Thermolabile Molecules Research Articles

Efficient Screening of Pharmaceutical Cocrystals by Microspacing In-Air Sublimation.

Cocrystal screening and single-crystal growth remain the primary obstacles in the development of pharmaceutical cocrystals. Here, we present a new approach for cocrystal screening, microspacing in-air sublimation (MAS), to obtain new cocrystals and grow high-quality single crystals of cocrystals within tens of minutes. The method possesses the advantages of strong designable ability of devices, user-friendly control, and compatibility with materials, especially for the thermolabile molecules. A novel drug-drug cocrystal of favipiravir (FPV) with salicylamide (SAA) was first discovered by this method, which shows improved physiochemical properties. Furthermore, this method proved effective in cultivating single crystals of FPV-isonicotinamide (FPV-INIA), FPV-urea, FPV-nicotinamide (FPV-NIA), and FPV-tromethamine (FPV-Tro) cocrystals, and the structures of these cocrystals were determined for the first time. By adjusting the growth temperature and growth distance precisely, we also achieved single crystals of 10 different paracetamol (PCA) cocrystals and piracetam (PIR) cocrystals, which underscores the versatility and efficiency of this method in pharmaceutical cocrystal screening.

Soft robotic patterning of liquids

Patterning of two or more liquids, either homogeneous in each phase or mixed with particles (including biological matter, such as cells and proteins), by controlling their flow dynamics, is relevant to several applications. Examples include dynamic spatial confinement of liquids in microfluidic systems (such as lab-on-a-chip and organ-on-a-chip devices) or structuring of polymers to modulate various properties (such as strength, conductivity, transparency and surface finishing). State-of-the-art strategies use various technologies, including positioners, shakers and acoustic actuators, which often combine limited versatility of mixing with significant inefficiency, energy consumption, and noise, as well as tendency to increase the temperature of the liquids. Here, we describe a new kind of robotic mixers of liquids, based on electro-responsive smart materials (dielectric elastomer actuators). We show for the first time how an efficient soft robotic device can be used to produce, via combinations of rotations and translations, various spatial patterns in liquids and maintain them stable for a few minutes. Moreover, we show that, as compared to a conventional orbital shaker, the new type of robotic device can mix liquids with a higher efficacy (~ 94% relative to ~ 80%, after 8 min of mixing) and with a significantly lower increase of the liquids’ temperature (+ 1 °C relative to + 5 °C, after 6 h of mixing). This is especially beneficial when mixing should occur according to controllable spatial features and should involve temperature-sensitive matter (such as biological cells, proteins, pre-polymers and other thermolabile molecules).

Kinetic study of some flavor and bioactive compounds during fermentation of Parkia biglobosa

Seeds of Parkia biglobosa, also called African locust bean seeds (ALBSs), are used to produce a condiment by subjecting them to cooking, alkaline fermentation, and drying. The objective of this work was to monitor the evolution of the composition of ALBS during spontaneous fermentation. Results showed that the seeds at initial time contained mainly proteins (45%) and lipids (36%). During fermentation, the conversion of this initial composition took place to produce ammonium, free amino acids, and short-chain fatty acids (SCFAs) that accounted for 2%, 10%, and 1.5% dry basis, respectively, which explains the typical flavor of fermented ALBS. This release was also concomitant with biogenic amine production. Along with ammonium and SCFA release, pyrazines were the major volatile organic compounds. Composition modulation was well represented by a Verhulst equation in which the dynamic could be modified by the initial composition of the seed, the pre-cooking step, and the fermentation conditions (temperature and aeration). Practical applications For nutritional and sustainability reasons, re-integration of vegetable in general and legumes, in particular in our diet, is a current trend. However, only dish that are appealing to consumers are concretely adopted. Fermentation of legumes is interesting in more than one way. It does not involve heat, so thermolabile molecules can be preserved and the financial and energetic costs are low. Biological interesting molecules are produced from the metabolism of microorganisms, and among them, volatile organic compounds that contribute to enhance the flavor of legume dishes. Soumbala is a meal and/or condiment obtained from the fermentation of African locust bean (Parkia biglobosa) seeds (ALBSs), a legume widely spread in west Africa. The final typical sensory properties are known to be due to the alkaline fermentation step by Bacillus species. This paper presents the kinetics of release of various compounds during spontaneous fermentation that may impact the sensory and nutritional properties of the final product. More precisely, the dynamic of the production of free amino acids, free fatty acids, volatile compounds, and biogenic amines are presented and discussed to give insights into the succession of important reactions to control the sensory and nutritional qualities of fermented ALBS (FALBS). This paper shows that further research are necessary to better understand and control the quality of not only FALBS but also similar protein-rich legumes to modulate the quality of the final products.

