P.D.G Wilson

Food structure and microbial growth

Abstract Many foods posses complex microstructures. The safety and quality of foods rely on the ability to control microbial growth in each microscopic location. This article summarizes the rationale and early results of a research programme aimed at predicting microbial growth within these structures using physicochemical techniques.

Simultaneous time-varying systemic appearance of oral and hepatic glucose in adults monitored with stable isotopes

The rates (and extent) of appearance of glucose in arterialized plasma from an oral glucose load and from liver (RaO, RaH) can be estimated in humans using radioisotopes, but estimates vary among laboratories. We investigated the use of stable isotopes and undertook 22 primed intravenous infusions of D-[6,6-2H2]glucose with an oral load including D-[13C6]glucose in healthy humans. The effective glucose pool volume (VS) had a lower limit of 230 ml/kg body weight (cf. 130 ml/kg commonly assumed). This VS in Steeles one-compartment model of glucose kinetics gave a systemic appearance from a 50-g oral glucose load per 70 kg body weight of 96 +/- 3% of that ingested, which compared with a theoretical value of approximately 95%. Maris two-compartment model gave 100 +/- 3%. The two models gave practically identical RaO and RaH at each point in time and a plateau in the cumulative RaO when absorption was complete. Less than 3% of 13C was recycled to [13C3]glucose, suggesting that recycling errors were practically negligible in this study. Causes of variation among laboratories are identified. We conclude that stable isotopes provide a reliable and safe alternative to radioactive isotopes in these studies.The rates (and extent) of appearance of glucose in arterialized plasma from an oral glucose load and from liver (RaO, RaH) can be estimated in humans using radioisotopes, but estimates vary among laboratories. We investigated the use of stable isotopes and undertook 22 primed intravenous infusions ofd-[6,6-2H2]glucose with an oral load includingd-[13C6]glucose in healthy humans. The effective glucose pool volume (VS) had a lower limit of 230 ml/kg body weight (cf. 130 ml/kg commonly assumed). This VS in Steeles one-compartment model of glucose kinetics gave a systemic appearance from a 50-g oral glucose load per 70 kg body weight of 96 ± 3% of that ingested, which compared with a theoretical value of ∼95%. Maris two-compartment model gave 100 ± 3%. The two models gave practically identical RaO and RaH at each point in time and a plateau in the cumulative RaO when absorption was complete. Less than 3% of13C was recycled to [13C3]glucose, suggesting that recycling errors were practically negligible in this study. Causes of variation among laboratories are identified. We conclude that stable isotopes provide a reliable and safe alternative to radioactive isotopes in these studies.

The effects of cell immobilization, pH and sucrose on the growth of Listeria monocytogenes Scott A at 10°C

A study into the growth of Listeria monocytogenes Scott A investigated the effects of cell immobilization, varying sucrose concentrations (as a humectant) and alteration of initial pH at 10°C. Three conclusions were drawn from this study. Firstly, it was found that the growth rate of the organism was affected by immobilization, increasing sucrose concentrations (up to 59% w/v) and the lowering of the pH (pH 7.0 to 4.6). Secondly it was found that the lower limits of growth, in terms of pH and water activity, were different for immobilized and planktonic cultures. Lastly, it was observed that the viable population increased when planktonic cells were incubated at 10°C in combination with either low pH (pH 4.6 and 4.8) or high sucrose concentrations (59% w/v). However, this was not observed with immobilized cells, which failed to initiate growth when incubated under these conditions. Such influences of immobilization on the effects of food preservation methods are an important consideration in the predictive modelling of the survival and growth of bacteria in foods.

Microgradients in bacterial colonies : use of fluorescence ratio imaging : a non-invasive technique

Fluorescence ratio imaging is a non-invasive technique for studying the formation of microgradients in immobilised bacterial colonies. These gradients can be quantified easily when combined with the gel cassette system designed at the Institute of Food Research, Norwich, UK. Colonies of Lactobacillus curvatus were observed using this technique and relevant pH gradients were present when the colonies reached a diameter of about 100 microm. These pH gradients were due to production of lactic acid by L. curvatus cells in the colonies. The spatial resolution of the images was about 1.5 microm (scale of bacterial cells) and therefore very suitable for observing local effects in colonies which ranged in sizes from 1 to 500 microm.

Weak acids: dissociation in complex buffering systems and partitioning into oils

A method is developed to predict pH changes in complex buffering systems resulting from the addition of weak acids. Such systems are often encountered in foods and microbiological growth media which contain proteins and proteinaceous digests. The ‘buffering capacity’ of the system is characterised by a strong acid titration curve, and the known dissociation constants of weak acids are used to predict pH changes resulting from the addition of either one or a mixture of weak acids to the system. Predictions by the method show close agreement with experimental data. The familiar Henderson–Hasselbalch equation describing the dissociation of weak acids is modified to account for partitioning of undissociated acid into an oil phase. The equation may be used in conjunction with the pH prediction method to calculate both the pH and the concentration of undissociated weak acid in the aqueous phase of an emulsion containing complex buffering species. The equations should find application wherever weak acids are used for microbiological control in complex systems such as foods and pharmaceutical preparations, for predicting pH changes resulting from microbial growth during the production of fermented food commodities (eg cheese and yoghurt) or during food spoilage, and in biotechnological processes ranging from industrial bioreactors to agricultural silage production. © 2000 Society of Chemical Industry

