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To improve the synthetic ability of E. coli L-isoleucine by combining λ-Red recombination with complex mutagenesis. Firstly, taking E. coli NXA as the original strain, λ-Red homologous recombination was used to knock out the coding gene brnQ of branched chain amino acid transport protein to obtain mutant strain E. coli NXA1. Secondly, E. coli NXA1 was subjected to multiple rounds of complex mutagenesis with atmospheric room temperature plasma (ARTP), ultraviolet (UV), and nitrosoguanidine (NTG), which was screened to obtain the mutant strain E. coli NXA2 with α-AB being structural analogue. The fermentation results showed that L-isoleucine titer of E. coli NXA1 was 2.76 g/L, which was 33.98% higher than E. coli NXA after fermentation for 40 hours at 37 ℃ and 200 r/min. The L-isoleucine titer of E. coli NXA2 was 3.22 g/L, which was 16.67% higher than E. coli NXA1 and 56.31% higher than E. coli NXA. After 20 continuous passages of E. coli NXA2, the good genetic stability could be reflected. The combination of λ-Red recombination and complex mutagenesis has a significant effect on improving the synthetic ability of E. coli L-isoleucine, laying a theoretical basis for the breeding of L-isoleucine high-producing strains.

L-isoleucine, an essential amino acid, is widely used in the pharmaceutical and food industries. However, the current production efficiency is insufficient to meet the increasing demands. In this study, we aimed to develop an efficient L-isoleucine-producing strain of Escherichia coli. First, accumulation of L-isoleucine was achieved by employing feedback-resistant enzymes. Next, a growth-coupled L-isoleucine synthetic pathway was established by introducing the metA - metB -based α-ketobutyrate-generating bypass, which significantly increased L-isoleucine production to 7.4 g/L. Upon employing an activity-improved cystathionine γ-synthase mutant obtained from adaptive laboratory evolution, L-isoleucine production further increased to 8.5 g/L. Subsequently, the redox flux was improved by bypassing the NADPH-dependent aspartate aminotransferase pathway and employing the NADH-dependent pathway and transhydrogenase. Finally, L-isoleucine efflux was enhanced by modifying the transport system. After fed-batch fermentation for 48 h, the resultant strain, ISO-12, reached an L-isoleucine production titer of 51.5 g/L and yield of 0.29 g/g glucose. The strains developed in this study achieved a higher L-isoleucine production efficiency than those reported previously. These strategies will aid in the development of cell factories that produce L-isoleucine and related products. [Display omitted] • Growth-coupled production of L-isoleucine was achieved in Escherichia coli. • Feedback regulation of key enzymes in L-isoleucine synthesis was abrogated. • A growth-coupled L-isoleucine synthetic pathway was established. • The redox flux was improved via NADH dependent pathways and transhydrogenase. • High production titer and yield of L-isoleucine was fulfilled. [ABSTRACT FROM AUTHOR]

l‐Isoleucine is subjected to density and sound velocity measurements in aqueous solutions of potassium (KCl) and sodium chlorides (NaCl) as well as water, from 288.15 to 298.15 K. Through the use of these experimental data, a variety of volumetric and acoustical parameters are calculated, including values for molar volume (Vm), available volume (Va), adiabatic compressibility (β), nonlinear parameters (B/A), Wada constant (W), Rao's constant (R), and van der Wall's constant (a). The findings shed light on many electrostatic and hydrophobic interactions within the binary system involving electrolytes, water, and amino acids. The interactions between the electrolytes and l‐isoleucine are thoroughly examined under varying electrolyte concentrations and temperatures, revealing many interactions. Positive transfer quantities demonstrate the ion‐hydrophilic and solute–solvent interaction in the aqueous medium, particularly between l‐isoleucine and NaCl and KCl. The influence of electrolyte type and temperature on these interactions is discernible. The culmination of the study's outcomes is elaborated through analyzing solute–solvent and solute–solute interactions. In particular, the interaction hierarchy is identified as l‐isoleucine + water + KCl > l‐isoleucine + water + NaCl > l‐isoleucine + swater. [ABSTRACT FROM AUTHOR]

This study consists of four steps. In the first, two different biocompatible organogelators were synthesized, starting with the L-isoleucine amino acid to obtain amide compounds. In the second step, the gelation potential of synthesized organogelators with fatty acid esters and organic solvents was investigated. These esters were chosen as gelation liquids due to their biocompatibility and also their penetration-enhancing properties when the drug is administered via the skin. After the minimum gel concentrations (MGCs) of the organogelators were determined, the melting point of gel Tg was found, and then, AHg gelation enthalpy values were found by means of the Van't Hoff equation. In addition to the gelation abilities and capacities of the organogelators being thus synthesized, their thermal stabilities were also determined. In the third stage of the study, the network which occurred during the formation of the gels was screened by an SEM device, and their characterizations were determined. In the study's fourth stage, the gels were loaded with ibuprofen and naproxen -- known for their non-steroidal antiinflammatory and analgesic effects -- and their drug-loading capacities were thus determined. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, EFSA was asked to deliver a scientific opinion on the safety and efficacy of l‐isoleucine produced by fermentation with Corynebacterium glutamicum CGMCC 20437 as a nutritional feed additive for use in feed and in water for drinking for all animal species. The production strain is non‐genetically modified, qualifies for the QPS approach to safety assessment when used for production purposes, is susceptible to the relevant antibiotics and contains no antimicrobial resistance genes of concern. No viable cells of the production strain were detected in the final product. The additive does not give rise to any safety concern regarding the production strain. l‐Isoleucine produced by fermentation with Corynebacterium glutamicum CGMCC 20437 is considered safe for the target species, the consumer and the environment. Regarding the use in water, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) reiterates its concerns over the safety for the target species of l‐isoleucine administered simultaneously via water for drinking and feed owing to the risk of nutritional imbalances and hygienic reasons. In the absence of data, the FEEDAP Panel is not in a position to conclude on the potential of l‐isoleucine produced by fermentation with Corynebacterium glutamicum CGMCC 20437 to be irritant to skin and/or eyes, or as a dermal sensitiser. Due to the high dusting potential, exposure by inhalation is likely. l‐Isoleucine produced by fermentation with Corynebacterium glutamicum CGMCC 20437 is considered as an efficacious source of the essential amino acid l‐isoleucine for non‐ruminant animal species. For the supplemental l‐isoleucine to be as efficacious in ruminants as in non‐ruminant species, it would require protection against degradation in the rumen. [ABSTRACT FROM AUTHOR]

