Growth-coupled production of L-isoleucine in Escherichia coli via metabolic engineering.

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.

Discovery of N-succinyl-L-isoleucine as a novel taste enhancer via multisensory techniques and molecular simulations.

As awareness of health implications associated with dietary habits elevates alongside an increase in living standards, the trend toward minimizing sodium chloride and monosodium glutamate consumption has gained prominence. This study explored a novel taste enhancer, N-succinyl-L-isoleucine (N-Suc-Ile), focusing on its synthesis, taste-enhancing effects, and taste-enhancing mechanism. The synthesis of N-Suc-Ile was achieved through enzymatic catalysis employing food-grade enzymes in an aqueous environment, achieving yielding 30.22 %. Quantitative descriptive analysis revealed that the addition of N-Suc-Ile significantly enhanced the intensities of umami (13.11-37.70 %), saltiness (11.11-38.10 %), and kokumi (12.73-49.10 %) in a concentration-dependent manner (0.25-1 mg/L). Furthermore, time-intensity results showed that the durations of both umami and saltiness were prolonged by 46.88 % and 50 %, respectively, with the addition of 1 mg/L N-Suc-Ile. Analysis using sigmoid curves further validated N-Suc-Ile's synergistic effects in enhancing umami and saltiness. Molecular docking and dynamic simulation indicated that N-Suc-Ile bound effectively and stably to taste receptors, enhancing the sensations of umami, saltiness, and kokumi. These findings underscored the potential of N-Suc-Ile as a taste enhancer, suggesting its application could ensure the taste of food products while aligning with health-conscious consumer preferences.

Study on solubility and solvation thermodynamics for the advancement of biorelevant activities of l-isoleucine and l-serine in aqueous ammonium chloride solutions in the temperature range of 288.15-308.15 K.

This study investigated the dissolution behavior of l-isoleucine and l-serine in an aqueous salt solution (ammonium chloride), examining how variations in temperature and electrolyte concentration affect their solubility. We conducted careful experiments and used mathematical calculations to explore interactions at a molecular level. We observed that the structure of these amino acids and salt concentration in the aqueous medium influence their interactions, which affects dissolution. In the presence of electrolytes, l-isoleucine demonstrated a salting-out effect whereas l-serine showed a salting-in effect. This work examines the solute-solvent interactions of these solutes in aqueous ammonium chloride solutions. l-isoleucine exhibits a nonspontaneous reaction with increasing salt concentrations whereas l-serine shows spontaneous behavior. Gibbs free energy analysis revealed greater stability of l-serine. The pH and conductance measurements showed how these factors influence solution properties. This insight helps us comprehend the nature and behavior of these molecules in different situations, which could be helpful in drug formulation or protein purification in the future.

Investigation of the synthesis, gelation potential, and drug-loading capacities of two novel amides.

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 T g was found, and then, ΔH g 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 anti-inflammatory and analgesic effects-and their drug-loading capacities were thus determined.

Safety and efficacy of a feed additive consisting of l-isoleucine produced with Corynebacterium glutamicum CGMCC 20437 for all animal species (Eppen Europe SAS).

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.

One-Pot Synthesis of Useful S-Substituted-l-cysteine Sulfoxides Using Genetically Engineered Escherichia coli.

S-Substituted-l-cysteine sulfoxides are valuable compounds that are contained in plants. Particularly, (+)-alliin and its degraded products have gained significant attention because of their human health benefits. However, (+)-alliin production has been limited to extraction from plants and chemical synthesis; both methods have drawbacks in terms of stability and safety. Here, we proposed the enzymatic cascade reaction for synthesizing (+)-alliin from readily available substrates. To achieve a one-pot (+)-alliin production, we constructed Escherichia coli coexpressing the genes encoding tryptophan synthase from Aeromonas hydrophila ssp. hydrophila NBRC 3820 and l-isoleucine hydroxylase from Bacillus thuringiensis 2e2 for the biocatalyst. Deletion of tryptophanase gene in E. coli increased the yield about 2-fold. Under optimized conditions, (+)-alliin accumulation reached 110 mM, which is the highest productivity thus far. Moreover, natural and unnatural S-substituted-l-cysteine sulfoxides were synthesized by applying various thiols to the cascade reaction. These results indicate that the developed bioprocess would enable the supply of diverse S-substituted-l-cysteine sulfoxides.

Identification and characterization of GLYCEROLIPASE A1 for wound-triggered JA biosynthesis in Nicotiana benthamiana leaves.

