G. Vaccari

Assessment of in-line near-infrared spectroscopy for continuous monitoring of fermentation processes

The application of NIR in‐line to monitor and control fermentation processes was investigated. Determination of biomass, glucose, and lactic and acetic acids during fermentations of Staphylococcus xylosus ES13 was performed by an interactance fiber optic probe immersed into the culture broth and connected to a NIR instrument. Partial least squares regression (PLSR) calibration models of second derivative NIR spectra in the 700–1800 nm region gave satisfactory predictive models for all parameters of interest: biomass, glucose, and lactic and acetic acids. Batch, repeated batch, and continuous fermentations were monitored and automatically controlled by interfacing the NIR to the bioreactor control unit. The high frequency of data collection permitted an accurate study of the kinetics, supplying lots of data that describe the cultural broth composition and strengthen statistical analysis. Comparison of spectra collected throughout fermentation runs of S. xylosus ES13, Lactobacillus fermentum ES15, and Streptococcus thermophylus ES17 demonstrated the successful extension of a unique calibration model, developed for S. xylosus ES13, to other strains that were differently shaped but growing in the same medium and fermentation conditions. NIR in‐line was so versatile as to measure several biochemical parameters of different bacteria by means of slightly adapted models, avoiding a separate calibration for each strain.

Near-Infrared Spectroscopy: A Tool for Monitoring Submerged Fermentation Processes Using an Immersion Optical-Fiber Probe

Near-infrared (NIR) spectroscopy has been developed as a noninvasive tool for the direct, real-time monitoring of glucose, lactic acid, acetic acid, and biomass in liquid cultures of microrganisms of the genera Lactobacillus and Staphylococcus. This was achieved employing a steam-sterilizable optical-fiber probe immersed in the culture (In-line Interactance System®). Second-derivative spectra obtained were subjected to partial least-squares (PLS) regression and the results were used to build predictive models for each analyte of interest. Multivariate regression was carried out on two different sets of spectra, namely whole broth minus the spectral subtraction of water, and raw spectra. A comparison of the two models showed that the first cannot be properly applied to real-time monitoring, so this work suggests calibration based on non-difference spectra, demonstrating it to be sufficiently reliable to allow the selective determination of the analytes with satisfactory levels of prediction (standard error of prediction (SEP) < 10%). Direct interfacing of the NIR system to the bioreactor control system allowed the implementation of completely automated monitoring of different cultivation strategies (continuous, repeated batch). The validity of the in-line analyses carried out was found to depend crucially on maintaining constant hydrodynamic conditions of the stirred cultures because both gas flow and stirring speed variations were found to markedly influence the spectral signal.

Substrate preference of Bifidobacterium adolescentis MB 239: compared growth on single and mixed carbohydrates.

The utilization of mono-, di-, and oligosaccharides by Bifidobacterium adolescentis MB 239 was investigated. Raffinose, fructooligosaccharides (FOS), lactose, and the monomeric moieties glucose and fructose were used. To establish a hierarchy of sugars preference, the kinetics of growth and sugar consumption were determined on individual and mixed carbohydrates. On single carbon sources, higher specific growth rates and cell yields were attained on di- and oligosaccharides compared to monosaccharides. Analysis of the carbohydrates in steady-state chemostat cultures, growing at the same dilution rate on FOS, lactose, or raffinose, showed that monomeric units and hydrolysis products were present. In chemostat cultures on individual carbohydrates, B. adolescentis MB 239 simultaneously displayed α-galactosidase, β-galactosidase, and β-fructofuranosidase activities on all the sugars, including monosaccharides. Glycosyl hydrolytic activities were found in cytosol, cell surface, and growth medium. Batch experiments on mixtures of carbohydrates showed that they were co-metabolized by B. adolescentis MB 239, even if different disappearance kinetics were registered. When mono-, di-, and oligosaccharides were simultaneously present in the medium, no precedence for monosaccharides utilization was observed, and di- and oligosaccharides were consumed before their constitutive moieties.

Fermentative production of l-lactic acid by Lactobacillus casei DSM 20011 and product recovery using ion exchange resins

In order to produce l(+)-lactic acid to be employed in poly-l-lactic acid polymer production, for biomedical applications, the strain Lactobacillus casei subsp. casei DSM 20011 was studied in a conventional batch mode using different initial concentrations of glucose. The results obtained showed that the initial glucose concentration exerts an influence on the fermentation pattern, modifying the different fermentation parameters. Nevertheless, the product yield remained at a constant value of 0.86 g·g−1. The proposed novel system of product recovery, based on the use of ion-exchange resins, gave high yields of pure lactic acid.

Enhanced production of L-(+)-lactic acid in chemostat by Lactobacillus casei DSM 20011 using ion-exchange resins and cross-flow filtration in a fully automated pilot plant controlled via NIR.

