Recuperação de carotenóides microbianos por diferentes técnicas de ruptura celular

O objetivo deste trabalho foi utilizar diferentes técnicas químicas (dimetilsulfóxido, ácido clorídrico, acético e lático), técnicas mecânicas (banho ultrassônico, abrasão com pérolas de vidro, maceração com terra diatomácea, ruptor ultrassônico e imersão em nitrogênio líquido) e técnica enzimática (preparado enzimático comercial Glucanex®) para a recuperação de carotenoides a partir da ruptura da parede celular das leveduras Sporidiobolus pararoseus e Rhodotorula mucilaginosa isoladas de amostras ambientais. Para isso a obtenção de biomassa foi realizada através de cultivos submersos no meio YM, a 25 °C, 180 rpm por 168 h. Para a ruptura celular, a operação de congelamento da biomassa (-18°C por 48 h) foi estudada. Os métodos de secagem convencional por ar forçado (35°C/48h) e liofilização (-80°C/48h, em ultrafreezer, seguido de liofilizador até alcançar 2% de umidade da amostra) também foram avaliados. Nas técnicas químicas aplicadas, o dimetilsulfóxido apresentou os melhores resultados para as duas leveduras, porém o seu uso é limitado devido a sua toxicidade. Para S. pararoseus, os maiores valores encontrados foram para o ácido clorídrico, seguido do acético e do lático, sendo detectada diferença entre eles quando aplicado o congelamento. Com R. mucilaginosa, os maiores valores foram encontrados para os ácidos acético e lático, seguido do ácido clorídrico, no qual o congelamento da biomassa também não influenciou a recuperação dos carotenoides. Dentre as técnicas mecânicas estudadas, para a levedura S. pararoseus, o banho ultrassônico e a abrasão com pérolas de vidro apresentaram os resultados mais promissores comparados ao DMSO (84,79±2,34 e 76,87±2,06 μg/g respectivamente), onde o processo de congelamento da biomassa não influenciou positivamente no percentual de extratibilidade e na concentração específica dos carotenoides quando utilizada estas técnicas. Com Rhodotorula mucilaginosa, o banho ultrassônico propiciou a recuperação da maior concentração específica de carotenoides (193,5±25,8 μg/g), sendo que o processo de congelamento também não influenciou positivamente no percentual de extratibilidade e na concentração específica dos carotenoides. Através do Delineamento Central Composto Rotacional (DCCR) 23 foi possível avaliar que a levedura S. pararoseus não demonstrou nenhum efeito sob as variáveis pH, temperatura e concentração de enzima. Assim, a melhor condição de trabalho escolhida foi pH 7,4, 30 ºC e concentração de enzima de 1,0 g/gcs, onde apresentou a concentração específica de 42,6 μg/g e volumétrica de 308 μg/L de carotenoides. Para R. mucilaginosa, a condição ótima foi definida como 1,0 g/gcs, pH 5,0 e temperatura de 30 ºC, onde foi encontrado 115,1±8,1 μg/g e 470,1±38,8 μg/L para a concentração específica e volumétrica de carotenoides, respectivamente. A utilização de técnicas combinadas empregando banho ultrassônico e lise enzimática não proporcionou melhorias nos resultados para ambas as leveduras. A liofilização provocou um ganho de 20% e 13,7% na concentração específica dos carotenoides das leveduras S. pararoseus e R. mucilaginosa, respectivamente, onde o congelamento da biomassa não influenciou significativamente (p<0,05) a recuperação de carotenoides provenientes das duas leveduras, podendo ser eliminada do processo. Assim, para S. pararoseus o banho ultrassônico e as pérolas de vidro apresentaram os melhores resultados na recuperação de carotenoides, e para R. mucilaginosa o melhor resultado foi alcançado com o banho ultrassônico.

The measurement of broadband ultrasonic attenuation in cancellous bone-a review of the science and technology

The measurement of broadband ultrasonic attenuation (BUA) in cancellous bone at the calcaneus was first described in 1984. The assessment of osteoporosis by BUA has recently been recognized by Universities UK, within its EurekaUK book, as being one of the “100 discoveries and developments in UK Universities that have changed the world” over the past 50 years, covering the whole academic spectrum from the arts and humanities to science and technology. Indeed, BUA technique has been clinically validated and is utilized worldwide, with at least seven commercial systems providing calcaneal BUA measurement. However, a fundamental understanding of the dependence of BUA upon the material and structural properties of cancellous bone is still lacking. This review aims to provide a science- and technology-orientated perspective on the application of BUA to the medical disease of osteoporosis.