Analytical evaluation of carotenoids, apocarotenoids, capsaicinoids, and phenolics to assess the effect of a protective treatment on chili peppers dried at different temperatures

The most common preservation process for chili peppers is drying, which inevitably causes oxidative degradation of thermolabile molecules. The aim of this research was to evaluate the potential protective effect exerted by an active ingredient based on grape seed oil, on pepper fruits. Grapeseed oil is rich in antioxidant compounds and was applied to pepper’s surface in form of a sol–gel product, before fruit thermal treatment. In this work, chili peppers samples were preventively treated with an active solution, and controls (untreated peppers), were submitted to a drying process performed at two different temperatures: 45 and 65 °C. Analysis of capsaicinoids, carotenoids, apocarotenoids, and phenolic content was performed to evaluate possible differences between the sets of samples. Oxidative stability of oil enriched with chili pepper powder aliquots was also measured to evaluate the antioxidant power of the samples. Obtained data showed that treated samples retained a higher amount of capsaicinoids and carotenoids. Oxidative stability of pepper powder was also higher for treated samples than for controls. Furthermore, the thermal treatment performed at 45 °C caused milder modifications than the 65 °C treatment. The applied pre-drying treatment can be proposed to prevent bioactive compounds loss and to enhance product stability and shelf-life.

Currently Applied Extraction Processes for Secondary Metabolites from Lippia turbinata and Turnera diffusa and Future Perspectives

The poleo (Lippia turbinata Griseb.) and damiana (Turnera diffusa Wild) are two of the most valued species in the Mexican semidesert due to their medicinal uses. The conventional essential oil extraction process is hydrodistillation, and for the extraction of antioxidants, the use of organic solvents. However, these techniques are time-consuming and degrade thermolabile molecules, and the efficiency of the process is dependent on the affinity of the solvent for bioactive compounds. Likewise, they generate solvent residues such as methanol, hexane, petroleum ether, toluene, chloroform, etc. Therefore, in recent years, ecofriendly alternatives such as ohmic heating, microwaves, ultrasound, and supercritical fluids have been studied. These methodologies allow reducing the environmental impact and processing times, in addition to increasing yields at a lower cost. Currently, there is no up-to-date information that provides a description of the ecofriendly trends for the recovery process of essential oils and antioxidants from Lippia turbinata and Turnera diffusa. This review includes relevant information on the most recent advancements in these processes, including conditions and methodological foundation.

Metabolism Dealing with Thermal Degradation of NAD+ in the Hyperthermophilic Archaeon Thermococcus kodakarensis.