One of the most important industrial requirements is to find the materials which have the tendency to withstand harsh environments, such as high pressure, high temperature, and radiation environments, so that they will maintain high efficiency in these conditions. Such materials are greatly required for aerospace, defense, and high-temperature molecular device fabrications. In the present context, we have chosen l-isoleucine samples to assess the crystallographic phase stability under dynamic shocked conditions. The stability result has been screened by a powder x-ray diffractometer (PXRD). Based on the observed diffraction results, it is well evidenced by the cleavage planes, such as (002) (003), (004), (005), and (006), that the applied shock waves do not induce any crystallographic phase transitions and lattice deformations even at 200 shocks. The unit cell dimensions of the l-isoleucine sample were a = 9.752 Å, b = 5.321 Å, c = 14.13 Å and V = 728 Å3 which were found to be the same under all numbers of shocked conditions. Hence, the l-isoleucine sample is strongly suggested for device fabrication, and previous static high-pressure experimental results also support this suggestion. [ABSTRACT FROM AUTHOR]

Abstract Background The roles of gut microbiota in the pathogenesis of SLE have been receiving much attention during recent years. However, it remains unknown how fecal microbiota transplantation (FMT) and microbial metabolites affect immune responses and lupus progression. Methods We transferred fecal microbiota from MRL/lpr (Lpr) mice and MRL/Mpj (Mpj) mice or PBS to pristane-induced lupus mice and observed disease development. We also screened gut microbiota and metabolite spectrums of pristane-induced lupus mice with FMT via 16S rRNA sequencing, metagenomic sequencing, and metabolomics, followed by correlation analysis. Results FMT from MRL/lpr mice promoted the pathogenesis of pristane-induced lupus and affected immune cell profiles in the intestine, particularly the plasma cells. The structure and composition of microbial communities in the gut of the FMT-Lpr mice were different from those of the FMT-Mpj mice and FMT-PBS mice. The abundances of specific microbes such as prevotella taxa were predominantly elevated in the gut microbiome of the FMT-Lpr mice, which were positively associated with functional pathways such as cyanoamino acid metabolism. Differential metabolites such as valine and L-isoleucine were identified with varied abundances among the three groups. The abundance alterations of the prevotella taxa may affect the phenotypic changes such as proteinuria levels in the pristane-induced lupus mice. Conclusion These findings further confirm that gut microbiota play an important role in the pathogenesis of lupus. Thus, altering the gut microbiome may provide a novel way to treat lupus.

Trehalose is commonly used as a protein stabilizer in spray dried protein formulations delivered via the pulmonary route. Spray dried trehalose formulations are highly hygroscopic, which makes them prone to deliquescence and recrystallization when exposed to moisture, leading to impairment in aerosolization performance. The main aim of this study was to investigate and compare the effect of hydrophobic amino acids (i.e. L-leucine and L-isoleucine) in enhancing aerosolization performance and in mitigating moisture-induced changes in spray dried trehalose formulations. Trehalose was spray dried with 20-60% w/w of amino acid (i.e. L-leucine or L-isoleucine). The spray dried formulations were stored at 25 °C/50% RH for 28 days. Solid state characterization and in vitro aerosolization performance studies were performed on the spray dried formulations before and after storage. The addition of 20-60% w/w of amino acid (i.e. L-leucine or L-isoleucine) improved the emitted fractions of spray dried trehalose formulations from a dry powder inhaler. However, ≥ 40% w/w of L-leucine/L-isoleucine was needed to prevent recrystallization of trehalose in the formulations when exposed to 25 °C/50% RH for 28 days. X-ray photoelectron spectroscopy (XPS) demonstrated that samples with 40-60% w/w L-isoleucine had more amino acid on the surfaces of the particles compared to their L-leucine counterparts. This may explain the greater ability of the L-isoleucine (40-60% w/w) samples to cope with elevated humidity compared to L-leucine samples of the same concentrations, as observed in the dynamic vapour sorption (DVS) studies. In conclusion, this study demonstrated that both L-leucine and L-isoleucine were effective in enhancing aerosolization performance and mitigating moisture-induced reduction in aerosolization performance in spray dried trehalose formulations. L-isoleucine proved to be superior to L-leucine in terms of its moisture protectant effect when incorporated at the same concentration in the formulations.

