In vivo measure of average bacterial cell size from a polarized light scattering function.

A particular combination of elements of the Mueller matrix for scattering of polarized light given by (S34 + S14)/(S11 + S13) identical to (S34/S11)++ is measured vs angle at a wavelength of 633 nm for randomly oriented suspensions of several species of bacteria in different stages of growth. (This combination of elements is dominated in the present measurements by the behavior of the normalized S34 matrix element, as is indicated by the notation defined on the right side of the equation.) The resulting graph in each case shows an oscillating function of angle. This function is compressed toward smaller angles when the bacteria are in the exponential phase of growth in comparison with results for a suspension of the same bacteria in the stationary (starving-smaller cells) phase of growth. Microscopic measurements were made to determine, for each case, the average dimensions of the bacterial population. Graphs were then plotted of the peak positions from the Mueller matrix function plots vs either cell length or cell diameter. The function was shown to be strongly correlated with cell diameter under the conditions of this experiment and poorly correlated with cell length. The measurements were shown to have a sensitivity to changes in average diameter of about 20 nm.     

Relation between particle size and shape of insecticidal suspensions and their contact toxicity; D.D.T. and rotenone suspensions against Oryzaephilus surinamensis L., with some time-mortality studies

Methods already worked out for the preparation and testing of aqueous D.D.T. suspensions against Tribolium castaneum , by a dipping method, have been applied to Oryzaephilus surinamensis , giving similar results, i.e. toxicity increases with increase in particle size.
In the same way, the precipitation of rotenone by exchange of solvents leads to the formation of simple aqueous suspensions. The theory of precipitation is described and methods are given of preparing five types of suspension: colloidal rotenone, a suspension of small elongated plates, a suspension containing small hexagonal plates in aggregates, and two suspensions containing hexagonal plate-shaped crystals of different sizes.
These were tested, by dipping, against Oryzaephilus surinamensis L. Within the range of crystal sizes up to about 150μ, toxicity is inversely related to the size of particle in suspension. The variation in median lethal concentration obtained in this way is of the order of 600 times. Crystal shape seems to be unimportant. Similar results were obtained with fine suspensions, using a spraying method.
The variation of mortality with time was also studied, using D.D.T. against Tribolium castaneum and rotenone against Oryzaephilus surinamensis. In the former case, both colloidal and crystalline D.D.T. show progressively increasing kills with the passage of time. Crystalline rotenone behaves similarly, but colloidal rotenone gives an initial paralytic effect, followed by recovery of the insects.     

Measuring diameters of rod-shaped bacteria in vivo with polarized light scattering.

The angular function for elements of the Mueller matrix for polarized light scattering from suspensions of microorganisms is known to be reproducible for different growths of a given bacterial strain in the log (or exponential) phase of growth. The reason for this, the stability of the size and shape distribution for cells, is briefly discussed. Experiments were performed using suspensions of two different strains of Escherichia coli cells in log phase and measuring the angular dependence of the Mueller matrix ratio S34/S11. Calculations were then performed using the coupled dipole approximation to model electromagnetic scattering from particles where the shape of an individual cell was approximated by a cylinder capped with hemispheres of the same radius as the cylinder. Using previously measured values for the length distribution and index of refraction of the cells, the calculated scattering curve was found to fit the measured curve very well. The values obtained for the cell diameters were quite close to diameters previously measured by optical microscopy. Thus this method provides a rapid and convenient method for monitoring bacterial diameters in vivo even when there is an appreciable distribution of bacterial lengths in the population.     

Feasibility of Focused Beam Reflectance Measurement (FBRM) for Analysis of Pharmaceutical Suspensions in Preclinical Development