Although many important discoveries have been made regarding the jasmonate signaling pathway, how jasmonate biosynthesis is initiated is still a major unanswered question in the field. Previous evidences suggest that jasmonate biosynthesis is limited by the availability of fatty acid precursor, such as ⍺-linolenic acid (⍺-LA). This indicates that the lipase responsible for releasing α-LA in the chloroplast, where early steps of jasmonate biosynthesis take place, is the key initial step in the jasmonate biosynthetic pathway. Nicotiana benthamiana glycerol lipase A1 (NbGLA1) is homologous to N. attenuata GLA1 (NaGLA1) which has been reported to be a major lipase in leaves for jasmonate biosynthesis. NbGLA1 was studied for its potential usefulness in a species that is more common in laboratories. Virus-induced gene silencing of both NbGLA1 and NbGLA2, another homolog, resulted in more than 80% reduction in jasmonic acid (JA) biosynthesis in wounded leaves. Overexpression of NbGLA1 utilizing an inducible vector system failed to increase JA, indicating that transcriptional induction of NbGLA1 is insufficient to trigger JA biosynthesis. However, co-treatment with wounding in addition to NbGLA1 induction increased JA accumulation several fold higher than the gene expression or wounding alone, indicating an enhancement of the enzyme activity by wounding. Domain-deletion of a 126-bp C-terminal region hypothesized to have regulatory roles increased NbGLA1-induced JA level. Together, the data show NbGLA1 to be a major lipase for wound-induced JA biosynthesis in N. benthamiana leaves and demonstrate the use of inducible promoter-driven construct of NbGLA1 in conjunction with its transient expression in N. benthamiana as a useful system to study its protein function.

Circabidian rhythm of sex pheromone reception in a scarab beetle.

Animals have endogenous clocks that regulate their behavior and physiology. These clocks rely on environmental cues (time givers) that appear approximately every 24 h due to the Earth's rotation; thus, most insects exhibit a circadian rhythm. One notable exception is the scarab beetle, Holotrichia parallela, a severe agricultural pest in China, Japan, South Korea, and India. Females emerge from the soil every other night, reach the canopy of host plants, evert an abdominal gland, and release a pheromone bouquet comprising l-isoleucine methyl ester (LIME) and l-linalool. To determine whether this circa'bi'dian rhythm affects the olfactory system, we aimed to identify H. parallela sex pheromone receptor(s) and study their expression patterns. We cloned 14 odorant receptors (ORs) and attempted de-orphanizing them in the Xenopus oocyte recording system. HparOR14 gave robust responses to LIME and smaller responses to l-linalool. Structural modeling, tissue expression profile, and RNAi treatment followed by physiological and behavioral studies support that HparOR14 is a sex pheromone receptor-the first of its kind discovered in Coleoptera. Examination of the HparOR14 transcript levels throughout the adult's life showed that on sexually active days, gene expression was significantly higher in the scotophase than in the photophase. Additionally, the HparOR14 expression profile showed a circabidian rhythm synchronized with the previously identified pattern of sex pheromone emission. 48 h of electroantennogram recordings showed that responses to LIME were abolished on non-calling nights. In contrast, responses to the green leaf volatile (Z)-3-henexyl acetate remained almost constant throughout the recording period.

L-Isoleucine reverses hyperammonemia-induced myotube mitochondrial dysfunction and post-mitotic senescence.

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.

Heat Capacities of N-Acetyl Amides of Glycine, L-Alanine, L-Valine, L-Isoleucine, and L-Leucine.

As a follow-up to our effort to establish reliable thermodynamic data for amino acids, the heat capacity and phase behavior are reported for N-acetyl glycine amide (CAS RN: 2620-63-5), N-acetyl-L-alanine amide (CAS RN: 15962-47-7), N-acetyl-L-valine amide (CAS RN: 37933-88-3), N-acetyl-L-isoleucine amide (CAS RN: 56711-06-9), and N-acetyl-L-leucine amide (CAS RN: 28529-34-2). Prior to heat capacity measurement, thermogravimetric analysis and X-ray powder diffraction were performed to determine decomposition temperatures and initial crystal structures, respectively. The crystal heat capacities of the five N-acetyl amino acid amides were measured by Tian-Calvet calorimetry in the temperature interval (266-350 K), by power compensation DSC in the temperature interval (216-471 K), and by relaxation (heat-pulse) calorimetry in the temperature interval (2-268 K). As a result, reference heat capacities and thermodynamic functions for the crystalline phase from 0 K up to 470 K were developed.

