Design of Model Apple Cells Suspensions: Rheological Properties and Impact of the Continuous Phase

The objective of this work is twofold: to develop a relevant model system to study plant cells suspensions’ rheology and to evaluate the impact of the continuous phase composition and viscosity on the rheological behaviour of apple cells suspensions. Model suspensions of individual or clustered apple cells were developed. Rheological behaviours of both type of suspensions were observed separately, suspending from 0.145 g/100mL to 3.48 g/100mL of particles in five model media and in the original apple serum. Our results show that model suspensions successfully reproduce the rheological behaviour of apple purees, following three concentration domains. In particular, cell clusters greatly reproduce the behaviour of bimodal apple purees, suggesting that clusters dominate the rheological behaviour of the whole puree. One of our main result is that continuous phase does not affect elastic properties of suspensions in the concentrated domain since they are essentially governed by particle interactions: G’ values are similar whatever the continuous phase. If the continuous phase has the main impact on diluted suspensions’ viscosity, its effect becomes smaller as particle concentration increases. A lubricating effect was observed in the concentrated domain for continuous phases containing polymers. Presence of polymers may help in structuring the network in the intermediate domain.     

Rheology of corn stover slurries at high solids concentrations--effects of saccharification and particle size

The rheological characteristics of untreated and dilute acid pretreated corn stover (CS) slurries at high solids concentrations were studied under continuous shear using plate-plate type measurements. Slurry rheological behavior was examined as a function of insoluble solids concentration (10-40%), extent of pretreatment (0-75% removal of xylan) and particle size (-20 and -80 mesh). Results show that CS slurries exhibit shear-thinning behavior describable using a Casson model. Further, results demonstrate that the apparent viscosity and yield stress increase with increasing solids concentration (which corresponds to a decrease in free water). Dilute acid pretreatment leads to lower viscosity and yield stresses at equivalent solids concentrations, as does smaller particle size. Taken together, these findings are consistent with the hypothesis that the availability of free water in the slurry plays a significant role in determining its rheological behavior. In particular, as the free water content of the slurry decreases, e.g., with increasing solids concentration, the greater interaction among particles likely increases the apparent viscosity and yield stress properties of the slurry. The results also suggest that the availability of free water, and thereby slurry rheological properties, depend on the chemical composition of the corn stover as well as its physical characteristics such as particle size and porosity. Hydrophilic polymers within the cell wall, such as xylan or pectin, or larger pores within bigger particles, facilitate sequestration of water in the solid phase resulting in decreased availability of free water. Thus, dilute acid pretreated slurries, which contain smaller size particles having significantly lower xylan content than slurries of untreated milled stover, exhibit much lower viscosities and yield stresses than untreated slurries containing large particles at similar solid concentrations.     

Rheological characterization of dilute acid pretreated softwood

Large-scale bioethanol production from lignocellulosic biomass will require high solids loading in the enzymatic hydrolysis step. However, slurries of pretreated lignocelluloses are complex fluids due to the fibrous nature, especially at high concentrations of water insoluble solids (WIS). A prerequisite for dealing with transport issues and for developing efficient full-scale processes is a fundamental understanding of the flow properties of pretreated lignocellulose. A comprehensive rheological characterization of dilute acid pretreated spruce has been carried out in this study, accounting for the effects of WIS concentration, particle size distribution (PSD), and the degree of enzymatic hydrolysis. The rheology of pretreated spruce slurries was found to be strongly dependent on the WIS concentration. The storage modulus (G'(LVR)) and yield stress showed typical power-law dependencies on volume fraction and WIS content. Milling of the pretreated material resulted in significantly higher yield stress and viscosity, likely due to narrower PSD, which suggests that the strength of the network of the coarsest fibers determines the rheology of these materials to a large extent. During enzymatic hydrolysis, yield stress and viscosity decreased dramatically, partly due to decreasing WIS content, but possibly also due to changes in fiber properties such as the chemical composition.     

