significant decrease of SPM to about 60% of the initial contents was observed in both vacuum-packaged and in modified atmosphere (20% CO₂ and 80% O₂)-stored breasts on day 21 at +2 °C. For both SPD and SPM, roasting, grilling and frying of fresh breasts caused losses of about 40–60% of the initial contents (higher than boiling and stewing). Similarly, losses of SPM, due to roasting of breasts frozen for 3 or 6 months, were higher than those caused by stewing. Putrescine was detected only sporadically and at levels close to the detection limit of 1.0 mg kg⁻¹ (fresh matter).