Molecular asymmetric imprinting polymerization in the cholesteric liquid crystal was carried out. An L-isoleucine derivative with fluorine was synthesized for a chiral inducer to induce the cholesteric liquid crystal from the nematic liquid crystal. A substance with multiple asymmetric centers can be obtained conveniently to use amino acid as chiral substance. The L-isoleucine derivative induced helical structure for 4-cyano-4'-hexylbiphenyl (6CB). Electrochemical preparation of π-conjugated polymers was carried out in the cholesteric liquid crystal. Fingerprint texture derived from the cholesteric liquid crystal was transcribed to the polymers. UV-vis and circular dichroism (CD) spectra of oxidized (doped) for and reduced (dedoped) form of the polymer were measured. Electrochemical polymerization in the helical liquid crystal produced the molecular aggregation imprinted chiral polymers, which have liquid crystal-like aggregation form, chirality with no stereogenic center via formation of polymer atropisomer, and electrochemical driven change in optical activity, as a form of electro-optically coloration active polymers. Finally, the polarons and bipolarons are in the form of chiral structure. These charge carriers in the polymers can be determined as chiral charge carrier "chiralions". [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on l‐isoleucine produced by Corynebacterium glutamicum KCCM 80185 for all animal species. In 2021, the FEEDAP Panel issued an opinion on the safety and efficacy of the product. In that assessment, the FEEDAP Panel could not exclude the potential presence of recombinant DNA derived from the genetically modified production organism in the additive. The applicant provided supplementary data to exclude the presence of recombinant DNA derived from the production organism in the final product. Based on the data provided, the FEEDAP Panel concluded that no DNA of the production strain C. glutamicum KCCM 80185 was detected in the additive. [ABSTRACT FROM AUTHOR]

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As one of the three important branched-chain amino acids, l-isoleucine has a wide range of applications in the fields of medicine, food, and feed. Currently, the production of l-isoleucine is well-studied by Corynebacterium glutamicum , while the autonomous and efficient production of l-isoleucine in Escherichia coli has not been reported. Here, we developed a production strategy that combined metabolic engineering with bacterial quorum sensing to achieve the efficient production of l-isoleucine. First, we enhanced the l-isoleucine synthesis pathway by overexpressing the genes ilvIH1 , CgilvA1 , and ygaZH . Second, the precursor supply was increased by knocking out the gene rhtC , while deletion of the gene livJ was implemented to maximize the accumulation of l-isoleucine. Finally, the artificial quorum sensing system was applied to the efficient production of l-isoleucine, and self-induced protein expression in E. coli was realized through self-regulation during fermentation. In this study, an l-threonine high-yielding strain of E. coli TWF106 was used as the starting strain, and the final strain TWF127/pST1011, pST1042-IH1ZHA1 obtained 49.3 g/L l-isoleucine with a yield of 0.32 g/g glucose and a productivity of 1.03 g/(L·h). This autonomous production strategy without the addition of inducers can also be used in other biosynthetic pathways to increase yields while also providing the possibility for various natural products to be applied to industrial production.

• Volumetric, Viscometrical, Conductometric, and Surface tension studies of amino acids in aq. [TCP][HCl] solutions. • Thermodynamics of viscous flow has been discussed using these measurements. • Study elucidates interactions of amino acids with [TCP][HCl] in aq. media. • L-Ile shows structure breaking and L-Pro shows structure making nature. • Computational and Spectroscopic studies have also verified these interactions. For efficient drug design and development, employing a holistic method that combines insights gathered from structural, thermodynamic, and biological backgrounds is essential. Understanding the energetic principles underlying molecular interactions requires employing various thermodynamic methods and physicochemical, spectroscopic, and computational analyses, particularly in the early stages of the drug development process to achieve the best possible energy interaction profile and maintain a strong pharmacological assay. The present study focused on the various molecular interactions of L-Isoleucine, an essential amino acid, and L-Proline a nonessential amino acid within varying molalities (0.001, 0.003, 0.005 mol.kg−1) in an aqueous solution of an antiplatelet drug namely ticlopidine hydrochloride [TCP][HCl] at five distinct temperature (293.15 K, 298.15 K, 303.15 K, 308.15 K, and 313.15 K) and pressure at 0.1 MPa. The volumetric properties like apparent molar volume (ϕ v), Limiting partial molar volume (ϕ v 0), Limiting the partial molar volume of transfer ( Δ ϕ v 0) , hydration number (n H), Hepler's constant (∂ ϕ E 0 /∂T) are calculated using density data; The viscosity B -coefficient, temperature derivative of the B -coefficient (d B /dT), Free activation energy of the solvent and solute (Δ μ 1 0 ≠ and Δ μ 2 0 ≠ ), entropy and enthalpy of activation (Δ S 2 0 ≠ and Δ H 2 0 ≠ ), and solvation number (n s) are all determined using the viscosity data; Walden product (Λ m 0 η 0) , limiting molar conductance (Λ m 0) , molar conductance (Λ m) was calculated using a specific conductance value; surface tension data calculates the Limiting Slope (∂σ/∂m). The UV–visible absorbance was measured, and the association constant was computed. The results of 1H NMR spectroscopy confirmed significant interactions among the protons of amino acids and Ticlopidine hydrochloride. These changes indicate a strong molecular interaction between them, which aligns with our theoretical studies. [ABSTRACT FROM AUTHOR]

