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Extraction experiments in the two-phase water-nitrobenzene system and γ-activity measurements were used to determine the stability constant of the enniatin B–K+ complex dissolved in nitrobenzene saturated with water as 105.5±0.2. Quantum mechanical calculations were applied to predict the most probable structure of this cationic complex species.

Forages are important components of dairy cattle rations but might harbor a plethora of mycotoxins. Ruminants are considered to be less susceptible to the adverse health effects of mycotoxins, mainly because the ruminal microflora degrades certain mycotoxins. Yet, impairment of the ruminal degradation capacity or high ruminal stability of toxins can entail that the intestinal epithelium is exposed to significant mycotoxin amounts. The aims of our study were to assess i) the mycotoxin occurrence in maize silage and ii) the cytotoxicity of relevant mycotoxins on bovine intestinal cells. In total, 158 maize silage samples were collected from European dairy cattle farms. LC-MS/MS-based analysis of 61 mycotoxins revealed the presence of emerging mycotoxins (e.g. emodin, culmorin, enniatin B1, enniatin B, and beauvericin) in more than 70% of samples. Among the regulated mycotoxins, deoxynivalenol and zearalenone were most frequently detected (67.7%). Overall, 87% of maize silages contained more than five mycotoxins. Using an in vitro model with calf small intestinal epithelial cells B, the cytotoxicity of deoxynivalenol, nivalenol, fumonisin B1 and enniatin B was evaluated (0&ndash, 200 &micro, M). Absolute IC50 values varied in dependence of employed assay and were 1.2&ndash, 3.6 &micro, M, 0.8&ndash, 1.0 &micro, M, 8.6&ndash, 18.3 &micro, M, and 4.0&ndash, 6.7 &micro, M for deoxynivalenol, nivalenol, fumonisin B1, and enniatin B, respectively. Results highlight the potential relevance of mycotoxins for bovine gut health, a previously neglected target in ruminants.

Totals of 158 corn and corn-based samples and 291 wheat and wheat-based samples from Shandong province, China in 2017 were analyzed for five mycotoxins including beauvericin (BEA), enniatin A (ENA), enniatin A1 (ENA1), enniatin B (ENB), and enniatin B1 (ENB1) by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). BEA was the predominant toxin detected, followed by ENB, ENA1, ENA, and ENB1. Corn and corn-based samples were more easily contaminated by BEA with an average concentration of 65.26 &micro, g/kg, compared with that in wheat and wheat-based samples (average = 0.41 &micro, g/kg). Concentrations of BEA, ENA, and ENB1 in corn kernels, flours, and flakes were significantly different (Kruskal&ndash, Wallis Test, p &lt, 0.05), as well as for BEA, ENA, ENB, and ENB1 in wheat kernels, flours, and noodles (Kruskal&ndash, Wallis test, p &lt, 0.05). Furthermore, 59.5% (94/158) and 59.8% (174/291) corn- and wheat-based samples were co-contaminated by at least two mycotoxins, respectively. Positive correlations in concentrations were observed in corn between levels of ENA and ENB1, ENA and ENB, ENA1 and ENB1, as well as in wheat between BEA and ENA, BEA and ENA1, BEA and ENB, BEA and ENB1, ENA and ENA1, ENA and ENB, ENA and ENB1, ENA1 and ENB, ENA1 and ENB1, and ENB and ENB1. These results demonstrate that co-contamination of BEA and enniatins (ENNs) in corn- and wheat-based samples from Shandong, China is very common. More data on the contamination of five mycotoxins in cereal and cereal-based samples nationwide are needed.