NAD+ is an important cofactor for enzymatic oxidation reactions in all living organisms, including (hyper)thermophiles. However, NAD+ is susceptible to thermal degradation at high temperatures. It can thus be expected that (hyper)thermophiles harbor mechanisms that maintain in vivo NAD+ concentrations and possibly remove and/or reuse undesirable degradation products of NAD+ Here we confirmed that at 85°C, thermal degradation of NAD+ results mostly in the generation of nicotinamide and ADP-ribose, the latter known to display toxicity by spontaneously linking to proteins. The hyperthermophilic archaeon Thermococcus kodakarensis possesses a putative ADP-ribose pyrophosphatase (ADPR-PPase) encoded by the TK2284 gene. ADPR-PPase hydrolyzes ADP-ribose to ribose 5-phosphate (R5P) and AMP. The purified recombinant TK2284 protein exhibited activity toward ADP-ribose as well as ADP-glucose. Kinetic analyses revealed a much higher catalytic efficiency toward ADP-ribose, suggesting that ADP-ribose was the physiological substrate. To gain insight into the physiological function of TK2284, a TK2284 gene disruption strain was constructed and examined. Incubation of NAD+ in the cell extract of the mutant strain at 85°C resulted in higher ADP-ribose accumulation and lower AMP production compared with those in experiments with the host strain cell extract. The mutant strain also exhibited lower cell yield and specific growth rates in a synthetic amino acid medium compared with those of the host strain. The results obtained here suggest that the ADPR-PPase in T. kodakarensis is responsible for the cleavage of ADP-ribose to R5P and AMP, providing a means to utilize the otherwise dead-end product of NAD+ breakdown.IMPORTANCE Hyperthermophilic microorganisms living under high temperature conditions should have mechanisms that deal with the degradation of thermolabile molecules. NAD+ is an important cofactor for enzymatic oxidation reactions and is susceptible to thermal degradation to ADP-ribose and nicotinamide. Here we show that an ADP-ribose pyrophosphatase homolog from the hyperthermophilic archaeon Thermococcus kodakarensis converts the detrimental ADP-ribose to ribose 5-phosphate and AMP, compounds that can be directed to central carbon metabolism. This physiological role for ADP-ribose pyrophosphatases might be universal in hyperthermophiles, as their homologs are widely distributed among both hyperthermophilic bacteria and archaea.

Complementary analytical liquid chromatography methods for the characterization of aqueous phase from pyrolysis of lignocellulosic biomasses.

In this work, two analytical liquid chromatography methods were developed and compared for the characterization of aqueous phases from pyrolysis of lignocellulosic biomasses. NanoLC electron ionization-mass spectrometry (EI-MS) represents a novel and useful tool for both separation and identification of semi/nonvolatile and thermolabile molecules. The use of nanoscale flow rates, the highly reproducibility, and high detailed information on EI spectra are the principal advantages of this technique. On the other hand, comprehensive 2D-LC, providing a two-dimensional separation, increases the overall peak capacity lowering the occurrence of peak coelutions. Despite the use of reversed phase modes in both dimensions, a satisfactory degree of orthogonality was achieved by the employment of a smart design of gradient elution strategies in the second dimension in combination with photodiode array detection (PDA) and atmospheric pressure chemical ionization-mass spectrometry (APCI-MS). Because of the absence of the preliminary extraction procedure, the fingerprint obtained for these samples results is independent of the extraction yield or contamination contrary to the gas chromatography-mass spectrometry (GC-MS) approach where a liquid-liquid extraction of the water phase is necessary. The main classes of identified compounds were phenols, ketones, furans, and alcohols. The synergistic information on the two powerful analytical approaches, e.g., NanoLC EI-MS and LC × LC, in the identification of such complex samples has never been investigated and fully benefit on the one hand from the superior degree of mass spectral information from EI-MS and on the other hand from enhanced LC × LC compound separation.

Supercritical Fluid Processing of Materials for Regenerative Medicine

In the field of regenerative medicine scaffolds, supercritical fluids (SCFs) were initially utilised to create microporous biodegradable polymeric foams. Supercritical carbon dioxide (scCO2) and other SCFs were desirable solvents as they were inexpensive, non-toxic, non-flammable, did not leave residues and could be reclaimed from other processes. Additionally, the plasticisation of polymers by scCO2 provided opportunities for the incorporation of thermolabile molecules. Whilst SCFs are still well utilised in the production of foamed matrices their versatility has enabled their use in other processes such as microparticle formation, polymer purification, sterilisation, tissue decellularisation and medical device coatings. Recent patents and innovative developments pertaining to the use of supercritical fluids in these processes are discussed herein. Keywords: Drug delivery, polymer processing, regenerative medicine, scaffolds, supercritical fluids, tissue engineering.