This study examined the use of focused beam reflectance measurement (FBRM) for qualitative and quantitative analysis of pharmaceutical suspensions with particular application to toxicology supply preparations for use in preclinical studies. Aqueous suspensions of ibuprofen were used as prototype formulations. Initial experiments were conducted to examine the effects of operational conditions including FBRM probe angle, probe location, and mixing (method and rate of mixing) on the FBRM analysis. Once experimental conditions were optimized, the homogeneity and sedimentation-redispersion of particles in the suspensions were assessed. Ibuprofen suspension under continuous agitation was monitored using FBRM for 60 h to study particle size change over time. Another study was performed to determine if particle count rates obtained by FBRM could be correlated to suspension concentration. The location and the angle of the FBRM probe relative to the beaker contents, and the rate and the method of mixing the suspension were found to be sensitive parameters during FBRM analysis. FBRM was able to monitor the process of particle sedimentation in the suspension. The attrition of ibuprofen particles was detectable by FBRM during prolonged stirring with an increase in the number of smaller particles and decrease in the number of larger particles. A strong correlation was observed between particle count rate by FBRM and ibuprofen concentration in the suspension. Also, change in content uniformity in the suspension at different locations of the beaker was represented by FBRM particle count. Overall, FBRM has potential to be a useful tool for qualitative and quantitative analysis of pharmaceutical suspensions.     

Shear Flow Behavior of Aqueous Suspensions of Potato Parenchyma Powder

The shear flow behavior of potato powder suspensions prepared from two different particle sizes and with a range of solids volume fraction (Φ) was studied. A concentrated sucrose solution was used as the continuous phase to maintain particle buoyancy. The shear flow properties were measured at 20, 50 and 80 °C. The suspensions obeyed a power-law equation in the dilute regime while the Herschel-Bulkley equation was the best fit for almost all semi-dilute and more concentrated suspensions. With increasing Φ, particle size and temperature, a gradual development of shear-thinning behavior was evident which coincided with an increase in the consistency index and the development of a yield stress in the suspensions. Potato powder suspensions therefore behave very differently to potato starch suspensions, with flow properties dominated by the effect of intra- and inter-cellular components in the potato powder particles that are transferred to the continuous phase and that alter suspension properties.     

Inertial migration based concentration factors for suspensions of Chlorella micro-algae in branched tubes

When a dilute suspension flows in the laminar regime through a tube, under certain conditions the suspended particles migrate radially to an equilibrium radial position. Branched tubes can use this radial concentration distribution to concentrate dilute suspensions. Suspensions of microalgae, Chlorella vulgaris, were pumped through tubes of various diameters for tube Reynolds number ranging from 47–1839 and photographed. Upstream particle concentration profdes were obtained by image analysis of the photographs. The dividing stream surfaces in branched tubes were obtained from the three-dimensional numerical solutions of the Navier-Stokes equations for steady, laminar, and homogeneous flow through tubes having one and two orthogonal branches. Concentration factors for Chlorella suspensions in branched tubes, predicted by a general method, fall in the range of 1.0–1.3     

CD of gels and suspensions: apparent CD in the soret region of sickle hemoglobin gels

CD spectra in the soret region of sickle-cell deoxyhemoglobin (deoxy-HbS) fiber gels are radically different from the CD of deoxy-HbS in solution. An explanation is found using the Stokes–Mueller representation of the interaction of a polarized beam with the instrument optical train and sample to derive expressions for the apparent CD of gels and suspensions of optically active molecules that consist of randomly oriented domains or particles that are linearly dichroic and linearly birefringent. These theoretical considerations show that the apparent CD spectra from such systems have contributions from the LD and birefringence of each domain even if no net linear birefringence and dichroism is apparent in the sample. Thus, the interpretation of the CD from gels and suspensions is problematic, unless it can be demonstrated that each domain or particle has extremely small absorbance or that the LD and birefringence of each is a very small fraction of the total absorbance. As a result, we conclude that the spectra of HbS gels are not due to the CD of the heme per se; rather, they also reflect the randomly oriented domain structure of the gels and the LD and linear birefringence associated with each domain.     

Low-temperature micronization of a peptide drug in fluid propellant: Case study cetrorelix

