Inactivation kinetics of Escherichia coli in cranberry juice during multistage treatment by electric fields.

The inactivation of Escherichia coli inoculated in cranberry juice by processing with radio frequency electric fields was studied. E. coli ATCC 35218 was chosen among three non-pathogenic strains based on its ability to survive in low pH cranberry juice. Studies were conducted by measuring the survival population when changing the electric field strength between 2.2 and 13.2 kV cm-1, number of treatment stages from 1 to 6 and flow rates between 13 and 25 L h-1 at moderate temperatures of 20, 30 and 40 °C. A minimum inactivation of 5-log reduction, as requested by the Food and Drug Administration (FDA), can be achieved by increasing the number of treatment stages, temperature or both. At 40 °C and 6 treatment stages, 6.57 ±â€¯0.02 log CFU ml-1 reduction in the initial population of E. coli (ATCC 35218) was obtained. At a constant electric field, increasing the temperature produced higher microbial inactivation, consuming lower radio frequency energy input, than increasing the number of treatment stages. Furthermore, a primary model that accounts for the combined effect of time and electric field is proposed. The model represented the sigmoidal curve composed of shoulder, log-linear and tailing sections as observed when changing electric fields. A secondary model that accounts for the effect of temperature and flow rate on the primary model constants is also proposed. The combined primary and secondary models were found to fit the data well with a high coefficient of determination (R2 = 0.965). The proposed model can be extended to kinetic models for pulsed electric fields.

Phenolic compounds of the Australian native herb Prostanthera rotundifolia and their biological activities.

The chemical identity and bioactivities of phenolic components of the Australian native herb Prostanthera rotundifolia were studied. Phenolic compounds were extracted with 80% (v/v) aqueous methanol and purified by liquid chromatography. The antioxidant capacity of the extract and its inhibiting activity against α-glucosidase, pancreatic lipase and hyaluronidase were determined. Phenolic compounds were identified by a combination of HPLC-PDA, LC-high resolution MS (LC-HRMS), LC-tandem MS (LC-MS/MS) and nuclear magnetic resonance (NMR) spectroscopy. Compared to spearmint, mint bush showed comparable antioxidant capacity, stronger inhibitory activity on pancreatic lipase and comparable and lower activity on α-glucosidase and hyaluronidase, respectively. Major compounds identified were verbascoside (48.8%), 4-methoxycinnamic acid (36.4%), p-coumaric acid glucose ester (9.2%) and 1-O-ß-d-glucopyranosyl sinapate (5.6%), while caffeic acid, p-coumaric acid, hesperidin and naringenin were present in trace quantities. 4-Methoxycinnamic acid, p-coumaric acid glucose ester and 1-O-ß-d-glucopyranosyl sinapate were identified for the first time in the genus of Prostanthera.

Identification and quantification of phenolics in Australian native mint (Mentha australis R. Br.).

Australian native mints have traditionally been used by the aboriginal people for natural remedies; however, their bioactive components have not been studied. Antioxidant capacity and composition of phenolic compounds of Mentha australis R. Br., Lamiaceae were investigated for the first time. Phenolic compounds were analyzed by HPLC photodiode array detector, liquid chromatography high resolution mass spectrometry, tandem mass spectrometry and nuclear magnetic resonance spectroscopy. Aqueous methanolic extract of the mint exhibited comparable antioxidant capacity to the common spearmint. Major compounds identified in the extract were rosmarinic acid (160.4 ± 0.85 µg mg(-1)purified extract), neoponcirin (145.0 ± 0.42 µg gallic acid equivalent(GAE) mg(-1)), narirutin (30.3 ± 0.02 µg GAE mg(-1)), chlorogenic acid (15.4 ± 0.05 µg mg(-1)) and biochanin A (9.6 ± 0.06 µg GAE mg(-1)), while minor compounds were caffeic acid, apigenin, hesperetin and naringenin. Neoponcirin and biochanin A were identified for the first time in the Mentha genus.

Clinical profile of young children with mental retardation and developmental delay in Hong Kong.

OBJECTIVE: To report the clinical profile of children with mental retardation and developmental delay diagnosed by the Child Assessment Service. DESIGN: Retrospective study. SETTING: Child Assessment Service, Department of Health, Hong Kong. PARTICIPANTS: Data pertaining to the children with mental retardation and developmental delay were drawn from an in-house clinical information system in the year 2004. MAIN OUTCOME MEASURES: Clinical profiles including: sources, reasons and age of referral, diagnosis, gender ratio, co-morbidities, and socio-economic background. RESULTS: In 2004, 23% (1463 of 6439) of Child Assessment Service referrals were diagnosed to have mental retardation or developmental delay. The Family Health Service was the major source of referral (64%). The majority (93%) of children were referred before the age of 6 years. The most common reason for referral was language delay (39%). More boys were affected (3 boys: 1 girl). The two most common co-morbidities were autistic spectrum disorders (33% in mental retardation and 19% in developmental delay) and discrepant language delay (17% in mental retardation and 47% in developmental delay). The socio-economic status of these families was higher than those in the general population. CONCLUSION: The data presented here provide information on the descriptive epidemiology of mental retardation and developmental delay among Hong Kong children. Since mental retardation and developmental delay are common developmental disabilities in Hong Kong, public health education to promote and ensure early screening and identification of cases is an important prelude to early training and guidance for families with children having these conditions.

