Cross-contamination of Escherichia coli O157:H7 and Listeria monocytogenes in the viable but non-culturable (VBNC) state during washing of leafy greens and the revival during shelf-life.

Some water disinfection treatments, such as chlorine and chlorine dioxide, used in the fresh-cut industry to maintain the microbiological quality of process water (PW), inactivate bacterial cells in the water but they also lead to the induction of an intermediate state between viable and non-viable known as viable but non-culturable (VBNC) state. Viable cells can participate in cross-contamination events but the significance of VBNC cells in PW, transfer to the product and potential resuscitation capacity during storage is unclear. The present study aims to determine first, if VBNC cells present in PW can cross-contaminate leafy greens during washing and secondly its potential revival during shelf-life. Process water characterized by a high chemical oxygen demand, due to the presence of high levels of organic matter, was inoculated with Listeria monocytogenes or Escherichia coli O157:H7. Inoculated PW was then treated for 1 min with chlorine dioxide (3 mg/L) or chlorine (5 mg/L) to generate VBNC cells. Absence of culturable cells was confirmed by plate count and VBNC cells by viability quantitative polymerase chain reaction (v-qPCR) complemented with two dyes, ethidium (EMA) and propidium (PMAxx) monoazide. Cross-contamination of shredded lettuce was demonstrated by monitoring the VBNC cells after washing the product for 1 min in the contaminated PW and during shelf life (15 days at 7 °C). In the case of L. monocytogenes, considering the total concentration of L. monocytogenes VBNC cells present in the PW, only a low proportion of cells were able to cross-contaminate the product during washing. VBNC L. monocytogenes cells were able to resuscitate on the product during shelf life, although levels of cultivable bacteria, close to the limit if detection (0.7 ± 0.0 log CFU/g), were only detected at the end of storage. On the other hand, VBNC cells of E. coli O157:H7 present in PW were not able to cross-contaminate shredded lettuce during washing. Moreover, when shredded lettuce was artificially inoculated with VBNC E. coli O157:H7, resuscitation of the VBNC cells during storage (15 days at 7 °C) was not observed. Based on the results obtained, injured L. monocytogenes cells present in the PW are able to be transferred to the product during washing. If VBNC L. monocytogenes cells present in leafy greens (shredded lettuce and baby spinach), they can resuscitate, although cultivable numbers remained very low. Taking all the results together, it could be concluded that under industrial conditions, VBNC cells can be transferred from water to product during washing, but their capacity to resuscitate in the leafy greens during storage is low.

Incidental findings on brain MRI in a cohort of diabetic patients.

OBJECTIVE: To study the frequency and management of incidental findings in nonenhanced brain MRI of a middle-aged population of type 2 diabetic patients. METHODS: We retrospectively analyzed the results of 289 brain MRI obtained from subjects between 40-75 years recruited from a previous study. Incidental findings were classified into three categories: (1) Vascular findings; (2) neoplastic findings; and (3) others. On the other side, we made a classification of referral findings. To compare our results, we reviewed the prevalence and evidence about management of both incidental and referral findings in other series. RESULTS: We found an overall prevalence of incidental findings of 10.4% (30/289). Incidental findings raised according to age. The most common incidental findings were: 7 vascular (2.4%), 6 calcifications (2.1%), 6 cystic (2.1%) and 5 neoplastic (1.7%) lesions. A percentage of 1.7% (5/289) were referral findings which required further clinical work-up. CONCLUSION: Incidental findings are relatively common in patients with type 2 diabetes. The most frequent are vascular findings, accordance with previous studies. Referral findings are uncommon. Clinical evidence about how to best manage the majority of incidental findings is lacking.

Chemical risks associated with ready-to-eat vegetables: quantitative analysis to estimate formation and/or accumulation of disinfection byproducts during washing.

