Endothelial dysfunction in cerebral microcirculation during hypothermic cardiopulmonary bypass in newborn lambs

OBJECTIVES: Inflammatory stimuli or mechanical stresses associated with hypothermic cardiopulmonary bypass could potentially impair cerebrovascular function, resulting in inadequate cerebral perfusion. We hypothesize that hypothermic cardiopulmonary bypass is associated with endothelial or vascular smooth muscle dysfunction and associated cerebral hypoperfusion. Therefore we studied the cerebrovascular response to endothelium-dependent vasodilator, acetylcholine, endothelium-independent nitric oxide donor, sodium nitroprusside, and vasoactive amine, serotonin, in newborn lambs undergoing hypothermic cardiopulmonary bypass (nasopharygeal temperature = 18 degrees C). METHODS: Studies were performed on 13 newborn lambs equipped with a closed cranial window, allowing for direct visualization of surface pial arterioles. Six animals were studied while undergoing hypothermic cardiopulmonary bypass, whereas seven served as nonbypass, warm (37 degrees C) controls. Pial arteriolar caliber (range = 111 to 316 microm diameter) was monitored using video microscopy. RESULTS: Topical application of acetylcholine caused a dose-dependent increase in arteriolar diameter in the control group that was absent in animals undergoing hypothermic cardiopulmonary bypass. Hypothermic cardiopulmonary bypass did not alter the vasodilation in response to sodium nitroprusside. Furthermore, the contractile response to serotonin was fully expressed during hypothermic cardiopulmonary bypass. CONCLUSIONS: The specific loss of acetylcholine-induced vasodilation suggests endothelial cell dysfunction rather than impaired ability of vascular smooth muscle to respond to nitric oxide. It is speculated that loss of endothelium-dependent regulatory factors in the cerebral microcirculation during hypothermic cardiopulmonary bypass may enhance vasoconstriction, and impaired cerebrovascular function may be a basis for associated neurologic injury during or after hypothermic cardiopulmonary bypass.     

Direct electron paramagnetic resonance monitoring of the peptide synthesis coupling reaction in polymeric support

This work demonstrates, for the first time, a time-resolved electron paramagnetic resonance (EPR) monitoring of a chemical reaction occurring in a polymeric structure. The progress of the coupling of a N^α-tert-butyloxycarbonyl-2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid (Boc-TOAC) spin probe to a model peptide-resin was followed through EPR spectra. Progressive line broadening of EPR peaks was observed, indicative of an increased population of immobilized spin probe molecules attached to the solid support. The time for spectral stabilization of this process coincided with that determined in a previous coupling study, thereby validating this in situ quantitative monitoring of the reaction. In addition, the influence of polymer swelling degree and solvent viscosity, as well as of the steric hindrance within beads, on the rate of coupling reaction was also addressed. A deeper evaluation of the latter effect was possible by determining unusual polymer parameters such as the average site-site distance and site-concentration within resin beads in each solvent system.     

Improved instrumentation for blood flow velocity measurements in the microcirculation of small animals

Microcirculation is the generic name of vessels with internal diameter less than 100 microm of the circulatory system, whose main functions are tissue nutrition and oxygen supply. In microcirculatory studies, it is important to know the amount of oxyhemoglobin present in the blood and how fast it is moving. The present work describes improvements introduced in a classical hardware-based instrument that has usually been used to monitor blood flow velocity in the microcirculation of small animals. It consists of a virtual instrument that can be easily incorporated into existing hardware-based systems, contributing to reduce operator related biases and allowing digital processing and storage. The design and calibration of the modified instrument are described as well as in vitro and in vivo results obtained with electrical models and small animals, respectively. Results obtained in in vivo studies showed that this new system is able to detect a small reduction in blood flow velocity comparing arteries and arterioles (p <0.002) and a further reduction in capillaries (p<0.0001). A significant increase in velocity comparing capillaries and venules (p <0.001) and venules and veins (p <0.001) was also observed. These results are in close agreement with biophysical principles. Moreover, the improvements introduced in the device allowed us to clearly observe changes in blood flow introduced by a pharmacological intervention, suggesting that the system has enough temporal resolution to track these microcirculatory events. These results were also in close conformity to physiology, confirming the high scientific potential of the modified system and indicating that this instrument can also be useful for pharmacological evaluations.     