Effects of cell wall components on the functionality of wheat gluten

Normal white wheat flours and especially whole meal flour contain solids from the inner endosperm cell walls, from germ, aleurone layer and the outer layers of cereal grains. These solids can prevent either gluten formation or gas cell structure. The addition of small amounts of pericarp layers (1–2%) to wheat flour had a marked detrimental effect on loaf volume. Microstructural studies indicated that in particular the epicarp hairs appeared to disturb the gas cell structure. The detrimental effects of insoluble cell walls can be prevented by using endoxylanases. It has been shown that some oxidative enzymes, naturally present in flour or added to the dough, will oxidise water-extractable arabinoxylans via ferulic acid bridges, and the resulting arabinoxylan gel will hinder gluten formation. The negative effects of water-unextractable arabinoxylans on gluten yield and rheological properties can be compensated by the addition of ferulic acid. Free ferulic acid can probably prevent arabinoxylan cross-linking via ferulic acid.     

Power Laws in the Elasticity and Yielding of Plant Particle Suspensions

The yielding and flow behaviour of plant suspensions are perhaps the most important rheological properties in process and product design for applications in paper, biofuel and food industries. Studies are reported here on the yield properties and flow behaviour of suspensions of plant particles with different shapes (clusters of cells, individual cells and cell fragments). Carrot and tomato were selected as model plant systems to prepare suspensions at particle dry mass concentrations ranging from 0.010 to 0.065. The flow behaviour was characterised by an apparent yield stress and shear thinning. The Herschel-Bulkley yield stress obtained from up and return flow curves was compared to the yield stress calculated from oscillatory measurements. The dependence of the yield stress values on particle dry mass concentration is approximately a power-law, with a fitted exponent of 3 ± 0.5 for all the suspensions, independently of the plant origin and particle shape. This same power-law behaviour was found for the elastic modulus G′, and in this case the exponent was 3 for carrot and 4 for the tomato suspensions. The yield strain, calculated from oscillatory measurements, decreased slightly with dry mass fraction, but did not follow a power-law. We discuss possible explanations for power law behaviour, and provide a model for G′ based on folded elastic sheets, which predicts an exponent of 3, similar to the values obtained for these suspensions.     

Shear Elastic Deformation and Particle Packing in Plant Cell Dispersions

The relationship between small amplitude oscillatory rheological properties and microstructure of plant cell suspensions was studied. Carrot, broccoli and tomato were selected as model plant systems to generate particles with different microstructures: clusters of cells with smooth or rough edges and single cells. By analysing the compressive stress undergone by the plant cells under centrifugation, and comparing this to oscillatory rheometry, agreement was found between the compressive stress required to compress the dispersions to higher insoluble solids dry mass fractions, and the elastic shear modulus of the plant dispersions. This indicated that centrifugation is acting as a crude rheological measurement on the samples, rather than measuring any well-defined “particle phase volume”. We estimated the theoretical critical dry mass fraction above which smooth, roughly spherical, elastically interacting particles would acquire a non-zero G′, and compared this with the experimental values. Our results give evidence that for the three vegetable suspensions considered here, the elastic rheology observed is not coming simply from the packing of smooth particles, but is dominated in the dilute limit by attractive forces or interaction of asperities, and in the concentrated limit by deformation and buckling acting together. Improved understanding of the particles and their packing would help in the structuring of food products without adding other texturising or stabilising agents.     