Fresh produce can become contaminated with disease-causing microorganisms and chemical contaminants at every step of the production and processing chain and in a variety of ways, including through contact with contaminated process water. Water quality is critical to prevent microbial and chemical risks in any of the postharvest and processing operations related to fresh and fresh-cut fruits and vegetables. The wash process requires high volumes of water, which are usually reduced by water reuse. To maintain the microbiological quality of the process water, intervention strategies are needed. Chemical disinfection is the most common method to maintain the microbial quality of process water. However, the use of chemicals leads to the formation/accumulation of disinfection byproducts (DBPs), which can be absorbed by the washed vegetables. This is the case of trihalomethanes (THMs) and chlorates. The presence of high concentrations of DBPs in vegetables has led to an intensive debate on current disinfection practices and how DBPs may enter the food supply chain, becoming a potential health risk for consumers. To assess the risk associated with the formation/accumulation of DBPs in process water, a quantitative analysis was done. Available data have been used to develop mathematical models to predict the formation/accumulation of DBPs (chlorates and THMs) in process water due to the use of chlorine-derived compounds. Preliminary models have been developed, but adjustments are still needed to refine them. The present study contributes more information related to the development of a mathematical model for the accumulation of chlorates and THMs in process water.

Irrigating Lettuce with Wastewater Effluent: Does Disinfection with Chlorine Dioxide Inactivate Viruses?

Reclaimed water obtained from urban wastewater is currently being used as irrigation water in water-scarce regions in Spain. However, wastewater can contain enteric viruses that water reclamation treatment cannot remove or inactivate completely. In the present study, greenhouse-grown baby lettuce ( L.) was irrigated with secondary treatment effluent from a wastewater treatment plant untreated and treated using chlorine dioxide (ClO). The effect of ClO treatment on the physicochemical characteristics and the presence of enteric viruses in irrigation water and lettuce was assessed. The presence of human noroviruses genogroups I and II (NoV GI and NoV GII), and human astroviruses (HAstV), was analyzed by real-time polymerase chain reaction (RT-qPCR). Additionally, to check for the loss of infectivity induced by the disinfection treatment, positive samples were re-analyzed after pretreatment with the intercalating dye PMAxx before RNA extraction and RT-qPCR. There were no significant differences in the proportion of positive samples and the concentration of enteric viruses between treated and untreated reclaimed water without PMAxx pretreatment ( > 0.05). A significantly lower concentration of NoV GI was detected in ClO-treated water when samples were pretreated with PMAxx ( < 0.05), indicating that inactivation was due to the disinfection treatment. Laboratory-scale validation tests indicated the suitability of PMAxx-RT-qPCR for discrimination between potentially infectious and ClO-damaged viruses. Although the applied ClO treatment was not able to significantly reduce the enteric virus load of the secondary effluent from the wastewater treatment plant, none of the lettuce samples analyzed ( = 36) was positive for the presence of NoV or HAstV.

Occurrence of enteric viruses in reclaimed and surface irrigation water: relationship with microbiological and physicochemical indicators.

AIMS: To assess the prevalence of enteric viruses in different irrigation water sources and in the irrigated produce, and the possible links with microbiological and physicochemical water characteristics. METHODS AND RESULTS: The prevalence and levels of Escherichia coli, Norovirus (NoV) genogroup I (GI) and II (GII), as well as Hepatitis A virus were assessed in three types of water: surface water (surface-W), reclaimed water subjected to secondary treatment (secondary-W) and reclaimed water subjected to tertiary treatment (tertiary-W), as well as in zucchini irrigated with these irrigation water sources. Chemical oxygen demand (COD), turbidity, total suspended solids, alkalinity and maximum filterable volume (MFV) were also measured in the water. Higher prevalence of NoV in secondary-W (GI 100%, GII 55·6%) and tertiary-W (GI 91·7%, GII 66·7%) compared with surface-W (GI 58·4%, GII 22·2%) was observed. Nov GI showed positive correlation with E. coli (Spearman's correlation coefficient = 0·68, P < 0·01), and with some physicochemical parameters such as COD (0·52, P < 0·01), turbidity (0·52, P < 0·01) and MFV (0·54, P < 0·01). Escherichia coli and enteric viruses were not detected in zucchini. CONCLUSION: There is a potential risk of contamination of crops with NoV when reclaimed water is used for irrigation. SIGNIFICANCE AND IMPACT OF THE STUDY: Increase the knowledge on the prevalence of enteric viruses in different irrigation water sources, and its consequences for fresh produce safety.

Suitability of different Escherichia coli enumeration techniques to assess the microbial quality of different irrigation water sources.