Ultrasonic measurement of fine head movements in a standard ophthalmic headrest

We aimed to investigate the naturally occurring horizontal plane movements of a head stabilized in a standard ophthalmic headrest and to analyze their magnitude, velocity, spectral characteristics, and correlation to the cardio pulmonary system. Two custom-made air-coupled highly accurate (±2 μm)ultrasound transducers were used to measure the displacements of the head in different horizontal directions with a sampling frequency of 100 Hz. Synchronously to the head movements, an electrocardiogram (ECG) signal was recorded. Three healthy subjects participated in the study. Frequency analysis of the recorded head movements and their velocities was carried out, and functions of coherence between the two displacements and the ECG signal were calculated. Frequency of respiration and the heartbeat were clearly visible in all recorded head movements. The amplitude of head displacements was typically in the range of ±100 μm. The first harmonic of the heartbeat (in the range of 2–3 Hz), rather than its principal frequency, was found to be the dominant frequency of both head movements and their velocities. Coherence analysis showed high interdependence between the considered signals for frequencies of up to 20 Hz. These findings may contribute to the design of better ophthalmic headrests and should help other studies in the decision making of whether to use a heavy headrest or a bite bar.

Evaluation cortical bone elasticity in response to pulse power excitation using ultrasonic technique

This paper presents the ultrasonic velocity measurement method which investigates the possible effects of high voltage high frequency pulsed power on cortical bone material elasticity. Before applying a pulsed power signal on a live bone, it is essential to determine the safe parameters of pulsed power applied on bone non-destructively. Therefore, the possible changes in cortical bone material elasticity due to a specified pulsed power excitation have been investigated. A controllable positive buck-boost converter with adjustable output voltage and frequency has been used to generate high voltage pulses (500V magnitude at 10 KHz frequency). To determine bone elasticity, an ultrasonic velocity measurement has been conducted on two groups of control (unexposed to pulse power but in the same environmental condition) and cortical bone samples exposed to pulsed power. Young’s modulus of cortical bone samples have been determined and compared before and after applying the pulsed power signal. After applying the high voltage pulses, no significant variation in elastic property of cortical bone specimens was found compared to the control. The result shows that pulsed power with nominated parameters can be applied on cortical bone tissue without any considerable negative effect on elasticity of bone material.

Power converters design and analysis for high power piezoelectric ultrasonic transducers

High power piezoelectric ultrasonic transducers have been widely exploited in a variety of applications. The critical behaviour of a piezoelectric device is encapsulated in its resonant frequencies because of its maximum transmission performance at these frequencies. Therefore power electronic converters should be tuned at those resonant frequencies to transfer electrical power to mechanical power efficiently. However, structural and environmental changes cause variations in the device resonant frequencies which can degrade the system performance. Hence, estimating the device resonant frequencies within the incorporated setup can significantly improve the system performance. This paper proposes an efficient resonant frequency estimation approach to maintain the performance of high power ultrasonic applications using the employed power converter. Experimental validations indicate the effectiveness of the proposed method.

A high cell count cascade full bridge converter for wide bandwidth ultrasonic transducer excitation

A nine level modular multilevel cascade converter (MMCC) based on four full bridge cells is shown driving a piezoelectric ultrasonic transducer at 71 and 39 kHz, in simulation and experimentally. The modular cells are small stackable PCBs, each with two fully integrated surface mount 22 V, 40 A MOSFET half-bridge converters, and include all control signal and power isolation. In this work, the bridges operate at 12 V and 384 kHz, to deliver a 96 Vpp 9 level waveform with an effective switching frequency of 3 MHz. A 9 pH air cored inductor forms a low pass filter in conjunction with the 3000 pF capacitance of the transducer load. Eight equally phase-displaced naturally sampled pulse width modulation (PWM) drive signals, along with the modulating sinusoid, are generated using phase accumulation techniques in a dedicated FPGA. Experimental time domain and FFT plots of the multilevel and transducer output waveforms are presented and discussed.