Fish meal replacement by plant protein substitution and guar gum addition in trout feed. Part II: Effects on faeces stability and rheology

In the explosively growing aquaculture industry, the challenge of sustainability includes a requirement to produce more fish than are consumed by stock. A promising avenue of research is the substitution of the fish meal in feeds with plant protein. However, there are inherent risks in the development of such feeds, and serious consideration should be given not only to nutritional content, but also to the mechanical quality of resulting faecal wastes. The present study uses a plate on plate rheometer running in different flow modes (creep, oscillation) to assess the rheological properties of wastes resulting from different diets.All faeces were shown to be thixotropic in nature, independent of diet composition. Details of dietary composition influence the consistency and the characteristic stresses at which faecal structure changes from a viscoelastic solid into a viscoelastic liquid. Furthermore, in linking active food components with mechanical properties of chyme faeces, rheological studies may be used to understand and counteract some problematic properties. Substituting 100% of fish meal with plant proteins leads to faeces that rapidly disintegrate into very fine solids which threaten the viability of aquacultural operations. Adding just 0.3% of the rapidly hydrating non-starch polysaccharide, guar gum (GG), significantly increased the viscosity and elastic modulus of the faeces. These mechanical improvements increase the size of the resulting particles and the effectiveness with which they can be removed, thereby leading to optimized water quality. GG addition was sufficiently effective to counter the stability and particle size effects of a 50% substitution of fish meal, but could not mitigate those of a 100% substitution, wherein dissolution effects of an unknown emulsifier are suspected.     

Experimental Measurement of Settling Velocity of Spherical Particles in Unconfined and Confined Surfactant-based Shear Thinning Viscoelastic Fluids

An experimental study is performed to measure the terminal settling velocities of spherical particles in surfactant based shear thinning viscoelastic (VES) fluids. The measurements are made for particles settling in unbounded fluids and fluids between parallel walls. VES fluids over a wide range of rheological properties are prepared and rheologically characterized. The rheological characterization involves steady shear-viscosity and dynamic oscillatory-shear measurements to quantify the viscous and elastic properties respectively. The settling velocities under unbounded conditions are measured in beakers having diameters at least 25x the diameter of particles. For measuring settling velocities between parallel walls, two experimental cells with different wall spacing are constructed. Spherical particles of varying sizes are gently dropped in the fluids and allowed to settle. The process is recorded with a high resolution video camera and the trajectory of the particle is recorded using image analysis software. Terminal settling velocities are calculated from the data.The impact of elasticity on settling velocity in unbounded fluids is quantified by comparing the experimental settling velocity to the settling velocity calculated by the inelastic drag predictions of Renaud et al.¹ Results show that elasticity of fluids can increase or decrease the settling velocity. The magnitude of reduction/increase is a function of the rheological properties of the fluids and properties of particles. Confining walls are observed to cause a retardation effect on settling and the retardation is measured in terms of wall factors.     

A simulation of the process of sedimentation of suspended solids in the Yoshii River estuary

A vertical plane 2D model has been applied to the Yoshii River estuary, Japan, as a means of simulating the process of sedimentation. The current velocity and isohaline distributions were well reproduced in the estuary and the model simulated the concentration distribution of suspended solids for 8 particle size classes by using a transport equation based only on physical processes. The changes of the particle size distribution in surface water were well reproduced by the model. The vertical velocity component plays an important role in the behaviour of suspended solids.     

Investigation of electrorheological properties of biodegradable modified cellulose/corn oil suspensions

Considerable scientific and industrial interest is currently being focused on a class of materials known as electrorheological (ER) fluids, which display remarkable rheological behaviour, being able to convert rapidly and repeatedly from a liquid to solid when an electric field (E) is applied or removed. In this study, biodegradable cellulose was modified and converted to their carboxyl salts. Modified cellulose is characterised by Fourier transform infrared (FTIR) spectroscopy, nuclear magnetic resonance (NMR) spectroscopy, energy dispersive spectroscopy (EDS), thermogravimetric analysis (TGA) and conductivity measurements. Suspensions of cellulose (C) and modified cellulose (MC) were prepared in insulated corn oil (CO). The effects of electric field strength, shear rate, shear stress, temperature, etc. of these suspensions onto ER activity were determined. Rheological measurements were carried out via a rotational rheometer with a high-voltage generator to investigate the effects of electric field strength and particle concentration on ER performance.The results show that the ER properties are enhanced by increasing the particle concentration and electric field strength. Also the cellulose-based ER fluids exhibit viscoelastic behaviour under an applied electric field due to the chain formation induced by electric polarization between particles.     