The use of fecal indicators such as Escherichia coli has been proposed as a potential tool to characterize microbial contamination of irrigation water. Recently, not only the type of microbial indicator but also the methodologies used for enumeration have been called into question. The goal of this study was to assess the microbial quality of different water sources for irrigation of zucchini plants by using E. coli as an indicator of fecal contamination and the occurrence of foodborne pathogens. Three water sources were evaluated including reclaimed secondary treated water (RW-2), reclaimed tertiary UV-C treated water (RW-3) and surface water (SW). The suitability of two E. coli quantification techniques (plate count and qPCR) was examined for irrigation water and fresh produce. E. coli levels using qPCR assay were significantly higher than that obtained by plate count in all samples of irrigation water and fresh produce. The microbial quality of water samples from RW-2 was well predicted by qPCR, as the presence of foodborne pathogens were positively correlated with high E. coli levels. However, differences in the water characteristics influenced the suitability of qPCR as a tool to predict potential contamination in irrigation water. No significant differences were obtained between the number of cells of E. coli from RW-2 and RW-3, probably due to the fact that qPCR assay cannot distinguish between viable and dead cells. These results indicated that the selection of the most suitable technique for enumeration of indicator microorganisms able to predict potential presence of fecal contamination might be influenced by the water characteristics.

The determination of copeptin levels helps management decisions among transient ischaemic attack patients.

BACKGROUND: Most approaches to transient ischaemic attack (TIA) triage use clinical scores and vascular imaging; however, some biomarkers have been suggested to improve the prognosis of TIA patients. METHODS: Serum levels of copeptin, adiponectin, neopterin, neuron-specific enolase, high-sensitivity C-reactive protein, IL-6, N-terminal pro-B-type natriuretic peptide, S100ß, tumour necrosis factor-alpha and IL-1α as well as clinical characteristics were assessed on consecutive TIA patients during the first 24 h of the onset of symptoms. RESULTS: Among 237 consecutive TIA patients, 12 patients (5%) had a stroke within 7 days and 15 (6%) within 90 days. Among all candidate biomarkers analysed, only copeptin was significantly increased in patients with stroke recurrence (SR) within 7 days (P = 0.026) but not within 90 days. A cut-off point of 13.8 pmol/l was established with a great predictive negative value (97.4%). Large artery atherosclerosis (LAA) [hazard ratio (HR) 12.7, 95% CI 3.2-50.1, P < 0.001] and copeptin levels ≥13.8 pmol/l (HR 3.9, 95% CI 1.01-14.4, P = 0.039) were independent predictors of SR at the 7-day follow-up. LAA was the only predictor of 90-day SR (HR 7.4, 95% CI 2.5-21.6, P < 0.001). ABCD3I was associated with 7- and 90-day SRs (P = 0.025 and P = 0.034, respectively). The association between copeptin levels and LAA had a diagnostic accuracy of 90.3%. CONCLUSIONS: Serum copeptin could be an important prognostic biomarker to guide management decisions among TIA patients. Therefore, TIA patients with copeptin levels below 13.8 pmol/l and without LAA have an insignificant risk of 7-day SR and could be managed on an outpatient basis.

Assessment of microbial risk factors and impact of meteorological conditions during production of baby spinach in the Southeast of Spain.

There is a timely need to evaluate the effect agricultural factors and meteorological conditions on fresh produce contamination. This study evaluated those risk factors and described, for the first time, the distribution of indicator microorganisms (Escherichia coli, Enterococcus, coliforms, and Enterobacteriaceae) and the prevalence of foodborne pathogens (Enterohaemorrhagic E. coli, Listeria monocytogenes and Salmonella spp.) in baby spinach grown in the Southeast of Spain. A longitudinal study was conducted on three farms (2011-2013). Results obtained for E. coli highlighted soil and irrigation water as important factors affecting the microbial safety of baby spinach. Significant differences in the proportion of E. coli positive samples were found between treated (46.1%) and untreated (100%) irrigation water. However, the microbial quality of irrigation water didn't affect E. coli prevalence in produce. All E. coli positive spinach samples were detected at the highest observed temperature range suggesting that ambient temperature affects the probability and extent of spinach contamination. Salmonella spp. was detected by RT-PCR in manure, soil, irrigation water and baby spinach but only two of them (manure and irrigation water) were confirmed by isolation in culture media. Salmonella RT-PCR positive samples showed higher levels of E. coli than Salmonella negative samples. This preliminary finding supports recent identification of E. coli as a suitable parameter for the hygiene criterion at the primary production of leafy greens.

