Nanosensors for Continuous and Noninvasive Monitoring of Mesenchymal Stem Cell Osteogenic Differentiation

Assessing mesenchymal stem cell (MSC) differentiation status is crucial to verify therapeutic efficacy and optimize treatment procedures. Currently, this involves destructive methods including antibody‐based protein detection and polymerase chain reaction gene analysis, or laborious and technically challenging genetic reporters. Development of noninvasive methods for real‐time differentiation status assessment can greatly benefit MSC‐based therapies. This report introduces a nanoparticle‐based sensing platform that encapsulates two molecular beacon (MB) probes within the same biodegradable polymeric nanoparticles. One MB targets housekeeping gene glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) as an internal reference, while another detects alkaline phosphatase (ALP), a functional biomarker. Following internalization, MBs are gradually released as the nanoparticle degrades. GAPDH MBs provide a stable reference signal throughout the monitoring period (18 days) regardless of differentiation induction. Meanwhile, ALP mRNA undergoes well‐defined dynamics with peak expression observed during early stages of osteogenic differentiation. By normalizing ALP‐MB signal with GAPDH‐MB, changes in ALP expression can be monitored, to noninvasively validate osteogenic differentiation. As proof‐of‐concept, a dual‐colored nanosensor is applied to validate MSC osteogenesis on 2D culture and polycaprolactone films containing osteo‐inductive tricalcium phospate.     

Quasilinearization and optimal control problems with a state inequality constraint

A quasilinearization algorithm is proposed to solve optimal control problems with a scalar control variable and a scalar state inequality constraint using a transformation technique. The optimal control problem is transformed into an unconstrained problem with an auxiliary control and higher dimension. Owing to the appearance of singular arcs in the transformed problem, this is made non-singular by adding a quadratic function of the auxiliary control to its performance index. A proposed quasilinearization algorithm is then used to generate a series of solutions of the modified transformed problems which converges to the solution of the original state-constrained problem in the limit. A distinct feature of the proposed algorithm is that each modified transformed problem yields a feasible trajectory of the original state-constrained problem. Two examples have been solved to illustrate the effectiveness of the proposed quasilinearization algorithm.     

Plant-wide control for the economic operation of modified single mixed refrigerant process for an offshore natural gas liquefaction plant

The marine operation of floating liquefied natural gas (FLNG) demands process compactness, flexibility, simplicity of operation, safety, and higher efficiency. The modified single mixed refrigerant (MSMR) process satisfies the FLNG process requirements and is accepted as a suitable technology for FLNG operation. The aim of this study was to develop a plant-wide control structure or strategy that can sustain the economic efficiency of the MSMR process. The NGL recovery and liquefaction units were integrated in the MSMR process to provide a compact plant structure with an efficient operation. Steady-state optimality analysis was intensively conducted in a rigorous dynamic simulation environment to determine the correct variable to sustain the economic efficiency of MSMR process. The results showed that the flow rate ratio of heavy and light mixed refrigerant (HK/LK ratio) is a promising self-optimizing controlled variable. Controlling this variable can sustain the MSMR optimality, even when the process is operated under off-design operating conditions or in the presence of disturbances. Based on the control structure tests, the control configuration with the HK/LK ratio loop showed excellent performance, maintaining the process stability against a range of disturbances. The proposed approach can also be applied to any cryogenic liquefaction technology for determining a possible optimizing controlled variable.     

Computer-based engineering of thermostabilized antibody fragments

We used the molecular modeling program Rosetta to identify clusters of amino acid substitutions in antibody fragments (scFvs and scAbs) that improve global protein stability and resistance to thermal deactivation. Using this methodology, we increased the melting temperature (T_m) and resistance to heat treatment of an antibody fragment that binds to the Clostridium botulinum hemagglutinin protein (anti-HA33). Two designed antibody fragment variants with two amino acid replacement clusters, designed to stabilize local regions, were shown to have both higher T_m compared to the parental scFv and importantly to retain full antigen binding activity after 2 hr of incubation at 70°C. The crystal structure of one thermostabilized scFv variants was solved at 1.6 Å and shown to be in close agreement with the RosettaAntibody model prediction.     