Aim of this study was to elaborate an efficient method for the micronization of the decapeptide cetrorelix (a GnRH-antagonist), in order to obtain a microsuspension as basis for other pharmaceutical preparations, such as e.g. inhalation aerosols. A modified pearl-mill coupled with a cryostat was used for the micronization of cetrorelix in fluid propellant and operated under different conditions. The obtained cetrorelix suspensions were analyzed for particle size distribution, purity of cetrorelix, and for metal contamination through abrasion from parts of the mill. The method allowed an effective micronization of cetrorelix. The mean particle size of the initial cetrorelix lyophilizate bulk ware was reduced from 52.5 μ (Volume Mean Diameter, VMD) down to 14.9, 6.1 and 3.1 μm, respectively, respectively. The HPLC analysis of all cetrorelix suspensions after micronization did not show signs of decomposition as compared to the initial product. The elementary analysis of the suspensions performed by inductively coupled plasma mass spectrometry revealed a negligible amount of contaminants in the suspension (Zr=max. 0.6 ppm; Fe, Cr, Ni, Ba, below limit of quantification, i.e.<0.14 ppm). The only appreciable contaminant. Aluminum (Al=1.1 ppm), was derived from the mechanical capping of aluminum canisters prior to analysis. The Zr determination in the suspension of 0.6 ppm, is still considered to be negligible as compared to the legally tolerated limit of air contamination. By low-temperature micronization in fluid propellant, fine drug suspensions of cetrorelix for pMDIs can be directly manufactured in one-step procedure without destruction of the peptide structure and without appreciable product contamination. Published: July 12, 2001.     

Optimizing the classification of biological tissues using machine learning models based on polarized data

Polarimetric data is nowadays used to build recognition models for the characterization of organic tissues or the early detection of some diseases. Different Mueller matrix-derived polarimetric observables, which allow a physical interpretation of a specific characteristic of samples, are proposed in literature to feed the required recognition algorithms. However, they are obtained through mathematical transformations of the Mueller matrix and this process may loss relevant sample information in search of physical interpretation. In this work, we present a thorough comparative between 12 classification models based on different polarimetric datasets to find the ideal polarimetric framework to construct tissues classification models. The study is conducted on the experimental Mueller matrices images measured on different tissues: muscle, tendon, myotendinous junction and bone; from a collection of 165 ex-vivo chicken thighs. Three polarimetric datasets are analyzed: (A) a selection of most representative metrics presented in literature; (B) Mueller matrix elements; and (C) the combination of (A) and (B) sets. Results highlight the importance of using raw Mueller matrix elements for the design of classification models.     

Nanometer-Scale Sizing Accuracy of Particle Suspensions on an Unmodified Cell Phone Using Elastic Light Scattering

We report on the construction of a Fourier plane imaging system attached to a cell phone. By illuminating particle suspensions with a collimated beam from an inexpensive diode laser, angularly resolved scattering patterns are imaged by the phone''s camera. Analyzing these patterns with Mie theory results in predictions of size distributions of the particles in suspension. Despite using consumer grade electronics, we extracted size distributions of sphere suspensions with better than 20 nm accuracy in determining the mean size. We also show results from milk, yeast, and blood cells. Performing these measurements on a portable device presents opportunities for field-testing of food quality, process monitoring, and medical diagnosis.     

Dispersibility in water of dried nanocrystalline cellulose

Dispersibility is important for nanocrystalline cellulose (NCC) because recovering the unique suspension and particle properties is essential after the product has been dried for storage or transport. It is our goal to produce dried NCC that redisperses in water to yield colloidal suspensions without the use of additives or a large energy input. In contrast with the as-prepared acidic form of NCC (H-NCC), suspensions of neutral sodium-form NCC (Na-NCC) dried by evaporation, lyophilization, or spray-drying are readily dispersible in water. Suspension properties and NCC particle size determined by light scattering were used as indicators of dispersion quality. The neutral counterion content, drying technique, freezing action, drying and redispersion concentrations, and moisture content in the dried NCC were all found to influence dispersibility. When a minimum of 94% of the H(+) counterion is exchanged for Na(+), the neutral salt form is fully dispersible in water even when fully dried. Mild sonication is generally sufficient to recover measured particle sizes identical to those in the never-dried Na-NCC sample. A threshold moisture content of 4 wt % was found, above which dried H-NCC is fully dispersible in water.     