The science and clinical translation of remote postconditioning

The treatment of reperfusion injury requires measures beyond timely reperfusion. Conventional postconditioning (PostC) of ischemic tissues offers a strategy to reduce reperfusion injury, but its adoption is challenged by requiring access and imposing additional ischemia to the ischemic organ. Generating protective signals by PostC in a tissue remote from the target organ such as the limb, i.e. remote PostC (rPostC), may present an alternative approach to exerting endogenous tissue protection. Because rPostC is only recently reported, the fundamental biology of rPostC is not well understood, and studies to date are largely observational. rPostC has been observed to reduce ischemia-reperfusion injury experimentally in heart, kidney, brain and skeletal muscle in multiple species, including rat, rabbit and pig. Both necrosis and apoptosis are reduced. As in remote ischemic preconditioning, rPostC requires a transfer or communication of protective factors or signals through humoral and/or neural pathways. Triggers of target organ protection include G-protein-coupled receptor ligands, metabolites of ischemia, or small thermolabile molecules. Some evidence suggests that reperfusion injury salvage kinases may be involved in rPostC, in agreement with both preconditioning and conventional PostC. Clinical studies investigating improvements in clinical outcomes or biomarkers with rPostC are encouraging.

VUV state-selected photoionization of thermally-desorbed biomolecules by coupling an aerosol source to an imaging photoelectron/photoion coincidence spectrometer: case of the amino acids tryptophan and phenylalanine

Gas phase studies of biological molecules provide structural and dynamical information on isolated systems. The lack of inter- or intra-molecular interactions facilitates the interpretation of the experimental results through theoretical calculations, and constitutes an informative complement to the condensed phase. However advances in the field are partially hindered by the difficulty of vaporising these systems, most of which are thermally unstable. In this work we present a newly developed aerosol mass thermodesorption setup, which has been coupled to a Velocity Map Imaging (VMI) analyzer operated in coincidence with a Wiley-McLaren Time of Flight spectrometer, using synchrotron radiation as a single photon ionization source. Although it has been previously demonstrated that thermolabile molecules such as amino acids can be produced intact by the aerosol vaporisation technique, we show how its non-trivial coupling to a VMI analyzer plus the use of electron/ion coincidences greatly improves the concept in terms of the amount of spectroscopic and dynamic information that can be extracted. In this manner, we report on the valence shell ionization of two amino acids, tryptophan and phenylalanine, for which threshold photoelectron spectra have been recorded within the first 3 eV above the first ionization energy using synchrotron radiation emitted from the DESIRS beamline located at SOLEIL in France. Their adiabatic ionization energies (IEs) have been measured at 7.40 ± 0.05 and 8.65 ± 0.02 eV, respectively, and their spectra analyzed using existing theoretical data from the literature. The IE values agree well with previously published ones, but are given here with a considerably reduced uncertainty by up to a factor of 5. The photostability of both amino acids is also described in detail, through the measurement of the state-selected fragmentation pathways via the use of threshold electron/ion coincidences (TPEPICO), with appearance energies for the different photofragments given for the vaporization temperatures studied, in correlation with the different molecular orbitals involved as identified from the Threshold Photoelectron Spectra (TPES).

Isolation and structural analysis of the cyclic fatty acid monomers formed from eicosapentaenoic and docosahexaenoic acids during fish oil deodorization

Long-chain polyunsaturated fatty acids (LC-PUFAs) present in fish oils are thermolabile molecules. Among the degradation reactions encountered, thermal cyclization occurs during refining or other heat treatments. Numerous studies have been carried out in the past to quantify and determine the structures of cyclic fatty acid monomers (CFAMs) formed from oleic, linoleic and linolenic acids in heated vegetable oils. Recently, much attention have been given to LC-PUFAs due to their potential health benefits. However, data on quantification of CFAMs formed from these fatty acids, such as eicosapentaenoic acid (EPA, cis-5, cis-8, cis-11, cis-14, cis-17 20:5) and docosahexaenoic acid (DHA, cis-4, cis-7, cis-10, cis-13, cis-16, cis-19 22:6), the two main LC-PUFAs in fish oils, are scarce. In the present study, structural analyses of CFAMs formed from EPA and DHA during the deodorization of fish oil are presented. Fish oil sample was deodorized at 250 °C for 3 h under a pressure of 1.5 mbar in a laboratory deodorizer. The CFAMs formed during heat treatment of fish oil were isolated by a combination of saponification, esterification, urea fractionations and column chromatography. Structural analyses of C20- and C22-CFAMs were achieved by gas-chromatography electronic-ionization mass-spectrometry (GC-EI-MS) of their 4,4-dimethyloxazoline (DMOX) derivatives. We identified seven out of 13 possible structures of hydrogenated CFAMs formed from EPA, and nine out of 16 possible structures of CFAM formed from DHA. Major CFAMs from both EPA and DHA were cyclohexyl isomers. All possible cyclohexyl isomers were found but only nine out of 18 of the cyclopentyl isomers were present in concentration sufficient for identification. Chemical mechanisms involved in the formation of polyunsaturated LC-PUFAs have been investigated. The results have shown that general principle involved in the cyclization of LC-PUFAs is same as that for the thermal cyclization of oleic, linoleic and α-linolenic acids.