Recently, the gelation of organic compounds whose molecular weights are low has been attracting considerable interest. It is known that gels are viscoelastic substances similar to liquids or solids. The main driving forces in the self-assembly of gelators include hydrogen bonding, n--n aggregation, van der Waals interactions, coordination gradients, charge transfer interactions and the other intermolecular non-covalent interactions. These interactions are non-covalent and result in the formation of supramolecular structures with nanocavities. Then, gels are formed by trapping liquid into the nano spaces in this structure. Besides contact lenses in daily life, it is one of the most beneficial systems with potential usages in the cosmetic industry, hair gels, toothpaste, shampoos and soaps, controlled drug release, sensors, food production, photography, dye removal, and tissue engineering. Gelators having low molecular mass display thermally reversible behavior. Thus, these gelators were not only academically interested but more importantly, they were of special attention because of their practical uses. For instance, studies on use of low molecular mass gelators for the removal of crude oil and domestic kitchen oil wastes poured into the sea are encountered in the literature. Herein, two low molecular mass tetraamide compounds derived from L-isoleucine were selected as organogelators. Fatty acid esters having different chain lengths such as ethyl laurate, ethyl myristate, ethyl palmitate, isopropyl laurate, isopropyl myristate, and isopropyl palmitate are exploited in the pharmaceutical and cosmetic industries as gelling liquids. Furthermore, xylene, anisole, liquid paraffin, n-dodecane, 1-decanol, diethylene glycol, and toluene have been chosen as common organic solvents. It was investigated whether the organogelators were gelled with these liquids and minimum gelling concentrations were determined. Then, Tg values of the gels were determined and AHg values of gelling enthalpy were obtained from van't Hoff correlation. In addition, the network structure of the gels was determined by the images taken with the SEM device. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety and efficacy of a feed additive consisting of l‐isoleucine produced by Corynebacterium glutamicum KCCM 80185 when used as a nutritional additive in feed and water for drinking for all animal species. The production strain is genetically modified, does not carry acquired antimicrobial resistance genes and no viable cells of the production strain were detected in the final product. The FEEDAP Panel could not exclude the presence of recombinant DNA from the production strain in the product. However, since no sequences of concern remain in the final production strain, the potential presence of recombinant DNA in the final product does not raise any safety concerns. The Panel concluded that the additive is safe for the target species, for the consumer and for the environment under the proposed conditions of use. Regarding the use in water, the FEEDAP Panel reiterated its concerns over the safety of l‐isoleucine administered simultaneously via water for drinking and feed owing to the risk of nutritional imbalances and hygienic reasons. The FEEDAP Panel concluded that l‐isoleucine produced by C. glutamicum KCCM 80185 is considered not toxic by inhalation, not irritant to skin or eyes and not a dermal sensitiser. However, due to the high dusting potential, exposure to dust per se might be a hazard for the user. l‐Isoleucine produced by C. glutamicum KCCM 80185 is considered as an efficacious source of the essential amino acid l‐isoleucine for non‐ruminant animal species. For the supplemental l‐isoleucine to be as efficacious in ruminants as in non‐ruminant species, it would require protection against degradation in the rumen. [ABSTRACT FROM AUTHOR]

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Chelating resin from naturally occurring polysaccharides like Tamarind Kernel Powder (TKP) has been synthesized. This TKP resin is highly selective, hydrophilic and biodegradable. Secondary amine derivative of tamarind kernel powder can be used as a metal ion exchanger and also as a scavenger for heavy metal ions from the effluent samples of metallurgical industries. A suitable and cost-effective method has been employed to reach the result of this experiment. L-isoleucine acts as the backbone on the tamarind kernel powder in dioxane medium. The selective chelating resins and ion exchange resins have been used in the removal of harmful heavy metal ions from effluent samples of metallurgical and mineral industries. [ABSTRACT FROM AUTHOR]

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In this study, l‐isoleucine production in Corynebacterium glutamicum WM001 was improved by deleting three genes in the genome, replacing the native promoter of ilvA in the genome, and overexpression of five genes in an alr‐based auxotrophic complementation expression system. The three genes deleted in the genome are alaT, brnQ, and alr. Deletion of alaT improved l‐isoleucine production by increasing the supply of pyruvate, whereas deletion of brnQ improved l‐isoleucine production by blocking the uptake of extracellular l‐isoleucine. Exchange of the native promoter of ilvA with promoter tac or tacM could contribute to l‐isoleucine production by increasing 2‐ketobutyric acid; tac is better than tacM for improving l‐isoleucine yield. Different combinations of the genes ilvBN, ppnK, lrp, and brnFE were overexpressed in an alr‐based auxotrophic complementation expression system to further improve l‐isoleucine production, and the best yield after 72‐H flask fermentation was obtained from the strain WM005/pYCW‐1‐ilvBN2‐ppnK1. Without addition of any antibiotics, WM005/pYCW‐1‐ilvBN2‐ppnK1 could produce 32.1 g/L l‐isoleucine after 72‐H fed‐batch fermentation, which is 34.3% increase compared with the original strain WM001. [ABSTRACT FROM AUTHOR]