Presented at Indoor Air Seminar (Helsinki, Finland) 15 March, 2018. ISBN 978-952-5236-46-0. Mittasimme aineiden (hometoksiinit; rakennus- ja saneeraustuotteiden kemikaalit, siivousaineet) kulkeutumista päästölähteistä ilmaan tutkimuskammioissa. Tutkittavat aineet annosteltiin kammioiden pohjalla oleviin avoimiin astioihin. Kammioiden ilman kosteutta säädettiin lisäämällä pohjalle vettä ja ilmankostutin- ja kuivainlaitteilla. Lämpötilaa, kosteutta ja TVOC-pitoisuutta monitoroitiin jatkuvatoimisilla antureilla. Kammioilman vesihöyry ja vesiaerosolit tiivistettiin nestemäiseksi vedeksi kuivaimen säiliöön. Kun tiivistevesi analysoitiin, havaittiin, että mikrobitoksiinit (okratoksiini A, enniatiini B) ja siivous- ja saneeraustuotteiden sisältämät antimikrobiset (PHMG) aineet, neutraalit (polyglykoli-alkoksyylit) ja kationiset tensidit (DDDMAC) siirtyivät pinnoilta ilmaan ja sieltä ilmankuivaimen kondenssivesisäiliöön. We measured mobility of nonvolatile fungal toxins and chemical substances in products used in building construction or maintenance into indoor air. The measurement was conducted in enclosed chambers provided with sensors for continuous monitoring of air humidity, temperature, organic substance (TVOC). Airborn humidicity was cyclically up/down adjusted using humidifier / dehumidifier. Substances tested for emission, were placed in open dishes on chamber floor. Airborn humidity was condensed into liquid water, harvested in dehumidifer storage container, and analysed at the end of each experiment.We found that nonvolatile substances such as fungal toxins (ochratoxin A), non-ionic and cationic tensides (polyoxyethylene alkanol ethers, didecyldimethyl ammonium chloride) and antimicrobial biocidal substances (PHMG) , contained in cleaning and sanitation products in buildings, translocated were recovered in the the dehumidifer condensate, thus indicating airborn migration.

Toxicogenic fungal moulds have been previously identified in Irish farm silages and are known to be capable of producing mycotoxins. Mycotoxins are detrimental to animal health and performance and can be transferred through feed to animal produce. The objectives of this study were to identify and quantify the challenge posed to livestock from mycotoxins in Irish silages and determine whether conventional chemical characteristics could be used as indicators of mycotoxin occurrence. Over the 2-year period, 300 silages were sampled on 150 farms and these consisted of round-baled grass silages (n = 115) and pit silages of either grass (n = 175) or maize (n = 10). There was no significant difference in mycotoxin concentrations across silage types in either year, except for baled silage containing higher concentrations of enniatin B compared with pit silage in Year 1. Conventional chemical characteristics of silages were generally not reliable predictors for mycotoxin incidences; however, dry-matter digestibility, crude protein, fermentation products and ash did predict the incidence of enniatins and beauvericin. The incidence and concentration of the twenty measured mycotoxins were generally low and individual mycotoxin concentrations recorded were considerably lower than current EU directive or guidance thresholds. Non-regulated mycotoxins measured were similar to or lower than concentrations reported in the literature. Based on current knowledge and the concentrations of individual mycotoxins detected in this study, the challenge to Irish livestock and livestock products from animal consuming silage is generally low. However, the additive or synergistic effects of multiple mycotoxins in silage are unknown.

In an effort to enforce the knowledge on mycotoxin occurrence and co-occurrence in dates and dried fruits, 228 samples purchased from Tunisian and Spanish markets were subjected to multi-mycotoxin liquid chromatography with tandem mass spectrometry (LC-MS/MS) analysis of 16 mycotoxins. At least one mycotoxin was detected in 160 samples (70%). The frequency of contaminated samples was 83%, 80%, 64%, 59% and 26% for dates, dried vine fruits, figs, apricots and plums, respectively but none of the analyzed samples contained detectable levels of fumonisins (FB1, FB2, FB3) or T-2 toxin (T2). The incidence and levels of mycotoxins varied in samples. The most prevalent mycotoxin was enniatin B (EnnB) (54%), followed by enniatinA1 (EnnA1) (36%); aflatoxins (AFs) (23%) and ochratoxin A (OTA) (22%). Thirteen samples contained AFs in levels that exceed the maximum limits established in EU legislation. The simultaneous contamination with 2–6 mycotoxins was observed in 51% of the analyzed samples. Lastly, intakes of the detected mycotoxins were calculated for average adult consumers and compared with the tolerable daily intakes (TDI). Data obtained were used to estimate the potential exposure levels.