Exploring L-isoleucine riboswitches for enhancing 4-hydroxyisoleucine production in Corynebacterium glutamicum.

OBJECTIVES: To explore an L-isoleucine (Ile)-induced biosensor for down-regulation of Ile synthesis pathway and enhancement of 4-hydroxyisoleucine (4-HIL) production in Corynebacterium glutamicum SN01. RESULTS: Four Ile-induced riboswitches (IleRSN) with different strength were screened from mutation library based on TPP riboswitch. Firstly, IleRSN were integrated into the chromosome of strain SN01 immediately upstream of ilvA gene. The 4-HIL titer of strains carrying PtacM-driven IleRS1 or IleRS3 (14.09 ± 1.07, 15.20 ± 0.93 g 4-HIL L-1) were similar with control strain S-D5I (15.73 ± 2.66 g 4-HIL L-1). Then, another copy of IleRS3-ilvA was integrated downstream of the chromosomal cg0963 gene in SN01-derived strain D-RS with down-regulated L-lysine (Lys) biosynthesis. The Ile supply and 4-HIL titer increased in ilvA two-copy strains KIRSA-3-D5I and KIRSA-3-9I, and Ile concentration was maintained less than 35 mmol L-1 under the control of IleRS3 during fermentation. The resulting strain KIRSA-3-9I produced 22.46 ± 0.96 g 4-HIL L-1. CONCLUSION: The screened IleRS was effective in the dynamic down-regulation of Ile synthesis pathway in C. glutamicum, and IleRSN with different strength can be applied in various conditions.

Semi-Rational Design of L-Isoleucine Dioxygenase Generated Its Activity for Aromatic Amino Acid Hydroxylation.

Fe (II)-and 2-ketoglutarate-dependent dioxygenases (Fe (II)/α-KG DOs) have been applied to catalyze hydroxylation of amino acids. However, the Fe (II)/α-KG DOs that have been developed and characterized are not sufficient. L-isoleucine dioxygenase (IDO) is an Fe (II)/α-KG DO that specifically catalyzes the formation of 4-hydroxyisoleucine (4-HIL) from L-isoleucine (L-Ile) and exhibits a substrate specificity toward L-aliphatic amino acids. To expand the substrate spectrum of IDO toward aromatic amino acids, in this study, we analyzed the regularity of the substrate spectrum of IDO using molecular dynamics (MD) simulation and found that the distance between Fe2+, C2 of α-KG and amino acid chain's C4 may be critical for regulating the substrate specificity of the enzyme. The mutation sites (Y143, S153 and R227) were also subjected to single point saturation mutations based on polarity pockets and residue free energy contributions. It was found that Y143D, Y143I and S153A mutants exhibited catalytic L-phenylalanine activity, while Y143I, S153A, S153Q and S153Y exhibited catalytic L-homophenylalanine activity. Consequently, this study extended the substrate spectrum of IDO with aromatic amino acids and enhanced its application property.

Scientific opinion on the presence of DNA in the feed additive consisting of l-isoleucine produced by Corynebacterium glutamicum KCCM 80185 for all animal species (CJ Europe GmbH).

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.

Improve L-isoleucine production in Corynebacterium glutamicum WM001 by destructing the biosynthesis of trehalose dicorynomycolate.

Trehalose dicorynomycolates are structurally important constituents of the cell envelope in Corynebacterium glutamicum. The genes treS, treY, otsA, mytA and mytB are necessary for the biosynthesis of trehalose dicorynomycolates. In this study, the effect of biosynthesis of trehalose dicorynomycolates on L-isoleucine production in C. glutamicum has been investigated by deleting the genes treS, treY, otsA, mytA, and mytB in the L-isoleucine producing C. glutamicum WM001. L-isoleucine production was slightly improved in the mutants ΔtreY, ΔotsA, and ΔtreYA, and not improved in the single deletion mutant ΔtreS , but significantly improved in the triple deletion mutant ΔtreSYA. Deletion of mytA or mytB in ΔtreSYA could further improve L-isoleucine production. However, deletion of both mytA and mytB in ΔtreSYA significantly decreased L-isoleucine production. The final L-isoleucine producing C. glutamicum WL001 was constructed by deletion of treS, treY, otsA, and mytB, insertion of lrp, and replacement of the native promoter of ilvA with the L-isoleucine sensitive promoter PbrnFE7. WL001 grew worse than the control WM001, but produced 36.1% more L-isoleucine after 72 h shake flask cultivation than WM001.