Prediction of myocardial infarction in patients with transient ischaemic attack.

BACKGROUND: Determinants of risk of myocardial infarction (MI) after transient ischaemic attack (TIA) are not well defined. The aim of our study was to determine the risk and risk factors for MI after TIA. METHODS: We prospectively recruited patients within 24 h of transient ischaemic cerebrovascular events between October 2006 and January 2013. A total of 628 TIA patients were followed for six months or more. MI and stroke recurrence (SR) were recorded. The duration and typology of clinical symptoms, vascular risk factors and aetiological work-ups were prospectively recorded and established prognostic scores (ABCD2, ABCD2I, ABCD3I, Essen Stroke Risk Score, California Risk Score and Stroke Prognosis Instrument) were calculated. RESULTS: Twenty-eight (4.5%) MI and 68 (11.0%) recurrent strokes occurred during a median follow-up period of 31.2 months (16.1-44.9). In Cox proportional hazards multivariate analyses, we identify previous coronary heart disease (CHD) (hazard ratio [HR] 5.65, 95% confidence interval [CI] 2.45-13.04, P < 0.001) and sex male (HR 2.72, 95% CI 1.02-7.30, P = 0.046) as independent predictors of MI. Discrimination for the prognostic scores only ranged from 0.60 to 0.71. The incidence of MI did not vary among the different aetiological subtypes. Positive diffusion weighted imaging (DWI) (7.5% vs 2.5%, P = 0.007), and ECG abnormalities (Q wave or ST-T wave changes) (13.6% vs 3.6%, P = 0.001) were associated to MI. CONCLUSION: According to our results, discrimination was poor for all previous risk prediction models evaluated. Variables such as previous CHD, male sex, DWI and ECG abnormalities should be considered in new prediction models.

N-terminal pro-brain natriuretic peptide level determined at different times identifies transient ischaemic attack patients with atrial fibrillation.

BACKGROUND AND PURPOSE: The etiological classification of patients with transient ischaemic attack (TIA) is a difficult endeavor and the use of serum biomarkers could improve the diagnostic accuracy. The aim of this study was to correlate atrial fibrillation, the main cardioembolic etiology (CE), with different serum biomarkers measured in consecutive TIA patients. METHODS: The concentrations of interleukin-6 (IL-6), tumor necrosis factor-alpha, neuron-specific enolase, high-sensitivity C-reactive protein, IL-1-α and the N-terminal pro-B type natriuretic peptide (NT-proBNP) were quantified in the serum of 140 patients with TIA and 44 non-stroke subjects. Measurements were performed at different times throughout evolution: within 24 h of symptoms onset and at days 7 and 90. RESULTS: With the exception of IL-6, all biomarkers were higher in TIA patients than in controls. NT-proBNP was significantly related to the presence or new diagnosis of AF at all time points analyzed. Furthermore, the baseline NT-proBNP level was significantly higher than values at the 7-day and 90-day follow-up. For this reason, different cut-off values were obtained at different times: 313 pg/ml at baseline [odds ratio (OR) = 18.99, P < 0.001], 181 pg/ml at 7 days (OR = 11.4, P = 0.001) and 174 pg/ml (OR = 8.46, P < 0.001) at 90 days. CONCLUSION: High levels of NT-proBNP determined during the first 3 months after a TIA were associated with AF. Consequently, this biomarker may be useful to reclassify undetermined TIA patients as having disease of CE.

Percepción del paciente mayor polimedicado sobre el rol del farmacéutico comunitario / Perception of the polymedicated elderly patient about the community pharmacist’s role