The Fusarium mycotoxin enniatin B (ENN B) is a so-called emerging mycotoxin frequently contaminating poultry feed. To investigate the impact of chronic ENN B exposure on animal health, broiler chickens were fed either a diet naturally contaminated with ENN B (2352 µg/kg) or a control diet (135 µg/kg) for 2, 7, 14, or 21 days. ENN B concentrations were determined in plasma and liver using a validated ultra-high performance liquid chromatography—tandem mass spectrometry UHPLC-MS/MS method. Liver was evaluated histologically, and the villus length and crypt depth of the duodenum, jejunum, and ileum were measured. Histopathology of the livers did not reveal major abnormalities. Feeding an ENN B-contaminated diet could possibly inhibit the proliferation of enterocytes in the duodenal crypts, but did not affect villus length, crypt depth, or villus length-crypt depth ratio of the jejunum and ileum. ENN B levels in plasma and liver were significantly higher in the ENN B-fed group and ranged between

Enniatins (Enns) are mycotoxins produced by Fusarium spp. and are widely distributed contaminants of cereals and derivate products. Among the different identified enniatins, Enn B is the most relevant analogue in cereals in Europe. Therefore, the aim of this study was to investigate for the first time the occurrence of Enn B and Enn B phase I metabolites in 300 human urine samples throughout an ultrahigh-performance liquid chromatography-high resolution mass spectrometry (UHPLC-Q-Orbitrap HRMS) methodology. Three different sample preparation procedures were evaluated and salting-out liquid-liquid extraction showed satisfactory validation results. Enn B was quantified in 83.7% of samples ranging from 0.006 to 0.391 ng/mL (average content: 0.016 ng/mL). In line with the in vitro observations with human liver microsomes, in the here analyzed samples the Enn B monooxygenated, N-demethylated and dioxygenated metabolites were tentatively found in 87.7%, 96.3% and 6.7% of samples. The data of this pilot biomonitoring survey indicate a frequent intake of enniatins in Italy, supporting further toxicological studies to provide better basis for understanding their potential effects in humans.

Beauvericin (BEA) and Enniatins (ENN) are mycotoxins produced by Fusarium fungi detected in food and feed; there are insufficient data to establish their reference values. To evaluate BEA and ENN oral toxicity, an integrated approach was applied. Among ENN, Enniatin B (ENNB) was selected as test substance. The approach is composed by: i) in vitro and acute in vivo genotoxicity tests; ii) a repeated-dose oral toxicity study focused on genotoxic, immune, endocrine, nervous endpoints and the reproductive/developmental toxicity screening. For BEA, all the genotoxicity endpoints yielded negative results excluding Comet assay in duodenum and kidney after repeated doses. BEA immunotoxicity was observed in female mice, concentrated in number and functional activity of effector T cells in the spleen. Based on the repeated-dose BEA study, the No Observed Adverse Effect Level (NOAEL) for female mice is 1 mg/kg b.w. per day (increased thyroid pycnotic nuclei and endometrial hyperplasia). In males, the NOAEL is 0.1 mg/kg b.w. per day (reduced colloid and altered T4 serum levels). Maternal NOAEL is 0.1 mg/kg b.w. per day (increased thymus weight), developmental NOAEL is 10 mg/kg b.w. per day. For ENNB, the results support a genotoxic effect in bone marrow and liver cells after acute treatment, but not after repeated exposure. Immunotoxic ENNB effects were observed in both genders, suggestive of a suppressive/inhibiting activity particularly evident in males. Based on the repeated-dose ENNB study, the NOAEL for females is 0.18 mg/kg b.w. per day (histomorphometrical effects on thymus, uterus and spleen). In male mice, the NOAEL is 1.8 mg/kg b.w. per day (enterocyte vacuolization in duodenum and increased Reactive Oxygen Species and reduced Glutathione brain levels). The maternal NOAEL is 1.8 mg/kg b.w. per day (decreased white pulp area and increased red/white pulp area ratio in spleen), developmental NOAEL is 18 mg/kg b.w. per day.