The Role of Polygalacturonase (PG) in the Ripening of Apple Fruits

Polygalacturonase (PG) activity and changes in respiratory intensity of apple fruits were investigated. The respiratory rate was decreased to a preclimacteric minimum from 30 Aug. to 20 Sept., Then increased to a climacteric peak (20–30 Sept.) and again drop down gradually with approaching the senescence stage. The PG activity was undetectable in a developing fruit until the onset climacteric phase. It rose rapidly after harvest, and reaching its highest level on 27 Oct. Just a month after the climacteric peak. The PG activity fell gradually. The amount of the fractions of pectic acid in fruits changed with the modifications of PG activity. With the ripening of fruits, the content of alcohol-soluble small molecules of pectic acids was increased from 12 to 13 5 mg/100 g of tissue, while the amount of alcohol- insoluble large molecules of pectic acids reduced from 530 to 280/100 g of tissue. PG activity would indicate the destruction of cell walls and the separation of cells. The onset of softening of fruits occurred 20 days after the rise of PG activity. It is supposed that the process of softening is directly controlled by PG activity.     

Kinetic model for microbial uptake of insoluble solid-state substrate

A kinetic model for anaerobic digestion of insoluble solid-state substrates was developed. Rate equations for cell growth and substrate consumption were derived based on the assumption that the microorganisms assimilate the substrate mainly at the point of contact where they grow. The model emphasizes effects of substrate particle size, organic loading, and cell concentration on the rates of cell growth and substrate utilization. Batch digestion of a stearic acid emulsion with a mean particle size of 2.0 mum and a biological sludge was conducted at 30 and 37 degrees C to verify the proposed model. Agreement between the experimental and calculated results indicated the validity of the model for describing the microbial degradation of insoluble solid-state substrates. Further examinationof the model revealed that with low cell substrate affinity or at low cell concentration, it coincided with a Michaelis-Menten type kinetics in which the effect of particle size was taken into consideration.     

Acid invertase is predominantly localized to cell walls of both the practically symplasmically isolated sieve element/companion cell complex and parenchyma cells in developing apple fruits

The acid invertase (β‐fructosidase, EC 3·2·1·26) was localized at subcellular level via immunogold electron microscopy in the phloem‐unloading zone of developing apple fruit. The enzyme (immunogold particles) was found to reside predominantly in the cell walls of the sieve element/companion cell (SE/CC) complex, phloem parenchyma cells and other parenchyma cells. There was almost no gold particle found in cytoplasm and vacuole. This distribution pattern remained unchanged throughout the growing season, but the enzyme numbers varied. The density of immunogold particles increased during fruit development. The immunoblotting of soluble and insoluble acid invertases provided a supporting proof for the assays of immunolocalization. The biochemical analysis showed a predominantly cell‐wall‐distributed activity of acid invertase that corresponds essentially with its amount distribution. The ultrastructural observations showed that there were numerous plasmodesmata between the parenchyma cells, but almost no plasmodesmium between the SE/CC complex and its surrounding parenchyma cells, practically resulting in the symplasmic isolation of the SE/CC complex. It is therefore suggested that the unloading pathway of sucrose from the SE/CC complex may be predominantly apoplasmic in the developing apple fruit, and that the unloaded sucrose may be hydrolysed by the functional acid invertase localized in the cell wall before it is loaded in sink cells.     

Investigation of settleability of biologically produced solids and biofilm morphology in moving bed bioreactors (MBBRs)

The objective of this work is to investigate the effects of surface area loading rates (SALRs) and hydraulic retention times (HRTs) in moving bed bioreactor (MBBR) systems on the morphology and thickness of the attached biofilm along with subsequent effects on particle size distribution and the settling characteristics of the biologically produced solids. The morphology of biofilm attached to the MBBR carriers changed from a porous biofilm to a biofilm with a more filamentous structure throughout the study at various operating conditions without observable correlation with SALR and HRT. Although, biofilm morphology did not demonstrate an effect on the biologically produced solids observed in this study, the thinnest biofilms resulted in the highest concentration of solids in the effluent. Furthermore, the particle size distribution analysis demonstrated that both higher SALRs and longer HRTs resulted in a shift towards larger-sized particles. Increases in SALR and HRT, independent of each other, also showed increases in effluent solid concentration and lower settleability of the solids.     