Detoxification of Aflatoxin B1 and M1 by Lactobacillus acidophilus and prebiotics in whole cow's milk

The detoxification ability of Lactobacillus acidophilus and prebiotics was evaluated by a Plackett‐Burman Design to examine the reduction of concentration and bioaccessibility of aflatoxin B₁ (AFB₁) and M₁ (AFM₁) in artificially contaminated whole cow's milk. Six variables were evaluated: AFB₁ (3.25–6.0 μg L^?1) or AFM₁ (1.0–2.0 μg L^?1) concentration; incubation time (0–6 h); and inulin, oligofructose, β‐glucan, and polydextrose concentrations (each between 0.00 and 0.75%). All runs achieved reductions of AFB₁ (13.53–35.53%) and AFM₁ (17.65–71.52%). Comparing with the positive control, the AFB₁ bioaccessibility ranged from 23.68 to 72.67% and for AFM₁ was 0%. The probiotic, isolated or combined with prebiotics, was efficient in mycotoxin reduction, while the combination of the two reduced the mycotoxin bioaccessibility. The best experimental condition was the highest concentration of AFB₁ (6.5 μg L^?1) and AFM₁ (2.0 μg L^?1), incubation time of 0 h and the addition of probiotic and inulin (0.75%).     

Original article: The effect of freezing at −18 °C and −70 °C with and without ascorbic acid on the stability of antioxidant in extracts of apple and orange fruits

The influence of low temperature and storage time on the antioxidant capacity of standard solutions and apple and orange extracts was evaluated. In addition, the effects of ascorbic acid (AA) addition to the fruit extracts in terms of antioxidant capacity, AA content and soluble and hydrolyzable polyphenol contents were also analysed. Polyphenol contents in both apple and orange extracts were stable during storage period, which reflected also in the antioxidant capacity stability. Freezing at ?18 °C did not result in different retention rates for polyphenols, AA and antioxidant capacity when compared to freezing at ?70 °C. However, vitamin C content in orange juice, without AA addition, slightly increased along the experimental period (10 days). Thus, this study shows that it may not be necessary to measure the antioxidant capacity immediately after the preparation of fruit extracts or antioxidant standard solutions.     

Validation of an HPLC Methodology for the Identification and Quantification of Biogenic Amines in Chicken Meat

This study validated a high performance liquid chromatography (HPLC) method to determine biogenic amines in chicken meat. For the identification of biogenic amines, an isocratic elution system coupled with a UV detector (254 nm) was used after a perchloric acid (5 %) extraction and benzoyl chloride derivatization of the samples. The standards of tyramine, putrescine, cadaverine, spermidine, and spermine were used for the following validation parameters: selectivity, linearity, accuracy, recovery, limit of detection and quantification, and robustness. Finally, chicken meat commercialized in two types of packaging was evaluated. The results showed an excellent selectivity and separation of all amines, r ²?>?0.99, relative standard deviation <5 %, recovery between 64 and 112 %, limits of detection and quantification between 0.03–1.25 and 0.15–5.00 μg?L^?1, respectively, and appropriate robustness for the proposed methodology. Moreover, both chicken meat commercial packages had similar values for all amines; only tyramine was significantly different (P?≤?0.05). The proposed method was suitable to detection and quantification of simultaneous five biogenic amines in chicken meat.     

Pressurisation of raw skim milk and of a dispersion of phosphocaseinate at 9 degrees C or 20 degrees C: effects on the distribution of minerals and proteins between colloidal and soluble phases

The effects of high pressure treatments (100-300 MPa; 15 min; 9 degrees C or 20 degrees C) on the distribution of minerals and proteins of raw skim milk (RSM) and of a dispersion of industrial phosphocaseinate (PC) were studied after separation of the micellar and soluble phases by ultracentrifugation (UCF). Whatever the temperature of high pressure treatments, the pressure-induced dissociation of the casein micelles was accompanied by calcium (Ca), phosphorus (P) and casein release from the micelles. The released Ca and P were or became bound to soluble proteins since progressive increases in Ca and P concentrations were observed in the UCF supernatants of RSM and of the PC dispersion but not in the ultrafiltrates from these UCF supernatants (free of soluble proteins). Simultaneously, alpha(S1-), alpha(S2-), beta- and kappa-caseins were progressively released from the micelles, as seen by electrophoretic analysis. The pressure-induced solubilisation of alpha(S1-) and alpha(S2-)caseins, essentially located in the core of the micelles, suggests that high pressure de-stabilized micelles including their internal structure.