Abstract Background Co-production of polyhydroxyalkanoate (PHA) and amino acids makes bacteria effective microbial cell factories by secreting amino acids outside while accumulating PHA granules inside. Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) is one of the PHAs with biocompatibility and fine mechanical properties, but its production is limited by the low level of intracellular propionyl-CoA. Results l-Isoleucine producing Corynebacterium glutamicum strain WM001 were analyzed by genome and transcriptome sequencing. The results showed that the most over-expressed genes in WM001 are relevant not only to l-isoleucine production but also to propionyl-CoA accumulation. Compared to the wild-type C. glutamicum ATCC13869, the transcriptional levels of the genes prpC2, prpD2, and prpB2, which are key genes relevant to propionyl-CoA accumulation, increased 26.7, 25.8, and 28.4-folds in WM001, respectively; and the intracellular level of propionyl-CoA increased 16.9-fold in WM001. When the gene cluster phaCAB for PHA biosynthesis was introduced into WM001, the recombinant strain WM001/pDXW-8-phaCAB produced 15.0 g/L PHBV with high percentage of 3-hydroxyvalerate as well as 29.8 g/L l-isoleucine after fed-batch fermentation. The maximum 3-hydroxyvalerate fraction in PHBV produced by WM001/pDXW-8-phaCAB using glucose as the sole carbon source could reach 72.5%, which is the highest reported so far. Conclusions Genome and transcriptome analysis showed that C. glutamicum WM001 has potential to accumulate l-isoleucine and propionyl-CoA pool. This was experimentally confirmed by introducing the phaCAB gene cluster into WM001. The recombinant strain WM001/pDXW-8-phaCAB produced high levels of PHBV with high 3-hydroxyvalerate fraction as well as l-isoleucine. Because of its high level of intracellular propionyl-CoA pool, WM001 might be used for producing other propionyl-CoA derivatives.

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Objective: The study aims at understanding the types of molecules present in one Ayurvedic formulation, Dantyarishtam by gas chromatography-mass spectrometry (GC-MS) analysis. Materials and Methods: Dantyarishtam was procured from standard Ayurvedic vendor at Chennai and was subjected to GC-MS analysis by standard procedure. Results: The GC-MS profile indicated the presence of some important biomolecules of medicinal importance such as heptanoic acid, L-Isoleucine, ethyl ester, Ethyl hydrogen succinate, 4-Ethyl hydrogen itaconate, 3-(3,3-Dimethyloxiran-2-yl)-2,2-dimethylpropionic acid, ethyl ester, pentanoic acid, 2-ethylhexyl ester, acetic acid, (3-methyl-2-nitrophenyl)methyl ester, Benzeneethanol, 4-hydroxy-, 2,5-di-tert-Butylaniline, Disulfide, di-tert-dodecyl, Methyl 4,7,10,13,16,19-docosahexaenoate, octadecanoic acid, 3-Hydroxy-4-decenoic acid, 9-(tetrahydropyran-2-yl)oxy-, 2-(trimethylsilyl)ethyl ester, and Z-10-Methyl-11-tetradecen1-ol propionate. Conclusion: The biomolecules present do have medicinal roles which represent the role of Dantyarishtam as a potent digestive formulation. [ABSTRACT FROM AUTHOR]

Abstract Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety and efficacy of l‐isoleucine produced by fermentation using Corynebacterium glutamicum KCCM 80189 when used as a nutritional additive in feed and water for drinking for all animal species. The production strain is not genetically modified. Viable cells of the production strain were not detected in the final additive. The additive does not give rise to any safety concern regarding the production strain. l‐Isoleucine produced using C. glutamicum KCCM 80189 is considered safe for the target species, for the consumer and for the environment. l‐Isoleucine produced by C. glutamicum KCCM 80189 is considered not toxic by inhalation, not irritant to skin or eyes and not a dermal sensitiser. Regarding the use in water, the FEEDAP Panel reiterates its concerns over the safety of l‐isoleucine administered simultaneously via water for drinking and feed owing to the risk of nutritional imbalances and hygienic reasons. l‐Isoleucine produced by C. glutamicum KCCM 80189 is considered as an efficacious source of the essential amino acid l‐isoleucine for non‐ruminant animal species. For the supplemental l‐isoleucine to be as efficacious in ruminants as in non‐ruminant species, it would require protection against degradation in the rumen.

Abstract Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on l‐isoleucine produced by Escherichia coli FERM ABP‐10641 (i) to renew the authorisation and to extend the use of the additive in water for drinking when used as a nutritional additive for all animal species and (ii) to evaluate a new use as a flavouring additive for all animal species. The applicant has provided evidence that the additive currently in the market complies with the existing conditions of authorisation. l‐Isoleucine does not give rise to any safety concern regarding the production strain and its genetic modification. Considering that the production process has not been substantially modified and that no adverse effects have been reported in the literature search, the FEEDAP Panel considers that there is no evidence to reconsider the conclusions reached in previous assessments. Therefore, the Panel considers that the additive l‐isoleucine produced by E. coli FERM ABP‐10641 remains safe for the target species, consumer and for the environment when used as a nutritional additive in feed. The Panel extends these conclusions to the use of the additive as a flavouring compound. Regarding the use in water, the FEEDAP Panel reiterates its concerns over the safety for the target species of l‐isoleucine administered simultaneously via water for drinking and feed owing to the risk of nutritional imbalances and hygienic reasons. l‐Isoleucine is not irritant to skin and eyes and is not a dermal sensitiser but is hazardous by inhalation. The Panel confirms the previous conclusions that the additive is considered as a source of available isoleucine for non‐ruminant animal species when used as a nutritional additive. It requires protection against degradation in the rumen to be as efficacious in ruminants as in non‐ruminant species. The Panel considers the use of the additive in water for drinking to be equally effective than the use in feed when used as a nutritional additive. Since l‐isoleucine is used in food as a flavouring compound, and its function in feed is essentially the same as that in food no further demonstration of efficacy is necessary.

Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on l‐isoleucine produced by Escherichia coli FERM ABP‐10641 (i) to renew the authorisation and to extend the use of the additive in water for drinking when used as a nutritional additive for all animal species and (ii) to evaluate a new use as a flavouring additive for all animal species. The applicant has provided evidence that the additive currently in the market complies with the existing conditions of authorisation. l‐Isoleucine does not give rise to any safety concern regarding the production strain and its genetic modification. Considering that the production process has not been substantially modified and that no adverse effects have been reported in the literature search, the FEEDAP Panel considers that there is no evidence to reconsider the conclusions reached in previous assessments. Therefore, the Panel considers that the additive l‐isoleucine produced by E. coli FERM ABP‐10641 remains safe for the target species, consumer and for the environment when used as a nutritional additive in feed. The Panel extends these conclusions to the use of the additive as a flavouring compound. Regarding the use in water, the FEEDAP Panel reiterates its concerns over the safety for the target species of l‐isoleucine administered simultaneously via water for drinking and feed owing to the risk of nutritional imbalances and hygienic reasons. l‐Isoleucine is not irritant to skin and eyes and is not a dermal sensitiser but is hazardous by inhalation. The Panel confirms the previous conclusions that the additive is considered as a source of available isoleucine for non‐ruminant animal species when used as a nutritional additive. It requires protection against degradation in the rumen to be as efficacious in ruminants as in non‐ruminant species. The Panel considers the use of the additive in water for drinking to be equally effective than the use in feed when used as a nutritional additive. Since l‐isoleucine is used in food as a flavouring compound, and its function in feed is essentially the same as that in food no further demonstration of efficacy is necessary. [ABSTRACT FROM AUTHOR]

Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety and efficacy of L-isoleucine produced by fermentation using Corynebacterium glutamicum KCCM 80189 when used as a nutritional additive in feed and water for drinking for all animal species. The production strain is not genetically modified. Viable cells of the production strain were not detected in the final additive. The additive does not give rise to any safety concern regarding the production strain. L-Isoleucine produced using C. glutamicum KCCM 80189 is considered safe for the target species, for the consumer and for the environment. L-Isoleucine produced by C. glutamicum KCCM 80189 is considered not toxic by inhalation, not irritant to skin or eyes and not a dermal sensitiser. Regarding the use in water, the FEEDAP Panel reiterates its concerns over the safety of L-isoleucine administered simultaneously via water for drinking and feed owing to the risk of nutritional imbalances and hygienic reasons. L-Isoleucine produced by C. glutamicum KCCM 80189 is considered as an efficacious source of the essential amino acid L-isoleucine for non-ruminant animal species. For the supplemental L-isoleucine to be as efficacious in ruminants as in non-ruminant species, it would require protection against degradation in the rumen. [ABSTRACT FROM AUTHOR]

In this work, the molecular structure, harmonic vibrational frequencies of L- isoleucine (LI), L-isoleucine Maleate (LIM) and L-isoleucine Oxalate (LIO) have been investigated. The optimized molecular geometries, first order hyperpolarizabilities and vibrational frequencies have been calculated using B3LYP/6-31G. The HOMO and LUMO energy calculation reveals that the charge transfer exist within the molecules. The Fourier Transform Infrared (FT-IR) of LIM and LIO are recorded in the regions 4000-400 cm-1. The observed and calculated wave numbers are found to be in good agreement. The experimental spectra also coincide satisfactorily with those of theoretically constructed spectra. [ABSTRACT FROM AUTHOR]

Trehalose is commonly used as a protein stabilizer in spray dried protein formulations delivered via the pulmonary route. Spray dried trehalose formulations are highly hygroscopic, which makes them prone to deliquescence and recrystallization when exposed to moisture, leading to impairment in aerosolization performance. The main aim of this study was to investigate and compare the effect of hydrophobic amino acids (i.e. L-leucine and L-isoleucine) in enhancing aerosolization performance and in mitigating moisture-induced changes in spray dried trehalose formulations. Trehalose was spray dried with 20–60% w/w of amino acid (i.e. L-leucine or L-isoleucine). The spray dried formulations were stored at 25 °C/50% RH for 28 days. Solid state characterization and in vitro aerosolization performance studies were performed on the spray dried formulations before and after storage. The addition of 20–60% w/w of amino acid (i.e. L-leucine or L-isoleucine) improved the emitted fractions of spray dried trehalose formulations from a dry powder inhaler. However, ≥ 40% w/w of L-leucine/L-isoleucine was needed to prevent recrystallization of trehalose in the formulations when exposed to 25 °C/50% RH for 28 days. X-ray photoelectron spectroscopy (XPS) demonstrated that samples with 40–60% w/w L-isoleucine had more amino acid on the surfaces of the particles compared to their L-leucine counterparts. This may explain the greater ability of the L-isoleucine (40–60% w/w) samples to cope with elevated humidity compared to L-leucine samples of the same concentrations, as observed in the dynamic vapour sorption (DVS) studies. In conclusion, this study demonstrated that both L-leucine and L-isoleucine were effective in enhancing aerosolization performance and mitigating moisture-induced reduction in aerosolization performance in spray dried trehalose formulations. L-isoleucine proved to be superior to L-leucine in terms of its moisture protectant effect when incorporated at the same concentration in the formulations. [ABSTRACT FROM AUTHOR]