Structure Elucidation and Quantification by HPLC Ion-Trap Multiple Mass Spectrometry

High-performance liquid chromatography combined with ion-trap mass spectrometry allows now to record mass spectra in the full-scan mode with sample quantities around 1 ng or less. In addition, the on-line generation of multiple daughter-ion spectra has become possible. This has revolutionised organic trace analysis. Small polar and/or thermolabile molecules such as biotoxins can be detected in the trace range and their structure unequivocally identified. In addition, a complete or partial structure elucidation of related structures is possible. This overview demonstrates the possibilities and limitations of HPLC-Msn with ion traps. Examples shown are the structure elucidation of aconitum alkaloids, trichothecenes and photochemically formed carbonyl compounds. This survey discusses also problems caused by system contamination and adsorption effects. Finally, the achievable performance of quantification in the trace range is briefly presented.

Capillary supercritical fluid chromatography of pyrethrins and pyrethroids with positive pressure and negative temperature gradients

Thermolabile molecules, such as pyrethrins and pyrethroids, are separated by supercritical fluid chromatography (SFC). Chromatography on a fused-silica capillary column is optimized by measuring the retention factor, peak resolution, and plate height. The separation of pyrethrins and pyrethroids by SFC was accomplished by the use of a positive pressure gradient and negative temperature gradient. The density of both gradients was comparable. Efficient separation of these pesticides was obtained with a linear isothermal pressure gradient of 0.2 MPa/min from 11.1 up to 22.3 MPa at a temperature of 90°C. Using a negative isobaric temperature gradient with a rate of −1.2°C/min from 130 down to 80°C, and holding 80°C for 10 min at 16.2 MPa, good results were obtained. The retention times of the analytes were constant.

Low-Temperature X-ray Structure Techniques for the Characterization of Thermolabile Molecules

Low-Temperature X-ray Structure Techniques for the Characterization of Thermolabile Molecules

Low-temperature x-ray structure techniques for the characterization of thermolabile molecules

Low-temperature x-ray structure techniques for the characterization of thermolabile molecules

Effect of heat-hardening on thermostability of acid phosphatase from wheat leaves

1. 1.|Acid phosphatase (APase) of hardened and non-hardened wheat seedlings is a two-component system with respect to their thermostability. 2. 2.|After heat-hardening the relative content of the thermostable component is increased. 3. 3.|Isoenzyme composition of APase was the same in hardened and non-hardened plants. 4. 4.|Sodium fluoride had a greater protective effect on thermoresistance of APase from non-hardened plants. 5. 5.|Analysis of the data obtained indicated that an increase in thermostability of APase produced by heat-hardening is attributed to both stabilization and partial elimination of thermolabile molecules of APase.

Effect of 5-fluorouracil on the gibberellic acid-induced formation of thermolabile α-amylase molecules in barley aleurone cells

IN the aleurone cells of barley seed gibberellic acid (GA) stimulates de novo synthesis of α-amylase1,2. From a study of the effect of 5-fluorouracil (FU) on the GA-evoked formation of thermolabile α-amylase molecules, Carlson3 has concluded that GA regulates a post-transcriptional control point. According to my interpretation, which differs from Carlson's conclusion, his data (Fig. 1, ref. 3) suggest that mRNAs for α-amylase are formed in the presence of GA.