Enhancement for the synthesis of bio-energy molecules (carbohydrates and lipids) in Desmodesmus subspicatus: experiments and optimization techniques.

Microalgae are regarded as renewable resources of energy, foods and high-valued compounds using a biorefinery approach. In the present study, we explored isolated microalgae (Desmodesmus subspicatus) for the production of bio-energy molecules (carbohydrate and lipid). Optimizations of media (BG-11) components have been made using the Taguchi orthogonal array (TOA) technique to maximize biomass, carbohydrate and lipid production. Optimized results showed that biomass, carbohydrates and lipid productivity increased by 1.3 times at optimal combinations of media components than standard BG-11 media. Further, the influence of various carbon and nitrogen sources as nutritional supplement with optimum media composition under different light intensities was investigated for productivity of carbohydrate and lipid. Results demonstrated that 1.5 times higher productivity of carbohydrate and lipids were achieved in the presence optimum BG-11 under a broad range of light intensities (84-504 µmol m-2 s-1). Among different nitrogen sources, glycine was found to give higher productivity (1.5 times) followed by urea. Use of the cellulose as a carbon source in the media significantly increases biomass (2.4 times), carbohydrates (2.3 times) and lipids (2.3 times) productivity. Investigations revealed that cultivating Desmodesmus subspicatus under optimum culture conditions has the potential for large-scale bio-ethanol and bio-diesel production.

A new methodology for measuring solid/liquid interfacial energy.

HYPOTHESIS: The interfacial energy γsl between a solid and a liquid designates the affinity between these two phases, and in turn, the macroscopic wettability of the surface by the fluid. This property is needed for precise control of fluid-transport phenomena that affect the operation/quality of commercial devices/products. Although several indirect or theoretical approaches can quantify the solid/liquid interfacial energy, no direct experimental procedure exists to measure this property for realistic (i.e. rough) surfaces. Makkonen hypothesized that the frictional resistance force per unit contact-line length is equal to the interfacial energy on smooth surfaces, which, however, are rarely found in practice. Consequently, the hypothesis that Makkonen's assumption may also hold for rough surfaces (which are far more common in practice) arises naturally. If so, a reliable and simple experimental methodology of obtaining γsl for rough surfaces can be put forth. This is accomplished by performing dynamic contact-angle experiments on rough surfaces that quantify the relationship between the frictional resistance force per unit contact-line length acting on an advancing liquid (Fp,a) and the surface roughness in wetting configurations. EXPERIMENT: We perform static and advancing contact-line experiments with aqueous and organic liquids on different hydrophilic surfaces (Al, Cu, Si) with varying Wenzel roughnesses in the range 1-2. These parameters are combined with the liquid's known surface tension to determine Fp,a. FINDINGS: Fp,a rises linearly with the surface roughness. Analysis based on existing theories of wetting and contact-angle hysteresis reveals that the slope of Fp,a vs.Wenzel roughness is equal to the solid/liquid interfacial energy, which is thus determined experimentally with the present measurements. Interfacial energies obtained with this experimental approach are within 12% of theoretically predicted values for several solid/liquid pairs, thereby validating this methodology.

Process intensification for the enhancement of growth and chlorophyll molecules of isolated Chlorella thermophila: A systematic experimental and optimization approach.

In our current work, we have optimized six physicochemical parameters (light intensity, light period, pH, inoculum size, culture period, and salt concentration) toward growth and chlorophyll synthesis using isolated fresh water microalgae Chlorella thermophila [contains ∼6% (w/w on dry biomass basis) chlorophyll]. Here, both experimental and computational [Taguchi orthogonal array (TOA), artificial neural network (ANN), and genetic algorithm (GA)] approaches were employed for the process intensification. Results revealed that the content of biomass and chlorophyll were enhanced by 118% and 95%, respectively, with productivity enhancement of 30% for biomass and 61% for chlorophyll from the optimization of physicochemical parameters. Further, optimum light intensity was found to be 128 µmol m-2 s-1 after conducting experiments in optimized chemical and physicochemical conditions, contributing to the enhancement of productivity of 46% for biomass and 106% for chlorophyll. Urea was found to be the most effective nitrogen source with an increase of 70% and 160% biomass and chlorophyll productivity, respectively. Moreover, sucrose as a carbon source contributed to an increase of 97% and 264% biomass and chlorophyll productivity.

Revisiting the supplementary relationship of dynamic contact angles measured by sessile-droplet and captive-bubble methods: Role of surface roughness.