Comportement rhéologique de biomatériaux pour l’ingénierie ostéoarticulaire et dentaire : matrices extracellulaires synthétiques et suspensions phosphocalciques

Injectable biomaterials are a particular field of biomaterials used for noninvasive surgical techniques (e.g. percutaneous surgery). The fundamental characteristic of this type of biomaterials is their rheological properties during implantation. In this context, the subject of this research work was to evaluate the rheological properties of two injectable biomaterials used in osteoarticular and dental tissue engineering: (i) a synthetic extracellular matrix and (ii) an injectable calcium phosphate suspension. The rheological properties of silated hydroxypropylmethylcellulose hydrogel were studied. It is shown that although silanization reduces the hydrodynamic volume in dilute solution, it does not affect significantly the rheological behavior of the concentrated solutions. In dilute solution, intrinsic viscosity of different HPMC-Si solutions before steam sterilization indicated that macromolecular chains occupied larger hydrodynamic volume compared to the sterilized HPMC-Si solutions. For the sterilized HPMC-Si concentrated solutions, the limiting viscosities decreased when the pH increased. This change, remarked in dilute and concentrated domain has been attributed to the formation of both intra- and intermolecular associations during the phase separation process of HPMC-Si during steam sterilization. The formation of HPMC-Si hydrogels from injectable aqueous solution was studied after neutralization. The study of the gelation process revealed the dependence of the final concentration of HPMC-Si hydrogel, pH and temperature on cross-linking kinetics and viscoelastic properties. An injectable calcium phosphate ceramic suspension was studied. This “ready-to-use” injectable bone substitute is consisting of an aqueous HPMC solution as matrix and calcium phosphate particles as fillers. The rheological characterization revealed the macromolecular behavior of the HPMC. The investigations of settling kinetics showed the dependence of the particle size and the HPMC concentration on the settling velocity and sediment compactness before and after sterilization. The rheological properties and injectability of this suspension were also studied. The suspensions showed a strongly increased viscosity as compared to the HPMC solution. The rheological proprieties of suspensions depend on the composition. A simple device has been used to characterize extrusion of the paste using a disposable syringe fitted with a needle. The injectability modeling was realized. A theoretical approach based on the capillary flow of non newtonian fluids was used to predict the necessary pressure for injection, on the basis of rheological properties and extrusion conditions. The theoretical estimation of the extrusion pressure showed a wall slip in the suspensions, so that the injection pressure is less than anticipated. The influence of wall slip leads, however, to a constant proportionality factor between theory and injection experiments.     

High-frequency transformation of undeveloped plastids in tobacco suspension cells

Although leaf chloroplast transformation technology was developed more than a decade ago, no reports exist of stable transformation of undeveloped plastids or other specialized plastid types, such as proplastids, etioplasts, or amyloplasts. In this work we report development of a dark-grown tobacco suspension cell model system to investigate the transformation potential of undeveloped plastids. Electron microscope analysis confirmed that the suspension cells carry plastids that are significantly smaller (approximately 50-fold less in volume) and have a very different subcellular localization and developmental state than leaf cell chloroplasts. Using antibiotic selection in the light, we demonstrated that both plastid and nuclear transformation of these cell suspensions is efficient and reproducible, with plastid transformation frequency at least equal to that of leaf chloroplast transformation. Homoplasmic plastid transformants are readily obtained in cell colonies, or in regenerated plants, providing a more consistent and versatile model than the leaf transformation system. Because of the uniformity of the cell suspension model, we could further show that growth rate, selection scheme, particle size, and DNA amount influence the frequency of transformation. Our results indicate that the rate-limiting steps for nuclear and plastid transformation are different, and each must be optimized separately. The suspension cell system will be useful as a model for understanding transformation in those plant species that utilize dark-grown embryogenic cultures and for characterizing the steps that lead to homoplasmic plastid transformation.     

Distinguishing structural features between Crohn's disease and gastrointestinal luminal tuberculosis using Mueller matrix derived parameters

Recently, the incidence of inflammatory bowel diseases, especially the Crohn's disease (CD) and gastrointestinal luminal tuberculosis (ITB), has grown rapidly worldwide. Currently there is no general gold standard to distinguish between CD and ITB tissues, which both have tuberculosis and surrounding fibrous structures. Mueller matrix imaging technique is suitable for describing the location, density and distribution behavior of such fibrous structures. In this study, we apply the Mueller matrix microscopic imaging to the CD and ITB tissue samples. The 2D Mueller matrix images of the CD and ITB tissue slices are measured using the Mueller matrix microscope developed in our previous study, then the Mueller matrix polar decomposition and Mueller matrix transformation parameters are calculated. To evaluate the distribution features of the fibrous structures surrounding the tuberculosis areas more quantitatively and precisely, we analyze the retardance related Mueller matrix derived parameters, which show clear different distribution behaviors between the CD and ITB tissues, using the Tamura image processing method. It is demonstrated that the Mueller matrix derived parameters can reveal the structural features of tuberculosis areas and be used as quantitative indicators to distinguish between CD and ITB tissues, which may be useful for the clinical diagnosis.      