Fecal microbiota from MRL/lpr mice exacerbates pristane-induced lupus.

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.

Internal standard metabolites for estimating origin blood volume of bloodstains.

The volume of blood leaked from blood vessels may change due to evaporation of water under the natural influence of the external environment. Bloodstains and dried blood spots (DBS), which describes blood dried in the external environment, are similar in their production and their metabolite quantification profiles. In both bloodstain metabolite analysis in the forensic science field and DBS metabolite analysis in the clinical field, it is important to determine the volume of the origin blood as this affects metabolite quantification results. Therefore, the purpose of this study is to discover the internal standard metabolites that have quantitatively proportional relationships with origin blood volume and maintain constant concentrations even as the age of the bloodstain increases. As a result, the concentrations of L-isoleucine and L-phenylalanine increased in proportion to the origin blood volume of the bloodstain. The differences in concentration of L-isoleucine were significant in all volume comparisons except in the comparison between 65 µL and 85 µL. The differences in concentration of L-phenylalanine were significant in all volume comparisons except between 65 µL and 45 µL and between 65 µL and 85 µL. In addition, it was confirmed that both metabolites tended to maintain constant concentrations without being affected by bloodstain age as the volume became smaller. These internal standard metabolites can be used for estimating the origin blood volume of bloodstains during metabolite analysis of bloodstains and DBS and could provide a volume criterion for standardization when comparing metabolite quantification between samples.

Development, validation, and uncertainty measurement of HPLC-DAD method for determination of some free amino acids in infant formula and medical food products for inborn errors of metabolism.

This research aims at determining some free amino acids in amino acid-based infant formulas and amino acid-modified medical foods for inborn errors of metabolism to prove their quality. A method based on high-performance liquid chromatography and diode array detection was developed and validated. Then, overall uncertainty was estimated by the bottom-up approach. Applying the weighted least squares regression method suggested good linearity with coefficient of determinations ≥ 0.9960. The limits of detection were calculated between 0.01 and 0.28 µg/mL. The most repetitive recovery values were obtained in the range of 91-108%, with RSDs ≤ 15%. The expanded uncertainties were below 20% for most amino acids. The contributions of linear regression and repeatability are two main factors in estimating overall uncertainty. The results offer this method as a simple and easy procedure for analyzing free amino acids in seven powdered medical foods designed for phenylketonuria, maple syrup urine disease, methylmalonic, and propionic acidemia.

The Escherichia coli Amino Acid Uptake Protein CycA: Regulation of Its Synthesis and Practical Application in l-Isoleucine Production.

Amino acid transport systems perform important physiological functions; their role should certainly be considered in microbial production of amino acids. Typically, in the context of metabolic engineering, efforts are focused on the search for and application of specific amino acid efflux pumps. However, in addition, importers can also be used to improve the industrial process as a whole. In this study, the protein CycA, which is known for uptake of nonpolar amino acids, was characterized from the viewpoint of regulating its expression and range of substrates. We prepared a cycA-overexpressing strain and found that it exhibited high sensitivity to branched-chain amino acids and their structural analogues, with relatively increased consumption of these amino acids, suggesting that they are imported by CycA. The expression of cycA was found to be dependent on the extracellular concentrations of substrate amino acids. The role of some transcription factors in cycA expression, including of Lrp and Crp, was studied using a reporter gene construct. Evidence for the direct binding of Crp to the cycA regulatory region was obtained using a gel-retardation assay. The enhanced import of named amino acids due to cycA overexpression in the l-isoleucine-producing strain resulted in a significant reduction in the generation of undesirable impurities. This work demonstrates the importance of uptake systems with respect to their application in metabolic engineering.

Safety and efficacy of a feed additive consisting of l-isoleucine produced by Corynebacterium glutamicum KCCM 80185 for all animal species (CJ Europe GmbH).

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.

Potato tuber-inducing activities of jasmonic acid and related-compounds (II).

New information is being accumulated for plant-derived oxylipins, such as jasmonic acid (JA) amino acid conjugates. However, these compounds have not being examined for their activity in promoting potato tuber formation. It was found that (-)-JA had the highest activity followed cis-(-)-OPDA, (+)-4, 5-didehydroJA, cis-(+)-OPDA-l-Ile, and (-)-JA-l-Ile, -Leu, -Phe, -Val, although iso-OPDA and 3,7-didehydroJA did not exhibit activity.