The presence of ultrasonic harmonics in the calls of the rifleman (Acanthisitta chloris)

Ultrasonic vocalisations (frequencies > 20 kHz) have been extensively studied in the context of echolocation by bats and other mammals (Sales & Pye 1974; Wilson & Hare 2004). Ultrasonic calls have also been recorded from birds, including the blue-throated hummingbird ( Lampornis clemenciae ) (Pytte et al. 2004), where it was first thought that individuals made use of high pitch calls to avoid masking by background noise in a visually obscured environment. Similarly, city-dwelling great tits ( Parus major ) use song with a higher minimum frequency (although not ultrasonic) compared to woodland birds to communicate with conspecifics to avoid the predominantly low-frequency background noise in the city (Slabbekorn & Peet 2003). The theory that birds use ultrasound to avoid noise masking was discarded when it was discovered that there was no corresponding auditory brainstem response (i.e. sensory perception) to the ultrasonic calls in the hummingbirds producing those calls.

Process considerations related to the microencapsulation of plasmid DNA via ultrasonic atomization

An effective means of facilitating DNA vaccine delivery to antigen presenting cells is through biodegradable microspheres. Microspheres offer distinct advantages over other delivery technologies by providing release of DNA vaccine in its bioactive form in a controlled fashion. In this study, biodegradable poly(D,L-lactide-coglycolide) (PLGA) microspheres containing polyethylenimine (PEI) condensed plasmid DNA (pDNA) were prepared using a 40 kHz ultrasonic atomization system. Process synthesis parameters, which are important to the scale-up of microspheres that are suitable for nasal delivery (i.e., less than 20 μm), were studied. These parameters include polymer concentration; feed flowrate; volumetric ratio of polymer and pDNA-PEI (plasmid DNA-polyethylenimine) complexes; and nitrogen to phosphorous (N/P) ratio. PDNA encapsulation efficiencies were predominantly in the range 82-96%, and the mean sizes of the particle were between 6 and 15 μm. The ultrasonic synthesis method was shown to have excellent reproducibility. PEI affected morphology of the microspheres, as it induced the formation of porous particles that accelerate the release rate of pDNA. The PLGA microspheres displayed an in vitro release of pDNA of 95-99% within 30 days and demonstrated zero order release kinetics without an initial spike of pDNA. Agarose electrophoresis confirmed conservation of the supercoiled form of pDNA throughout the synthesis and in vitro release stages. It was concluded that ultrasonic atomization is an efficient technique to overcome the key obstacles in scaling-up the manufacture of encapsulated vaccine for clinical trials and ultimately, commercial applications.

Creation of protein loaded biodegradable microparticles via ultrasonic atomization suitable for nasal delivery

Improved biopharmaceutical delivery may be achieved via the use of biodegradable microspheres as delivery vehicles. Biodegradable microspheres offer the advantages of maintaining sustained protein release over time whilst simultaneously protecting the biopharmaceutical from degradation. Particle samples produced by ultrasonic atomization were studied in order to determine a feed stock capable of producing protein loaded poly-ε-caprolactone (PCL) particles suitable for nasal delivery (i.e., less than 20 μm). A 40 kHz atomization system was used with a 6 mm full wave atomization probe. The effect of solids percent, feed flow rate, volumetric ratio of the polymer stock to the protein stock, and protein concentration in the protein stock on particle size characteristics were determined. It was shown that feed stocks containing 100 parts of 0.5 or 1.0% w/v PCL in acetone with one part 100 mg ml -1 BSA and 15 mg ml -1 PVA produced particles with a mass moment diameter (D[4,3]) of 13.17 μm and 9.10 μm, respectively in addition to displaying high protein encapsulation efficiencies of 93 and 95%, respectively. The biodegradable PCL particles were shown to be able to deliver encapsulated protein in vitro under physiological conditions.

Straightforward biodegradable nanoparticle generation through megahertz-order ultrasonic atomization

Simple and reliable formation of biodegradable nanoparticles formed from poly-ε-caprolactone was achieved using 1.645 MHz piston atomization of a source fluid of 0.5% w/v of the polymer dissolved in acetone; the particles were allowed to descend under gravity in air 8 cm into a 1 mM solution of sodium dodecyl sulfate. After centrifugation to remove surface agglomerations, a symmetric monodisperse distribution of particles φ 186 nm (SD=5.7, n=6) was obtained with a yield of 65.2%. © 2006 American Institute of Physics.