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Abstract This study experimentally investigated the electrostatic characteristic of L-isoleucine powder, which is an amino acid powder. L-isoleucine is a crystal powder that has been used in industrial production sites. The volume resistivity, charge-to-mass ratio, and minimum ignition energy (MIE) or L-isoleucine powder were tested in this experiment. The results show that L-isoleucine powder had a volume resistivity as high as 1.2 × 1013 Ωm, classified as highly electrifying, with a high risk of electrostatic charge. The actual charge-to-mass ratio was also as high as 6.4 nC/g on average. Added to this, with a MIE of 4 mJ, it is a material very sensitive to electrostatic discharge. Graphical abstract Unlabelled Image Highlights • Volume resistivity of L-isoleucine powder was classified as highly electrifying. • The actual charge-to-mass ratio of L-isoleucine powder was as high as 6.4 nC/g. • L-isoleucine powder can be ignited with 4 mJ of electrostatic discharges. [ABSTRACT FROM AUTHOR]

As one of the three branched-chain amino acids essential for human body, L-isoleucine is widely used in food, medicine, and feed industries. At present, L-isoleucine is mainly produced by microbial fermentation, and the main production strain is Corynebacterium glutamicum. The biosynthetic pathway of L-isoleucine in C. glutamicum is complex, and the activity of key enzymes and the transcription of key genes in the pathway are strictly regulated. The intracellularly synthesized L-isoleucine is secreted by transporters, and the activity of the transporters is also regulated. These intricate regulatory mechanisms increase the difficulty to engineer the L-isoleucine-producing C. glutamicum. This article focuses on the mechanism of L-isoleucine biosynthesis, secretion, and regulation in C. glutamicum and reviews the various metabolic engineering strategies for improving L-isoleucine production efficiency in C. glutamicum. [ABSTRACT FROM AUTHOR]

Perturbations in the metabolism of ammonia, a cytotoxic endogenous metabolite, occur in a number of chronic diseases, with consequent hyperammonemia. Increased skeletal muscle ammonia uptake causes metabolic, molecular, and phenotype alterations including cataplerosis of (loss of tricarboxylic acid cycle (TCA) cycle intermediate) α-ketoglutarate (αKG), mitochondrial oxidative dysfunction, and senescence-associated molecular phenotype (SAMP). L-Isoleucine (Ile) is an essential, branched-chain amino acid (BCAA) that simultaneously provides acetyl-CoA as an oxidative substrate and succinyl-CoA for anaplerosis (providing TCA cycle intermediates). Our multiomics analyses in myotubes and skeletal muscle from hyperammonemic mice and human patients with cirrhosis showed perturbations in BCAA transporters and catabolism. We, therefore, determined if Ile reverses hyperammonemia-induced impaired mitochondrial oxidative function and SAMP. Studies were performed in differentiated murine C2C12 myotubes that were early passage, late passage (senescent), or those depleted of LAT1/SLC7A5 and human induced pluripotent stem cell-derived myotubes (hiPSCM). Ile reverses hyperammonemia-induced reduction in the maximum respiratory capacity, complex I, II, and III functions in early passage murine myotubes and hiPSCM. Consistently, low ATP content and impaired global protein synthesis (high energy requiring cellular process) during hyperammonemia are reversed by Ile in murine myotubes and hiPSCM. Lower abundance of critical regulators of protein synthesis in mTORC1 signaling, and increased phosphorylation of eukaryotic initiation factor 2α are also reversed by Ile. Genetic depletion studies showed that Ile responses are independent of the amino acid transporter LAT1/SLC7A5. Our studies show that Ile reverses the hyperammonemia-induced impaired mitochondrial oxidative function, cataplerosis, and SAMP in a LAT1/SLC7A5 transporter-independent manner. [Display omitted] [ABSTRACT FROM AUTHOR]

Cysteine synthase A (CysK) catalyzes the last reaction of l‐cysteine synthesis in bacteria, but its moonlighting functions have been revealed recently. In this study, CysK was overexpressed in Corynebacterium glutamicum IWJ001, an l‐isoleucine producer. Compared with the control IWJ001/pDXW‐8, IWJ001/pDXW‐8‐cysK cells grew fast during log phase, and produced 26.5% more l‐isoleucine in flask fermentation and 23.5% more l‐isoleucine in fed‐batch fermentation. The key genes aspC, lysC, hom, thrB, ilvA, and ilvBN involved in l‐isoleucine biosynthesis were all upregulated in IWJ001/pDXW‐8‐cysK, compared with IWJ001/pDXW‐8. In addition, IWJ001/pDXW‐8‐cysK cells were longer and thicker than IWJ001/pDXW‐8 cells. Compared with IWJ001/pDXW‐8, the membrane permeability increased 15.8% and biofilm formation ability decreased 71.3% for IWJ001/pDXW‐8‐cysK cells. The results demonstrate that CysK overexpression in C. glutamicum is a good approach to enhance l‐isoleucine production. [ABSTRACT FROM AUTHOR]