Development and Validation of a Discriminative Dissolution Test for Nimesulide Suspensions

The dissolution test for oral dosage forms has recently widened to a variety of special dosage forms such as suspensions. For class II drugs, such as nimesulide (NMS), this study is very important because formulation problems may compromise drug bioavailability. In the present work, tests with four brands of commercially available NMS (RA, TS, TB, and TC) have been performed in order to study their dissolution at different conditions. The suspensions have been characterized relatively to particle size, pH, and density besides NMS assay and the amount of drug in solution in the suspension vehicles. The dissolution study was conducted using the following media: simulated intestinal fluid, pH 6.8, containing polysorbate 80 (P80) or sodium lauryl sulfate (SLS); phosphate buffer, pH 7.4, with P80 and aqueous solution of SLS. Concerning the quantitative analysis, the UV–VIS spectrophotometry could have been used in substitution to high-performance liquid chromatography since the methodology had been adequately validated. The influence of the drug particle size distribution was significant on the dissolution profiles of NMS formulations, confirming to be a factor that should be strictly controlled in the development of oral suspensions.     

Periodic cell aggregation in suspensions of Dictyostelium discoideum

Abstract. Periodic activities of Dictyostelium discoideum can be observed in cell suspension as two types of oscillations in the light-scattering properties, spike-shaped and sinusoidal. Responses of suspended cells to applied chemoattractants are also reflected by transient changes in light scattering. Alterations in the light-scattering properties are due to structural changes such as changes in cell shape and/or changes in the size of cell aggregates. Therefore, changes in the aggregation state during autonomous oscillations and during attractant-induced responses were investigated. In order to be able to withdraw multiple samples and larger sample volumes from optically monitored cell suspensions, a photometer comprising glass fiber optics immersable in a cell suspension was constructed. Samples were fixed with formaldehyde and photographed. The aggregation state of the samples was quantified by counting the number of particles (cells and cell aggregates) per volume. Folic acid elicited in suspensions of undifferentiated cells a transient decrease in the number of particles per volume as did cAMP in suspensions of preaggregation cells. Periodic changes in the number of particles per volume occurred synchronously with spike-shaped and sinusoidal oscillations. The relative amplitude of the oscillations in particle number was larger during sinusoids than during spikes. Photographs showed periodic changes in the aggregate size during sinusoidal oscillations. In each cycle, the cell-aggregation phase was followed by a phase of partial disaggregation. The recurring loosening of cell-cell contacts may be relevant for sorting out the different cell types. The potential role of contact site as synchronizer and as constituent of an oscillator is discussed.     

Experimental test of absorption flattening correction for circular dichroism of particle suspensions

There are well established theoretical models for correction for absorption flattening of circular dichroism (CD) measurements on particle suspensions. However, these have not been directly tested experimentally. We describe a test system with the chiral tris(ethylenediamine)Co(III) complex dissolved in water trapped inside sephadex particles, suspended in 1-butanol. Independent measurements of particle size distribution, volume fraction, and the absorbance of the suspension are used to calculate the required CD correction. The corrected CD signal is found to agree rather well with that for the same amount of Co-complex dispersed uniformly throughout the sample cell. This holds for different particle volume fractions and Co-complex concentrations inside the particles. The correction seems to work despite a substantial scattering contribution to the absorbance, which is not considered in the theoretical models.     

The effect of particle characteristics on the beam attenuation coefficient and output from an optical backscatter sensor

Experiments were conducted to measure optical backscatter and beam transmission of suspensions of 180, 150 and 90 μm sand, and 40 μm clay, in a recirculation tank designed to house an optical backscatterance sensor (O.B.S.) and a beam transmissometer. Particle size was determined using gravimetric techniques and Coulter counter. By contriving known sediment distributions from the fractionated sediment samples, it was found that both the O.B.S. and beam transmissometer responded approximately linearly to narrow band and broad band particle suspensions. The beam transmissometer showed greater sensitivity to the fine-grain fraction of a poly-disperse suspension than the O.B.S.