Introducción: Actualmente casi un 80% del gasto sanitario español corresponde a las personas mayores de 65 años que, debido a su pluripatología, se convierten en pacientes polimedicados. El farmacéutico comunitario es uno de los agentes de salud que puede ayudar a controlar este gasto y mejorar la calidad del uso de los medicamentos debido a su cercanía y accesibilidad, aunque actualmente sólo se le reconozca un rol dispensador de medicamentos. Objetivos: Explorar la percepción que tiene el paciente mayor polimedicado sobre el rol del farmacéutico. Material y métodos: Se utilizó un método exploratorio basado en la investigación cualitativa, usando como medio de obtención de información la entrevista semiestructurada y como marcos teóricos la Teoría del Rol complementada con la Teoría de las Creencias en Salud. Resultados: Los pacientes mayores polimedicados identifi can con el rol de profesional sanitario principalmente a su médico de cabecera y al personal de enfermería, relegando al farmacéutico a funciones de índole menor y, sobre todo, con un rol dispensador de medicamentos y productos sanitarios, aunque resaltan la gran accesibilidad y la confianza que tienen con este profesional. Conclusión: Los pacientes mayores polimedicados en España esperan del farmacéutico un buen trato y un rol dispensador. Esto puede deberse a la baja expectativa que se tiene del farmacéutico para realizar servicios profesionales distintos de los clásicos (dispensación). Los farmacéuticos deberían empezar a cambiar su rol actual implantando servicios para que la percepción del paciente cambie(AU)

Introduction: Nowadays, almost 80 percent of the health expenditure in Spain corresponds to people over 65 years of age as a result of their multiple pathology, have become polymedicated patients. Community pharmacists are community health workers who can help to control this expense and improve the quality of use of the medications due to the proximity and approachability although nowadays they are only known in their role as the medication dispenser. Materials and methods: An exploratory method was used, based on the qualitative research and semi-structured interviews were conducted as a means to get information, combining both the Role Theory and Health Belief Model as theoretical frameworks. Results: Polymedicated elderly patients tend to associate the role of health care professionals with their general practitioners and community nurses, giving less priority to the function of the pharmacists. Pharmacists are often relegated to a minor role and particularly to the supplying of drugs and healthcare products, that is, their dispensary role. However, patients are likely to highlight the great accessibility and trust they have towards these professionals. Conclusion: The polymedicated elderly patients expect from the pharmacist in Spain is a good rapport with their patients, besides a dispensary role. This could be a consequence of the low expectations that they have of the pharmacists concerning the services they provide. Pharmacists are assumed not to offer other services different to the classical services (dispensing). They should start changing their current role by introducing new services in order to change the perception that patients have about them(AU)

Patterns of diffusion-weighted magnetic resonance imaging associated with etiology improve the accuracy of prognosis after transient ischaemic attack.

OBJECTIVE: diffusion-weighted magnetic resonance imaging (DWI) is a sensitive diagnostic tool for detecting acute ischaemic lesions in patients with transient ischaemic attacks (TIAs). The additional predictive value of DWI lesion patterns is not well known. METHODS: two hundred and fifty-four consecutive patients with TIA underwent DWI within 7 days of symptom onset. The presence and pattern of acute ischaemic lesions were related to clinical features, etiology, and stroke recurrence at seven- and 90-day follow-up. RESULTS: diffusion-weighted images abnormalities were identified in 117 (46.1%) patients. The distribution of DWI lesions was cortical, 31; subcortical, 32; scattered lesions in one arterial territory (SPOT) 42; and in multiple areas, 12. SPOT were significantly associated with motor weakness, large-artery atherosclerosis (LAA), and the cardioembolic subtype of TIA. Single cortical lesions were also associated with cardioembolism, whereas subcortical acute lesions were associated with recurrent episodes, dysarthria, and motor weakness. During follow-up, seven patients had a stroke within 7 days (2.8%, 95% CI 2.9-6.4%), and 12 had a stroke within 3 months (4.7%%, 95% CI 2.9-6.4%). In the Cox logistic regression model, the combination of LAA and positive DWI remained as independent predictors of stroke recurrence at 90-day follow-up (HR 5.78, 95 CI 1.74-19.21, P = 0.004). CONCLUSION: according to our results, MRI, including DWI, should be considered a preferred diagnostic test when investigating patients with potential TIAs. The combination of neuroimaging and vascular information could improve prognostic accuracy in patients with TIA.

Potential microbial risk factors related to soil amendments and irrigation water of potato crops.