HYPOTHESIS: Quantitative characterization of surface wettability through contact angle (CA) measurement using the sessile droplet (SD) or captive bubble (CB) methods is often limited by the intrinsic wetting properties of the substrate. Situations may arise when an extreme surface wettability may preclude using one of the two methods for predicting the behaviors of droplets or bubbles on the surface. This warrants a relationship between the dynamic CAs measured via the SD and CB methods. While the two dynamic CAs (e.g., the advancing CA of SD and receding CA of CB) add up to 180° on a smooth surface, the simple geometric supplementary principle may not apply for rough surfaces. EXPERIMENTS: We perform a systematic wettability characterization of solid substrates with varying degrees of roughness using the sessile-droplet and captive-bubble methods, and interpret the experimental observations using a theoretical model. FINDINGS: The dynamic contact angles measured by the sessile-droplet and captive-bubble methods deviate from the supplementary principle as the surface roughness is increased. We present a theoretical explanation for this disparity and predict the values of the contact angles using prevalent thermodynamic models of wetting and contact-angle hysteresis on rough substrates. The theoretical prediction is in good agreement with the experimental observations.

Disparities of Single-Particle Growth Rates in Buried Versus Exposed Ritonavir Crystals within Amorphous Solid Dispersions.

Seeded growth rates of ritonavir in copovidone at 75% relative humidity (RH) and 50 °C were evaluated by single-particle tracking second harmonic generation (SHG) microscopy and found to be ∼3-fold slower for crystallites at the surface compared to the bulk. The shelf lives of final dosage forms containing amorphous solid dispersions (ASDs) are often dictated by the rates of active pharmaceutical ingredient crystallization. Upon exposure to elevated RH, the higher anticipated water content near the surfaces of ASDs has the potential to substantially impact nucleation and growth kinetics relative to the bulk. However, quantitative assessment of these differences in growth rates is complicated by challenges associated with discrimination of the two contributions (supersaturation and molecular mobility) in ensemble-averaged measurements. In the present study, "sandwich" materials were prepared, in which sparse populations of ritonavir single-crystalline seeds were pressed between two similar ASD films to assess bulk crystallization rates. These sandwich materials were compared and contrasted with analogously prepared "open-faced" samples, without the capping film, to assess the surface crystallization rates. Single-particle analysis by SHG microscopy time-series during in situ crystallization produced average growth rates of 3.8 µm/h for bulk columnar crystals with a particle-to-particle standard deviation of 0.9 µm/h. In addition, columnar crystal growth rates for surface particles were measured to be 1.3 µm/h and radiating crystal growth rates for surface particles were measured to be 1.0 µm/h, both with a particle-to-particle deviation of 0.4 µm/h. The observed appearance of radiating crystals upon surface seeding is attributed to reduced ritonavir solubility upon water adsorption at the interface, leading to higher defect densities in crystal growth. Despite substantial differences in crystal habit, correction of the surface growth rates by a factor of 4 from geometric effects resulted in relatively minor but statistically significant differences in the growth kinetics for the two local environments. These results are consistent, with viscosity being a relatively weak function of water absorption coupled with primarily diffusion-limited growth kinetics.

In Situ Crystal Growth Rate Distributions of Active Pharmaceutical Ingredients.

Single-particle tracking of crystal growth performed in situ enables substantial improvements in the signal-to-noise ratio (SNR) for recovered crystal nucleation and growth rates by nonlinear optical microscopy. Second harmonic generation (SHG) is exquisitely sensitive to noncentrosymmetric crystals, including those produced by many homochiral active pharmaceutical ingredients (APIs). Accelerated stability testing at elevated temperatures and relative humidity informs design of pharmaceutical formulations. In the present work, we demonstrate reduction in the Poisson noise associated with the finite number of particles present in a given field of view through continuous monitoring during stability testing. Single-particle tracking enables recovery of crystal growth rates of individual crystallites and enables unambiguous direct detection of nucleation events. Collectively, these capabilities provide significant improvements in the signal-to-noise for nucleation and crystal growth measurements, corresponding to approximately an order of magnitude reduction in anticipated measurement time for recovery of kinetics parameters.

Characterization of Phase Transformations for Amorphous Solid Dispersions of a Weakly Basic Drug upon Dissolution in Biorelevant Media.