Effects of microprojectile bombardment on embryogenic suspension cell cultures of maize (Zea mays L.) used for genetic transformation

We have investigated the interaction between tungsten and gold microprojectiles with suspension-culture cells of maize used for genetic transformation. Particle size measurements were evaluated before and after DNA precipitation to determine mean particle size and the effect of DNA precipitation on particle aggregation. Following particle bombardment, metal foils were examined by scanning electron microscopy to visualize dispersion of individual particles and aggregates. Particle penetration into suspension-culture cell clusters was examined in paraffin-embedded bombarded cells serially sectioned and viewed with light microscopy and by energy dispersive X-ray microanalysis. Acridine-orange-stained bombarded cells were examined to observe cellular response to particle penetration. Transient expression of reporter genes C1 and B and GUS, (-glucuronidase) were used to assess effects of particle bombardment on embryogenic cell types. Autoradiographic analysis of the transformable suspension cell culture SC82 (see Gordon-Kamm et al. 1990, Plant Cell 2, 603–618) was conducted to evaluate the S-phase and mitotic indices in embryogenic and nonembryogenic cells throughout a subculture passage and in response to DNA/particle delivery. The results of these investigations are discussed relative to cytodifferentiation of suspension cell clusters and recovery of transformed clonal sectors.Abbreviations GUS -glucuronidase - FAA formaldehyde-acetic acid-alcohol - SEM scanning electron microscopy     

Fractionation of cell-wall preparations from grass leaves by centrifuging in non-aqueous density gradients.

1. Dried preparations of cell walls from perennial-ryegrass (Lolium perenne) and Italian-ryegrass (L. multiflorum) leaves were suspended in mixtures of carbon tetrachloride with light-petroleum (b.p. 45--50 degrees C) or alcohols and layered on density gradients formed from the same solvents. 2. On centrifugation, the cell walls become distributed throughout a suitably chosen gradient. Fractions corresponding to various regions of the gradient were separated, examined under the microscope and analysed. 3. Cell-wall preparations made from leaf material ground in liquid N2, or in a triple roll mill, showed considerable heterogeneity in particle size, and their behaviour in the density gradient was variable, although there was a general indication that walls derived from vascular bundles were less dense than those from sclerenchyma. 4 Treatment in a vibratory ball mill decreased the size of the particles and produced a more uniform material, but made it impossible to distinguish the origins of the particles. This material behaved more reproducibly in the density gradient. 5. Some fractionations were also made by successive centrifugation in media of increasing relative density. 6. Analyses of the fractions obtained by each method indicated that the less dense had a greater proportion of xylose in the polysaccharide components, and higher contents of acetyl groups and lignin, confirming the close relationship between these components in plant cell walls. 7. The results show that there are differences in polysaccharide composition between the cell-wall types in the grass leaf, the vascular tissue being richer in hemicellulose relative to cellulose than the sclerenchyma.     

Oxidation of Phenolic Compounds by Peroxidase in the Presence of Soluble Polymers