AIMS: This study assesses the potential microbial risk factors related to the use of soil amendments and irrigation water on potato crops, cultivated in one traditional and two intensive farms during two harvest seasons. METHODS AND RESULTS: The natural microbiota and potentially pathogenic micro-organisms were evaluated in the soil amendment, irrigation water, soil and produce. Uncomposted amendments and residual and creek water samples showed the highest microbial counts. The microbial load of potatoes harvested in spring was similar among the tested farms despite the diverse microbial levels of Listeria spp. and faecal coliforms in the potential risk sources. However, differences in total coliform load of potato were found between farms cultivated in the autumn. Immunochromatographic rapid tests and the BAM's reference method (Bacteriological Analytical Manual; AOAC International) were used to detect Escherichia coli O157:H7 from the potential risk sources and produce. Confirmation of the positive results by polymerase chain reaction procedures showed that the immunochromatographic assay was not reliable as it led to false-positive results. CONCLUSIONS: The potentially pathogenic micro-organisms of soil amendment, irrigation water and soil samples changed with the harvest seasons and the use of different agricultural practices. However, the microbial load of the produce was not always influenced by these risk sources. Improvements in environmental sample preparation are needed to avoid interferences in the use of immunochromatographic rapid tests. SIGNIFICANCE AND IMPACT OF THE STUDY: The potential microbial risk sources of fresh produce should be regularly controlled using reliable detection methods to guarantee their microbial safety.

Antioxidant capacity and phenolic content of spinach as affected by genetics and maturation.

Spinach leaves harvested at three maturity stages from eight commercial cultivars (CC) and eight advanced breeding lines (ABL) were evaluated for oxygen radical absorbing capacity (ORAC), total phenolics, and flavonoid composition and content. ABL had higher levels of total phenolics, total flavonoids, and ORAC than CC. Midmaturity spinach leaves had higher levels of total phenolics, total flavonoids, and antioxidant capacity than immature and mature leaves. The contents of individual flavonoids varied in response to maturation, with the predominant glucuronated flavones decreasing and patuletin and spinacetin derivatives increasing. Both total phenolics and total flavonoids correlated well with ORAC (r(xy)() = 0.78 and 0.81, respectively) demonstrating that flavonoids were major contributors to antioxidant capacity. Our results indicate that spinach genotypes should be harvested at the midmaturity stage for consumers to benefit from elevated levels of health promoting flavonoids present in the leaves. Additionally, plant breeders can select for increased phenolic content to increase antioxidant capacity of spinach genotypes.

Heavy metals in the vicinity of a chlor-alkali factory in the Upper Negro River ecosystem, Northern Patagonia, Argentina.

A study on heavy metal contents was performed in sediments and biota of the Upper Negro River (Alto Valle) aquatic system, Northern Patagonia, Argentina. The irrigation system of the Neuquén and Negro Rivers runs alongside these rivers for 150 km, supporting intensive agricultural and economical activities, mainly related to fruit production. A mercury cell chlor-alkali factory operated between 1951 and 1995. Close attention was given to the surroundings of the plant, located next to the Main Irrigation Channel, and to the PII drainage channel which received the plant's effluents between 1951 and 1979. From 1979 until its closure, the effluents were pumped above a ravine to a series of evaporation and decantation pools. Mercury and other heavy metals and metalloids (Ag, As, Ba, Co, Cr, Cs, Ni, Sb, Se, U and Zn) contents were measured for bottom sediments of the river and irrigation and drainage channels, for two widespread species of macrophytes (Potamogeton pectinatus and Myriophyllum brasiliensi), and for liver and muscle of native fish Odontesthes microlepidotus. River bed sediments show no evidence of heavy metal accumulation, however, biota might indicate that contaminants are entering the rivers. Mercury was the only element accumulated in the Main Irrigation channel sediments, the highest contents occurring in the surroundings of the nowadays shut-down chlor-alkali plant, returning to background values approximately 40 km downstream the plant. At the plant site, sediments from the center of the channel showed a decrease in Hg content in the upper 10 cm layer, ranging from 0.8 to 3.4 microg g(-1), and from 2.8 to 13.7 microg g(-1) in the next 10 cm lower layer. Conversely, the PII drainage channel sediments showed accumulation of Hg (2-4 microg g(-1)), distributed uniformly at different depths and along the channel, until its mouth at Negro river. Mercury contents of macrophytes downstream the chlor-alkali plant are higher than the baseline for the area, and macrophytes and fish liver from the PII drainage channel present the highest content in this element. The drainage channel system showed different degrees of impact, those channels flowing through densely populated areas being the most affected.

Antioxidant capacity and phenolic content of spinach as affected by genetics and growing season.