Sodium azide and metal chelator effects on 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) radical scavenging compounds from methylene blue photosensitized lard

Thermal oxidation of edible oils can generate 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) radical scavenging compounds from oxidized lipids (RSOLs). However, effects of photosensitization on the formation of RSOLs have not been reported yet. Methylene blue (MB) photosensitization and involvement of singlet oxygen and transition metals on the RSOL formations were determined in stripped lard oils. RSOLs were formed in lard containing MB and visible light irradiation only. Addition of sodium azide decreased RSOLs with concentration dependent manner, which implies singlet oxygen was involved on the RSOL formation. Ethylenediammetetraacetic acid (EDTA), a well known metal chelator, accelerated the formation of RSOLs through protecting the decomposition of MB photosensitizer. Results from p‐anisidine values showed that RSOLs from photosensitization may not be formed from the same pathways compared to thermal oxidation. Practical application: Understanding mechanisms of lipid oxidation can help extend the shelf‐life of foods. Photosensitization plays important roles in accelerating the rates of lipid oxidation. The results of this study showed that foods containing photosensitizers can generate radical scavenging compounds from oxidized lipids (RSOLs) under visible light irradiation and singlet oxygen is involved in the formations of these compounds. However, these compounds may not share the same pathways with thermally oxidized lipids. Metal chelating agents accelerated the rates of lipid oxidation and formation of RSOLs which implies that metal chelators can act as prooxidant. Careful considerations are necessary on the addition of metal chelators because non‐polar photosensitizers can act a prooxidant.     

Application of solid phase-microextraction (SPME) and electronic nose techniques to differentiate volatiles of sesame oils prepared with diverse roasting conditions

Headspace volatiles of sesame oil (SO) from sesame seeds roasted at 9 different conditions were analyzed by a combination of solid phase microextraction (SPME)-gas chromatography/mass spectrometry (GC/MS), electronic nose/metal oxide sensors (MOS), and electronic nose/MS. As roasting temperature increased from 213 to 247 °C, total headspace volatiles and pyrazines increased significantly (P < 0.05). Pyrazines were major volatiles in SO and furans, thiazoles, aldehydes, and alcohols were also detected. Roasting temperature was more discrimination factor than roasting time for the volatiles in SO through the principal component analysis (PCA) of SPME-GC/MS, electronic nose/MOS, and electronic nose/MS. Electronic nose/MS showed that ion fragment 52, 76, 53, and 51 amu played important roles in discriminating volatiles in SO from roasted sesame seeds, which are the major ion fragments from pyrazines, furans, and furfurals. SO roasted at 213, 230, and 247 °C were clearly differentiated from each other on the base of volatile distribution by SPME-GC/MS, electronic nose/MOS, and electronic nose/MS analyses. Practical Application: The results of this study are ready to apply for the discriminating samples using a combinational analysis of volatiles. Not only vegetable oils prepared from roasting process but also any food sample possessing volatiles could be targets for the SPME-GC/MS and electronic nose assays. Contents and types of pyrazines in sesame seed oil could be used as markers to track down the degree of roasting and oxidation during oil preparation.     

Characterization of brewing microorganisms isolated from Korean traditional <Emphasis Type="Italic">nuruk</Emphasis> for <Emphasis Type="Italic">Cheongju</Emphasis> production

The objective of this study was to select and identify yeasts and molds isolated from traditional nuruk and to investigate their brewing characteristics for Cheongju production. The yeast strains Y190 (Accession ID-KACC 93251P), Y263 (Accession ID-KACC 93252P), and Y270 (Accession ID-KACC 93253P) showing high alcohol and flavor productivity were isolated and identified by phylogenetic inference based on an internal transcribed spacer 2 region sequence analysis. In addition, Aspergillus oryzae 83-10 (Accession ID-KACC 93254P) showing the highest enzyme activity was isolated. This study provides basic data for the production of Korean Cheongju and assesses the applicability of these three yeast strains and A. oryzae 83-10 isolated from traditional nuruk.     

Chromatographic analysis of ceramide III ofSaccharomyces cerevisiae

Ceramide was prepared by the cultivation ofSaccharomyces cerevisiae from cell extracts by solvent exfraction and analyzed by NP-HPLC using a UV detector. The mobile phase was composed of hexane, methanol, and IPA. From the experimental conditions, the composition of mobile phase was 72/5/23 (hexane/IPA/methanol, vol%). Quantitative analysis of ceramide was performed. Based on the analytical conditions, the effect of cultivation temperature for the production of ceramide was investigated and the optimum cultivation temperature was found to be 35°C. This paper is dedicated to Dr. Youn Yong Lee on the occasion of his retirement from Korea Institute of Science and Technology.