The kinetics of Coprinus cinereus peroxidase-catalyzed 1-naphthol, 2-naphthol, and 4-hydroxybiphenyl oxidation was investigated. The initial rates of the naphthols' and 4-hydroxybiphenyl oxidations were linearly dependent on enzyme concentration. The rates depended on substrate concentration and saturated at concentrations above 100 microM of hydrogen peroxide, 25-50 microM of naphthols, and 10 microM of 4-hydroxybiphenyl. At the peroxide concentration 100 microM calculated K(m) and the maximal rate (V(max)) were 74.7 microM and 0.53 microM/sec or 175 microM and 2.0 microM/sec for 1- or 2-naphthol, respectively, and 29.68 microM and 0.42 microM/sec for 4-hydroxybiphenyl. Kinetic measurements of exhaustive naphthol and 4-hydroxybiphenyl oxidation showed that peroxidase is inactivated during the oxidation of the substrates. Different factors and additives, water soluble polymers and albumins (PEG, PEI, PL, BSA, HSA), influenced the initial naphthols and 4-hydroxybiphenyl oxidation rates, peroxidase inactivation rates, and the degree of the substrate conversion. Addition of albumin increased turnover number of naphthols oxidation 1.5-4 times. Light scattering increase was observed when peroxidase-catalyzed oxidation reaction was investigated and suggested that insoluble particles were formed during the process. The addition of polymers, change of concentration and ionic strength of the solution as well as the number of other factors influenced the observed light scattering. The number of particles formed during peroxidase-catalyzed naphthols' and 4-hydroxybiphenyl oxidation and their distribution according to size in the interval 2.5-300 microm were detected by particle counting in solutions.     

Short-term Studies of the Uptake of Nitrogen by Young Apple Trees after Soil Application of Ammonium Nitrate

Young trees of apple rootstock MM.104 were supplied with ammoniumnitrate (0.4 g in 100 ml water) via the soil at selected timesduring the annual growth cycle. Appreciable amounts of nitrogenwere taken up within 40 h in summer and autumn, leading to increasesin the soluble nitrogen fraction. Corresponding changes werenot detected in the insoluble nitrogen fraction. The increasesin soluble nitrogen were not dependent on carbohydrate status,plant size, root mass or on whether or not shoot extension hadceased. Amounts of increase were profoundly influenced by priornitrogen status as modified by a small pre-treatment boost ofammonium nitrate given in July.     

Interactions between apple cell walls and native apple polyphenols: quantification and some consequences

Cell walls were prepared from apple parenchyma by a phenol:buffer procedure. Polyphenols were extracted from freeze-dried apple parenchyma by methanol and water:acetone, and purified by preparative HPLC. Interactions were quantified by bringing into contact suspended cell walls and polyphenol solutions. Hydroxycinnamic acids and (-)-epicatechin did not bind to cell walls. Binding of procyanidins was fast and reached up to 0.6 g per g cell walls. The amounts of procyanidins bound increased with the initial concentration and with DPn. Procyanidins could be partially desorbed by buffer, more being desorbed in the presence of dissolved cell wall polysaccharides. They were totally desorbed using 8 M urea or acetone:water. Higher polymers were bound selectively from procyanidin mixtures, and very high average DPn were obtained in extensively washed complexes. Binding of procyanidins inhibited enzymic degradation of the cell walls.     

Cell wall composition of the carrageenophyte Kappaphycus alvarezii (Gigartinales, Rhodophyta) partitioned by wet sieving

The cortical and medullary cells of Kappaphycus alvarezii fractions were screened by wet sieving after aqueous extraction of carrageenans. The cell populations obtained showed a clear partition between these two cell types. The main monosaccharide in hydro-insoluble cell walls was cellulosic glucose (70% dry weight), the crystallinity of which was shown torange from 20% in the cortical cells to 45% in the large medullary cells (over 250 μm diameter). Minor monosaccharides in the insoluble fraction were galactose, 3,6-anhydrogalactose (indicating presence of residual carrageenans), mannose and xylose. However, the major part of the remaining galactose probably originated from another galactoglycan strongly linked to insoluble polymers in the large medullarycell walls. The mannose concentration was maximum in the cortical cells and decreased with increasing size of the medullary cells. Thus, besides cellulose, two other types of polysaccharides were detected in insolublecell walls, mannoglycans and galactoglycans in cortical and medullary cellwalls, respectively. This revised version was published online in August 2006 with corrections to the Cover Date.