Total phenolic content and antioxidant capacity of 11 commercial cultivars and 15 advanced breeding lines of spinach were determined over two growing seasons known to vary in biotic and abiotic stresses. Flavonoid composition and content of fall-grown commercial cultivars and advanced breeding lines were also determined. Over-winter spinach, which was planted in late fall and harvested in the spring, had much higher levels of total phenolics and antioxidant capacity than spinach planted in early fall and harvested in late fall, indicating that growing conditions, as well as biotic and abiotic stresses, influenced phenolic metabolism. Genotype also appeared to play an important role in affecting phenolic metabolism and antioxidant capacity in spinach. Advanced breeding lines of spinach, which show increased disease resistance, had higher levels of total phenolics, individual and total flavonoids, and antioxidant capacity than commercial cultivars. Our results indicate that plant breeders can select for increased phenolic content to increase antioxidant capacity in spinach cultivars, or the crops can be grown in different seasons or under certain stress conditions to elevate levels of antioxidants.

HPLC-DAD-ESIMS analysis of phenolic compounds in nectarines, peaches, and plums.

The phenolic compounds of 25 peach, nectarine, and plum cultivars were studied and quantified by HPLC-DAD-ESIMS. Hydroxycinnamates, procyanidins, flavonols, and anthocyanins were detected and quantified. White and yellow flesh nectarines and peaches, and yellow and red plums, were analyzed at two different maturity stages with consideration of both peel and flesh tissues. HPLC-MS analyses allowed the identification of procyanidin dimers of the B- and A-types, as well as the presence of procyanidin trimers in plums. As a general rule, the peel tissues contained higher amounts of phenolics, and anthocyanins and flavonols were almost exclusively located in this tissue. No clear differences in the phenolic content of nectarines and peaches were detected or between white flesh and yellow flesh cultivars. There was no clear trend in phenolic content with ripening of the different cultivars. Some cultivars, however, had a very high phenolic content. For example, the white flesh nectarine cultivar Brite Pearl (350-460 mg/kg hydroxycinnamates and 430-550 mg/kg procyanidins in flesh) and the yellow flesh cv. Red Jim (180-190 mg/kg hydroxycinnamates and 210-330 mg/kg procyanidins in flesh), contained 10 times more phenolics than cultivars such as Fire Pearl (38-50 mg/kg hydroxycinnamates and 23-30 mg/kg procyanidins in flesh). Among white flesh peaches, cultivars Snow King (300-320 mg/kg hydroxycinnamates and 660-695 mg/kg procyanidins in flesh) and Snow Giant (125-130 mg/kg hydroxycinnamates and 520-540 mg/kg procyanidins in flesh) showed the highest content. The plum cultivars Black Beaut and Angeleno were especially rich in phenolics.

In vitro availability of flavonoids and other phenolics in orange juice.

Hand-squeezed navel orange juice contains 839 mg/L phenolics, including flavanones, flavones, and hydroxycinnamic acid derivatives. The flavanones are the main phenolics in the soluble fraction (648.6 mg/L) and are also present in the cloud fraction (104.8 mg/L). During refrigerated storage of fresh juice (4 degrees C), 50% of the soluble flavanones precipitate and integrate into the cloud fraction. Commercial orange juices contain only 81-200 mg/L soluble flavanones (15-33%) and the content in the cloud is higher (206-644 mg/L) (62-85%), showing that during industrial processing and storage the soluble flavanones precipitate and are included in the cloud. An in vitro simulation of orange juice digestion shows that a serving of fresh orange juice (240 mL) provides 9.7 mg of soluble hesperidin (4'-methoxy-3',5,7-trihydroxyflavanone-7-rutinoside) and 4.7 mg of the C-glycosylflavone vicenin 2 (apigenin, 6,8-di-C-glucoside) for freshly squeezed orange juice, whereas pasteurized commercial juices provide 3.7 mg of soluble hesperidin and a higher amount of vicenin 2 (6.3 mg). This means that although orange juice is a very rich source of flavanones, only a limited quantity is soluble, and this might affect availability for absorption (11-36% of the soluble flavanones, depending on the juice). The flavanones precipitated in the cloud are not available for absorption and are partly transformed to the corresponding chalcones during the pancreatin-bile digestion.