Niacin deficiency causes oxidative stress in rat bone marrow cells but not through decreased NADPH or glutathione status.

Niacin (vitamin B(3)), in the form of NADPH, is required for the regeneration of glutathione (GSH), which is the substrate of GSH peroxidase. In this study, we examined the effect of dietary niacin deficiency on protein and DNA oxidation in bone marrow cells of Long-Evans rats. Western blotting was used to measure 2,4-dinitrophenylhydrazine-reactive protein carbonyl products, and the Biotrin OxyDNA method was used to measure 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG). The levels of both protein carbonyls and 8-oxodG were increased by 50% in niacin-deficient bone marrow cells. To examine whether this oxidant damage involves altered metabolism of pyridine nucleotides and glutathione, both oxidized and reduced forms of pyridine nucleotides (NAD(+), NADH, NADP(+), NADPH) and glutathione (GSSG and GSH) were quantified in total and nucleated bone marrow cells. NAD and NADP(+) levels were decreased 80% and 22%, respectively, by niacin deficiency. NADPH and GSH were not depleted by niacin deficiency, showing that oxidant injury was not due directly to impairment of this pathway. Oxidative stress, of uncertain etiology, may play a role in the observed genomic instability and sensitivity to leukemogenesis in bone marrow cells during niacin deficiency.

Morphological and biochemical analysis of Rac1 in three-dimensional epithelial cell cultures.

Rho GTPases are critical regulators of epithelial morphogenesis. A powerful means to investigate their function is three-dimensional (3D) cell culture, which mimics the architecture of epithelia in vivo. However, the nature of 3D culture requires specialized techniques for morphological and biochemical analyses. Here, we describe protocols for 3D culture studies with Madin-Darby Canine Kidney (MDCK) epithelial cells: establishing cultures, immunostaining, and expressing, detecting, and assaying Rho proteins. These protocols enable the regulation of epithelial morphogenesis to be explored at a detailed molecular level.

Formation of multicellular epithelial structures.

The kidney is primarily comprised of highly polarized epithelial cells. Much has been learned recently about the mechanisms of epithelial polarization. However, in most experimental systems the orientation of this polarity is determined by external cues, such as growth of epithelial cells on a filter support. When Madin-Darby canine kidney (MDCK) cells are grown instead in a three-dimensional (3D) collagen gel, the cells form hollow cysts lined by a monolayer of epithelial cells, with their apical surfaces all facing the central lumen. We have found that expression of a dominant-negative (DN) form of the small GTPase Rac1 causes an inversion of epithelial polarity, such that the apical surface of the cells instead faces the periphery of the cyst. This indicates that the establishment of polarity and the orientation of polarity can be experimentally separated by growing cells in a 3D collagen gel, where there is no filter support to provide an external cue for orientation. DN Rac1 causes a defect in the assembly of laminin into its normal basement membrane network, and addition of a high concentration of exogenous laminin rescues the inversion of polarity caused by DN Rac1.

ERK and MMPs sequentially regulate distinct stages of epithelial tubule development.

Epithelial cells undergo tubulogenesis in response to morphogens such as hepatocyte growth factor (HGF). To organize into tubules, cells must execute a complex series of morphogenetic events; however, the mechanisms that underlie the timing and sequence of these events are poorly understood. Here, we show that downstream effectors of HGF coordinately regulate successive stages of tubulogenesis. Activation of extracellular-regulated kinase (ERK) is necessary and sufficient for the initial stage, during which cells depolarize and migrate. ERK becomes dispensable for the latter stage, during which cells repolarize and differentiate. Conversely, the activity of matrix metalloproteases (MMPs) is essential for the late stage but not the initial stage. Thus, ERK and MMPs define two regulatory subprograms that act in sequence. By inducing these reciprocal signals, HGF directs the morphogenetic progression of tubule development.

Analysis of membrane traffic in polarized epithelial cells.

Spatial asymmetry is fundamental to the structure and function of most eukaryotic cells. A basic aspect of this polarity is that the cell's plasma membrane is divided into discrete domains. The best studied and simplest example of this occurs in epithelial cells, which line exposed body surfaces. Epithelial cells use two pathways to send proteins to the cell surface. Newly made proteins can travel directly from the trans-Golgi network (TGN) to either the apical or basolateral surface. Alternatively, proteins can be sent to the basolateral surface and then endocytosed and transcytosed to the apical surface. Epithelial cells grown on porous filters adopt a typical polarized morphology; transfected epithelial cells can be used to biosynthetically characterize the trafficking patterns of a given protein. These cells can also be used to study delivery to a particular surface and to localize the protein by immunofluorescence.