Due to the lack of suitable in-process sensors, on-line monitoring of fermentation processes is restricted almost exclusively to the measurement of physical parameters only indirectly related to key process variables, i.e., substrate, product, and biomass concentration. This obstacle can be overcome by near infrared (NIR) spectroscopy, which allows not only real-time process monitoring, but also automated process control, provided that NIR-generated information is fed to a suitable computerized bioreactor control system. Once the relevant calibrations have been obtained, substrate, biomass and product concentration can be evaluated on-line and used by the bioreactor control system to manage the fermentation. In this work, an NIR-based control system allowed the full automation of a small-scale pilot plant for lactic acid production and provided an excellent tool for process optimization. The growth-inhibiting effect of lactic acid present in the culture broth is enhanced when the growth-limiting substrate, glucose, is also present at relatively high concentrations. Both combined factors can result in a severe reduction of the performance of the lactate production process. A dedicated software enabling on-line NIR data acquisition and reduction, and automated process management through feed addition, culture removal and/or product recovery by microfiltration was developed in order to allow the implementation of continuous fermentation processes with recycling of culture medium and cell recycling. Both operation modes were tested at different dilution rates and the respective cultivation parameters observed were compared with those obtained in a conventional continuous fermentation. Steady states were obtained in both modes with high performance on lactate production. The highest lactate volumetric productivity, 138 g L(-1) h(-1), was obtained in continuous fermentation with cell recycling.

Minimum energy consumption in sugar production by cooling crystallisation of concentrated raw juice

Abstract The sugar manufacturing process based on cooling crystallisation of concentrated raw juice is considered. Micro-filtration and softening of raw juice makes it possible to obtain white sugar by three-or four-stage cooling crystallisation. Prior to crystallisation, raw juice should be concentrated by multistage evaporation in a pressure range below the atmospheric pressure. The preferred evaporator arrangement is backward feed. As the temperature of vapours and condensates leaving the evaporator station is low, the opportunities for heat recovery are limited. In order to save energy, vapour recompression can be applied. For a given evaporator structure, the energy consumption depends on the parameters of the evaporation process. The minimum energy consumption of the novel process should thus be determined by simultaneously carrying out energy targeting and optimisation of evaporation. It is assumed that the consumption of utilities in the sugar factory is balanced with the supply of heat and power from the factory’s own power plant. Energy systems employing various CHP technologies and various evaporator stations optionally combined with vapour compressors are studied. A range of feasible solutions is defined in terms of minimum energy consumption and combined heat transfer area of the evaporator station and the heat exchanger network.

Growth of sucrose crystals and tapering effect studied by holographic interferometry

Effects of dopant raffinose on the growth of sucrose crystals are studied by means of holographic interferometry. By the distribution of hologram fringes the selective doping effect is measured on {100}, {110}, and {110} forms. An interpretation is put forward for the tapering effect in wedge shaped crystals.

Equilibrium and growth forms of sucrose crystals in the {h0l} zone: 1. Theoretical treatment of {101}-d form

Abstract Observed {h0l} growth forms of the sucrose crystal are compared with calculated ones, on the basis of attachment energy plots. Agreement is found for all forms only if a modification of character of the {101}-d face is taken into consideration. Calculation of equilibrium and growth forms have been carried out using two potential for dispersion and hydrogen bond energy respectively. The same method is applied for studying the spiral growth shape on the (101) face and its field of validity is discussed: at low driving force values, the attachment energy at the steps is the relevant energy, determining the shape of spirals on the (101) face of sucrose.

Detection of oligosaccharides in sugar products using planar chromatography

Abstract Oligosaccharides, and in particular raffinose and kestoses, are of great importance not only in the field of cane and beet processing but also in respect of the analyses of a number of agricultural raw materials and trade products. The authors judge it to be useful to have at ones disposal a simple and reliable analytical technique to be used for the detection of oligosaccharides in different materials. Modern planar chromatography can be utilized, bearing in mind that HPTLC (high performance thin layer chromatography) plates, Automated Multiple Development, completely automated elution systems, sample positioning and spots detection apparatuses, are available. Details on the analytical methodology adopted for molasses and other sugar products are presented and discussed also giving statistical data about its accuracy and precision. Some examples of practical application of the proposed methodology are described.

Growth morphology of sucrose crystals The role of glucose and fructose as habit-modifiers

Abstract The influences of glucose (G) and fructose (F) on the habit of sucrose crystals were analysed through stereochemical compatibility between their molecules and surface sites of the crystal. First neighbour interactions (H-bondings) and the terrace–ledge–kink (TLK) surface model are adopted. Notwithstanding the simple qualitative model, this method of analysis appears to be satisfactory to account for the preferential adsorption upon specific crystal forms. A more complex model for adsorption in the kink sites, together with the behaviour as disruption tailor-made additives are proposed for both G and F, to highlight why high concentrations are needed to induce macroscopic habit variations. X-ray powder diffractometry (XRPD) shows the systematic effect of the impurities on the bulk structure of the sucrose crystal.