Gout and hyperuricemia are highly prevalent metabolic diseases caused by high level of uric acid. Amino acids (AAs) involve in various biochemical processes including the biosynthesis of uric acid. However, the role of AAs in discriminating gout from hyperuricemia remains unknown. Here, we report that the plasma AAs profile can distinguish acute gout (AG) from asymptomatic hyperuricemia (AHU). We established an LC-MS/MS-based method to measure the plasma AAs without derivatization for the AG and AHU patients, and healthy controls. We found that the plasma profiling of AAs separated the AG patients from AHU patients and controls visually in both principal component analysis and orthogonal partial least-squares discriminant analysis (OPLS-DA) models. In addition, L-isoleucine, L-lysine, and L-alanine were suggested as the key mediators to distinguish the AG patients from AHU and control groups based on the S-plot analysis and variable importance in the projection values in the OPLS-DA models, volcano plot, and the receiver operating characteristic curves. In addition, the saturation of monosodium urate in the AA solutions at physiologically mimic status supported the changes in plasma AAs facilitating the precipitation of monosodium urate. This study suggests that L-isoleucine, L-lysine, and L-alanine could be the potential markers to distinguish the AG from AHU when the patients have similar blood levels of uric acid, providing new strategies for the prevention, treatment, and management of acute gout. [ABSTRACT FROM AUTHOR]

[Display omitted] • Investigated solubilities of L-isoleucine and L-serine in aqueous – (NH 4) 2 SO 4 electrolyte solutions. • L-serine exhibited higher solubility than L-isoleucine in water and water-electrolyte mixtures. • L-serine showed enhanced solubility (salting-in effect) with increasing electrolyte concentration. • L-isoleucine had reduced solubility (salting-out effect) at a specific temperature. • The larger size and hydrophobic side chain of L-isoleucine resulted in weaker interactions with electrolyte ions. The study aimed to measure the solubilities of L-serine and L-isoleucine in an aqueous ammonium sulfate [(NH 4) 2 SO 4 ] solution at different temperatures (288.15 to 308.15 K) using a gravimetric technique. In pure aqueous media L-serine solubility is 4.1218 in mol•kg−1 of water while for L-isoleucine is 0.3094 mol•kg−1 at 298.15 K. The obtained solubility data was used to estimate the thermodynamic properties of the amino acids, such as the standard Gibbs free energy and standard entropy change during the solvation process. The analysis involved investigating the short range chemical interactions between the amino acids and the electrolyte solution, which significantly influenced the solvation thermodynamics. The study found that L-serine exhibited a salting in effect, meaning its solubility increased in the presence of ammonium sulfate, while L-isoleucine experienced a salting out effect, resulting in decreased solubility with the electrolyte. Additionally, solubility parameters were employed to assess the chemical strength of the amino acids and understand their solute–solvent interactions and relative solubilities in the solution. Overall, this experimental investigation provided valuable insights into the solvation behavior and chemical characteristics of L-serine and L-isoleucine in the presence of an electrolyte solution. [ABSTRACT FROM AUTHOR]

A ligand-exchange micellar electrokinetic capillary electrophoresis system with copper(II)-L-isoleucine complexes as the chiral selector incorporated in micelles of sodium dodecyl sulfate (SDS) was developed for the enantioseparation of ofloxacin and its four related substances (impurities A, C, E, and F). The effects of important parameters affecting separation such as buffer pH, SDS concentration, chiral selector concentration, and organic additive were investigated in detail. Under optimum experimental conditions, enantioseparation of ofloxacin, impurities A, C, E, and F enantiomers was accomplished with resolutions of 4.28, 2.83, 3.40, 3.58, and 2.46, respectively. Further, simultaneous separation of impurities A, C, E, and F enantiomers was achieved using 10 mmol/L NH4OAc as the running buffer containing 4 mmol/L copper sulfate,20 mmol/L L-isoleucine, 20 mmol/L SDS, and 5% methanol at pH 8.5. To the best of our knowledge, the simultaneous enantioseparation of four impurities of ofloxacin has not been reported previously. [ABSTRACT FROM AUTHOR]

• Solubility of L-isoleucine in three kinds of binary solvent mixtures were determined using gravimetric method. • The experimental solubility data of L-isoleucine were correlated and analyzed by three thermodynamic models. • Dissolution thermodynamic properties of L-isoleucine were calculated and discussed. Thermodynamic properties, which play an important role in design and optimization of crystallization processes, are the essential data for the study of crystal nucleation and growth. In this study, the solubility of L-isoleucine were firstly determined by gravimetric method in three binary solvent mixtures including water + (ethanol, acetonitrile or DMSO) in the temperature range of 303.20 K to 338.05 K at atmospheric pressure. The effects of temperatures and solvent types on the dissolution process of L-isoleucine were investigated. It was found that the solubility of L-isoleucine increased dramatically with the increasing of the mole fraction of water in each binary solvent mixture and increased monotonically with the increasing of temperature. To extend the applicability of the solubility data, the experimental solubility data of L-isoleucine were fitted and analyzed using the modified Apelblat equation, the CNIBS/R-K model and the Jouyban-Acree model. Finally, based on the Van't Hoff equation and the experimental solubility data, the apparent dissolution thermodynamic properties of L-isoleucine including apparent dissolution enthalpy, apparent dissolution entropy, and apparent Gibbs free energy change of dissolution process were calculated and discussed. [ABSTRACT FROM AUTHOR]