A dual-labeled fluorescent probe for visualization of dextranase activity in a simulated food digestion system.

Molecular bioimaging of enzyme activity is rapidly emerging as a powerful strategy for accurate disease diagnostics. This work aims to prove that bioimaging of enzyme activity in food digestion with a fluorescent probe is feasible. In this study, a dual-labeled fluorescent probe with dextran-tetramethylrhodamine (TMR)-biotin conjugate (DTB) as the enzyme-cleavable unit, and biotin-(5-fluorescein) conjugate (FB) as the reference unit, was developed. It was immobilized in the agarose gel (the model food matrix) for the fluorescence quantification of dextranase activity. The probe manifested significantly ratiometric fluorescent signals (Igreen/Ired) in response to the enzyme-active reaction. Linear relationships of Igreen/Ired were obtained against the dextranase concentration ratio (C/C0). Igreen/Ired increased more rapidly with a greater dextranase diffusion rate, also supported by the more significant diffusion coefficient of fluorescently labeled dextranase in 0.5 wt% agarose gel (1.87 × 10-6 cm2 s-1). Our work provides more mechanistic evidence for enzyme activity imaging in food digestion.

Controlling the rheological properties of W1/O/W2 multiple emulsions using osmotic swelling: Impact of WPI-pectin gelation in the internal and external aqueous phases.

The purpose of this study was to create W1/O/W2 Multiple Emulsions by controlled osmotic swelling, and gelation of whey protein isolate (WPI) and high methoxy pectin (HMP) microspheres in internal and external acidic aqueous phases. Three different kinds of W1/O/W2 multiple emulsions (ME) were prepared, with 8 wt.% Polyglycerol polyricinoleate (PGPR) in their oil phases, with WPI and HMP in internal and external aqueous phases (250 mM NaCl, pH 3.5): (i) ME1: The inner aqueous phase (W1) contained 40% buffer solution, while W2 consisted of 10% WPI and 2% HMP; (ii) ME2: W1 contained 10% WPI, with 2% HMP (250 mM NaCl) in W2; (iii) ME3: 10% WPI and 2% HMP in W1, while W2 contained 1% Tween 80. The original multiple emulsions were diluted by different factors (1:0 to 1:5 with citrate buffer solution), and subject to thermal treatment from 25 to 90 °C to compare their microstructural and rheological properties. It was observed that the ME1 emulsion had higher viscosity and shear modulus than for other emulsions. After dilution however, the shear viscosity of ME3 was higher than ME1 and ME 2 at intermediate shear rates, which showed that the emulsions were osmotically well controlled in internal aqueous phases. Optical and confocal microscopy also supported our rheological measurements with evidence of WPI-HMP gelation, and osmotic swelling, in original and in swollen multiple emulsions. The results of this work may provide useful information about the encapsulation of bioactive compounds, in internal and external aqueous phases, for the development of healthier reduced-fat products in food industry.

Quantification of the oxygen uptake rate in a dissolved oxygen controlled oscillating jet-driven microbioreactor.

BACKGROUND: Microbioreactors have emerged as a new tool for early bioprocess development. The technology has advanced rapidly in the last decade and obtaining real-time quantitative data of process variables is nowadays state of the art. In addition, control over process variables has also been achieved. The aim of this study was to build a microbioreactor capable of controlling dissolved oxygen (DO) concentrations and to determine oxygen uptake rate in real time. RESULTS: An oscillating jet driven, membrane-aerated microbioreactor was developed without comprising any moving parts. Mixing times of ∼7 s, and kLa values of ∼170 h-1 were achieved. DO control was achieved by varying the duty cycle of a solenoid microvalve, which changed the gas mixture in the reactor incubator chamber. The microbioreactor supported Saccharomyces cerevisiae growth over 30 h and cell densities of 6.7 gdcw L-1. Oxygen uptake rates of ∼34 mmol L-1 h-1 were achieved. CONCLUSION: The results highlight the potential of DO-controlled microbioreactors to obtain real-time information on oxygen uptake rate, and by extension on cellular metabolism for a variety of cell types over a broad range of processing conditions. © 2015 The Authors. Journal of Chemical Technology & Biotechnology published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Oxygen transfer characteristics of miniaturized bioreactor systems.

Since their introduction in 2001 miniaturized bioreactor systems have made great advances in function and performance. In this article the dissolved oxygen (DO) transfer performance of submilliliter microbioreactors, and 1-10 mL minibioreactors was examined. Microbioreactors have reached k(L) a values of 460 h(-1) , and are offering instrumentation and some functionality comparable to production systems, but at high throughput screening volumes. Minibioreactors, aside from one 1,440 h(-1) k(L) a system, have not offered as high rates of DO transfer, but have demonstrated superior integration with automated fluid handling systems. Microbioreactors have been typically limited to studies with E. coli, while minibioreactors have offered greater versatility in this regard. Further, mathematical relationships confirming the applicability of k(L) a measurements across all scales have been derived, and alternatives to fluorescence lifetime DO sensors have been evaluated. Finally, the influence on reactor performance of oxygen uptake rate (OUR), and the possibility of its real-time measurement have been explored.