Splanchnic retention of intraduodenal and intrajejunal glucose in healthy adults

Estimates of the spanchnic retention and appearance in the systemic circulation of orally administered glucose vary among laboratories even after recently identified sources of error have been accounted for [Livesey, G., P. D. G. Wilson, J. R. Dainty, J. C. Brown, R. M. Faulks, M. A. Roe, T. A. Newman, J. Eagles, F. A. Mellon, and R. Greenwood. Am. J. Physiol. 275 ( Endocrinol. Metab. 38): E717-E728, 1998]. We questioned whether, in healthy humans,d-glucose delivered intraluminally to the midjejunum appeared systemically as extensively as that delivered intraduodenally. Subjects were infused over a period of 90 min with 50 g of glucose in 1 liter of isotonic saline (incorporating 0.5 gd-[13C6]glucose) per 70 kg of body weight. Infusions were via enteral tubes terminating ∼15 and 100 cm postpylorus. The systemic appearance of glucose was monitored by means of a primed-continuous intravenous infusion ofd-[6,6-2H2]glucose. Whereas 98 ± 2% ( n = 7) of the duodenally infused glucose appeared in the systemic circulation, only 35 ± 9% ( n = 7) of midjejunally infused glucose did so, implying that 65 ± 9% was retained in the splanchnic bed. Either glucose was less efficiently absorbed at the midintestinal site or hepatic glucose sequestration was increased 10-fold, or both. The proximal intestine plays a key role in the delivery of glucose to the systemic circulation, and the distal intestine potentially delivers more glucose to the liver.Estimates of the spanchnic retention and appearance in the systemic circulation of orally administered glucose vary among laboratories even after recently identified sources of error have been accounted for [Livesey, G., P. D. G. Wilson, J. R. Dainty, J. C. Brown, R. M. Faulks, M. A. Roe, T. A. Newman, J. Eagles, F. A. Mellon, and R. Greenwood. Am. J. Physiol. 275 (Endocrinol. Metab. 38): E717-E728, 1998]. We questioned whether, in healthy humans, D-glucose delivered intraluminally to the midjejunum appeared systemically as extensively as that delivered intraduodenally. Subjects were infused over a period of 90 min with 50 g of glucose in 1 liter of isotonic saline (incorporating 0.5 g D-[13C6]glucose) per 70 kg of body weight. Infusions were via enteral tubes terminating approximately 15 and 100 cm postpylorus. The systemic appearance of glucose was monitored by means of a primed-continuous intravenous infusion of D-[6,6-2H2]glucose. Whereas 98 +/- 2% (n = 7) of the duodenally infused glucose appeared in the systemic circulation, only 35 +/- 9% (n = 7) of midjejunally infused glucose did so, implying that 65 +/- 9% was retained in the splanchnic bed. Either glucose was less efficiently absorbed at the midintestinal site or hepatic glucose sequestration was increased 10-fold, or both. The proximal intestine plays a key role in the delivery of glucose to the systemic circulation, and the distal intestine potentially delivers more glucose to the liver.

Zinc Metabolism in Non-insulin Dependent Diabetes Mellitus

Low plasma zinc concentrations and increased zinc excretion have been reported in patients with non-insulin dependent diabetes mellitus (NIDDM). This study was designed to compare zinc metabolism in ten NIDDM patients with ten healthy controls, using stable isotopes of zinc. Zinc absorption from a standard breakfast labelled extrinsically with 3 mg 67Zn was measured using a fecal monitoring technique and thermal ionization quadrupole mass spectrometry. One hour after the test meal each volunteer was given 0.5 mg 70Zn (as citrate) intravenously and blood samples taken at intervals for up to 6 days. Isotopic enrichment was measured and the data entered into a simple kinetic model to estimate the size and rate of turnover of exchangeable pools of zinc. Urinary zinc excretion was higher in the male diabetics than controls (p < 0.05), but not in females. Since the efficiency of absorption and endogenous losses of zinc were very variable between individuals, and there was only a limited amount of data, it was not possible to draw conclusion about the effect of NIDDM on zinc absorption and excretion. However, the fact that there were no differences in exchangeable zinc pool sizes suggests that zinc metabolism is not altered in NIDDM.

Batch growth of Salmonella typhimurium LT2: stoichiometry and factors leading to cessation of growth

Salmonella typhimurium LT2 was grown in batch culture (trypticase soy broth, with 0.3%(w/v) yeast extract, 1% (w/v) glucose and 0.5% (w/v/) NaCl, 20 degrees C) at a range of initial pH (4.4, 4.8, 5.0 and 7.0). The consumption of oxygen and glucose was found to be independent of initial pH, and stoichiometric with growth. Mean yield coefficients of 6.9 x 10(-15) and 15.5 x 10(-15) mol oxygen/cell were estimated. Calculation of the instantaneous state of carbon during the cultivation showed stoichiometric conversion of glucose into biomass, carbon dioxide and organic acids. The concentration of the undissociated form of the primary acidic product (acetic acid) was shown to be the factor limiting growth.

Production of 13C-labelled β-carotenefrom Dunaliella salina

A 4.5 litre photobioreactor was developed and used to produce 13 C-labelled b-carotene by feeding 13 CO2 to growing ells of the marine alga Dunaliella salina. Three runs were carried out using different feeding regimes producing b-carotene containing 40, 56 and 76 Atom% 13 C. Approximately 145 mg b-carotene was recovered from each run. Incorporation of 13 C into b-carotene ranged from 1.4 to 2.8%. The labelled b-carotene produced would allow multiple tracer experiments for the study of human carotenoid metabolism.

On-line estimation of biomass using dynamic oxygen balancing

A method for the on-line estimation of biomass is proposed. The method is based on a liquid-phase dynamic mass balance for oxygen during a sequence of “no aeration” transients. The method has the advantage that no samples need be withdrawn and the biomass need not be homogeneously distributed in the vessel. The technique thus has applications in the estimation of immobilised biomass and plant organ culture, e.g. “hairy roots”.