PURPOSE: The overall goal of this study was to investigate the dissolution performance and crystallization kinetics of amorphous solid dispersions (ASDs) of a weakly basic compound, posaconazole, dispersed in a pH-sensitive polymeric matrix consisting of hydroxypropyl methylcellulose acetate succinate (HPMC-AS), using fasted-state simulated media. METHODS: ASDs with three different drug loadings, 10, 25 and 50 wt.%, and the commercially available tablets were exposed to acidic media (pH 1.6), followed by transfer to, and dissolution in, intestinal media (pH 6.5). Parallel single stage dissolution experiments in only simulated intestinal media were also performed to better understand the impact of the gastric stage. Different analytical methods, including nanoparticle tracking analysis, powder x-ray diffraction, second harmonic generation and two-photon excitation ultraviolet fluorescence microscopy, were used to characterize the phase behavior of these systems at different stages of dissolution. RESULTS: Results revealed that all ASDs exhibited some degree of drug release upon suspension in acidic media, and were also vulnerable to matrix crystallization. Upon transfer to intestinal media conditions, supersaturation was observed. This was short-lived for some dispersions due to the release of the crystals formed in the acid immersion stage which acted as seeds for crystal growth. Lower drug loading ASDs also exhibited transient formation of amorphous nanodroplets prior to crystallization. CONCLUSIONS: This work emphasizes the significance of assessing the impact of pH change on dissolution and provides a fundamental basis of understanding the phase behavior kinetics of ASDs of weakly basic drugs when formulated with pH sensitive polymers.

Kinetic Modeling of Accelerated Stability Testing Enabled by Second Harmonic Generation Microscopy.

The low limits of detection afforded by second harmonic generation (SHG) microscopy coupled with image analysis algorithms enabled quantitative modeling of the temperature-dependent crystallization of active pharmaceutical ingredients (APIs) within amorphous solid dispersions (ASDs). ASDs, in which an API is maintained in an amorphous state within a polymer matrix, are finding increasing use to address solubility limitations of small-molecule APIs. Extensive stability testing is typically performed for ASD characterization, the time frame for which is often dictated by the earliest detectable onset of crystal formation. Here a study of accelerated stability testing on ritonavir, a human immunodeficiency virus (HIV) protease inhibitor, has been conducted. Under the condition for accelerated stability testing at 50 °C/75%RH and 40 °C/75%RH, ritonavir crystallization kinetics from amorphous solid dispersions were monitored by SHG microscopy. SHG microscopy coupled by image analysis yielded limits of detection for ritonavir crystals as low as 10 ppm, which is about 2 orders of magnitude lower than other methods currently available for crystallinity detection in ASDs. The four decade dynamic range of SHG microscopy enabled quantitative modeling with an established (JMAK) kinetic model. From the SHG images, nucleation and crystal growth rates were independently determined.

Variation in Supersaturation and Phase Behavior of Ezetimibe Amorphous Solid Dispersions upon Dissolution in Different Biorelevant Media.

The delivery of poorly water-soluble drugs using amorphous solid dispersions (ASDs) has been widely acknowledged as a promising strategy for enhancing oral bioavailability. Upon dissolution, ASDs have accelerated dissolution rates and yield supersaturated solutions leading to higher apparent solubilities. Understanding the complex phase behavior of ASDs during dissolution is crucial for developing an effective formulation. Since the absorption of a lipophilic, high permeability drug is determined primarily by the intraluminal dissolution process and the final concentration achieved, there is a need for evaluation in biorelevant dissolution media that simulate both fasting and fed gastrointestinal states. In this study, using ezetimibe as a model drug, three different ASDs were prepared using poly(acrylic acid) (PAA), polyvinylpyrrolidone (PVP), and hydroxypropyl methylcellulose acetyl succinate (HPMC-AS). Dissolution of ASDs was carried out in sodium phosphate buffer, fed-state simulated intestinal fluid (FeSSIF), and Ensure Plus to evaluate the impact of different dissolution media on release profile, supersaturation, and phase behavior. The supersaturation level and crystallization kinetics varied among the dispersions and were found to be highly dependent on the medium employed. The presence of solubilizing additives in biorelevant media greatly affected the generation and stabilization of supersaturated solutions. Second harmonic generation microscopy was found to enable the detection of crystals in all media including the highly turbid Ensure Plus system. In conclusion, it is important to evaluate the impact of complex biorelevant media on the dissolution performance of ASDs to better design supersaturating formulations for oral delivery.

Isolation and Tandem Mass Spectrometric Identification of a Stable Monolayer Protected Silver-Palladium Alloy Cluster.

A selenolate-protected Ag-Pd alloy cluster was synthesized using a one-pot solution-phase route. The crude product upon chromatographic analyses under optimized conditions gave three distinct clusters with unique optical features. One of these exhibits a molecular peak centered at m/z 2839, in its negative ion mass spectrum assigned to Ag5Pd4(SePh)12(-), having an exact match with the corresponding calculated spectrum. Tandem mass spectrometry of the molecular ion peak up to MS(9) was performed. Complex isotope distributions in each of the mass peaks confirmed the alloy composition. We find the Ag3Pd3(-) core to be highly stable. The composition was further supported by scanning electron microscopy, energy-dispersive spectroscopy, and X-ray photoelectron spectroscopy.