The waste management occupational environment is recognized by the simultaneous presence of several substances and biologic agents. Therefore, workers are exposed simultaneously to multiple contaminants. Occupational exposure to aflatoxin B1 in one Portuguese waste sorting plant was already reported. However, besides this mycotoxin, data regarding fungal contamination showed that exposure to other mycotoxins could be expected. A study was developed to analyze if exposure to other mycotoxins besides aflatoxin B1 was occurring in the workers from the waste sorting plant previously assessed and to discuss how these findings need to be considered in the risk assessment process. In addition to aflatoxin B1 detected previously by ELISA, two additional mycotoxins and one mycotoxin degradation product were detected and quantified by a multi-mycotoxin HPLC-MS/MS approach: Enniatin B and ochratoxin A as well as 2’R-ochratoxin A. Besides the confirmation of co-exposure to several mycotoxins, results probably indicate different exposure routes for the mycotoxins reported.

Emerging Fusarium and Alternaria mycotoxins gain more and more interest due to their frequent contamination of food and feed, although in vivo toxicity and toxicokinetic data are limited. Whereas the Fusarium mycotoxins beauvericin, moniliformin and enniatins particularly contaminate grain and grain-based products, Alternaria mycotoxins are also detected in fruits, vegetables and wines. Although contamination levels are usually low (mu g/kg range), higher contamination levels of enniatins and tenuazonic acid may occasionally occur. In vitro studies suggest genotoxic effects of enniatins A, A1 and B1, beauvericin, moniliformin, alternariol, alternariol monomethyl ether, altertoxins and stemphyltoxin-III. Furthermore, in vitro studies suggest immunomodulating effects of most emerging toxins and a reproductive health hazard of alternariol, beauvericin and enniatin B. More in vivo toxicity data on the individual and combined effects of these contaminants on reproductive and immune system in both humans and animals is needed to update the risk evaluation by the European Food Safety Authority. Taking into account new occurrence data for tenuazonic acid, the complete oral bioavailability, the low total body clearance in pigs and broiler chickens and the limited toxicity data, a health risk cannot be completely excluded. Besides, some less known Alternaria toxins, especially the genotoxic altertoxins and stemphyltoxin III, should be incorporated in risk evaluation as well.

Bipolaris oryzae is a phytopathogenic fungus causing a brown spot disease in rice, and produces substance that strongly perturbs motility and membrane integrities of boar spermatozoa. The substance was isolated from the liquid culture of the fungal strain using extraction and a multi-step semi-preparative HPLC procedures. Based on the results of mass spectrometric and 2D NMR techniques, the bioactive molecule was identified as ophiobolin A, a previously described sesterterpene-type compound. The purified ophiobolin A exhibited strong motility inhibition and viability reduction on boar spermatozoa. Furthermore, it damaged the sperm mitochondria significantly at sublethal concentration by the dissipation of transmembrane potential in the mitochondrial inner membrane, while the plasma membrane permeability barrier remained intact. The study demonstrated that the cytotoxicity of ophiobolin A toward somatic cell lines is higher by 1–2 orders of magnitude compared to other mitochondriotoxic mycotoxins, and towards sperm cells unique by replacing the progressive motility by shivering tail beating at low exposure concentration.

The contamination levels of beauvericin and four enniatins, A, A1, B and B1, in 207 samples of wheat flour and corn grits on the Japanese market were determined by an analytical method based on LC-MS/MS. The toxins were extracted from samples with acetonitrile–water (85:15, v/v) and then purified with C18 cartridges. The method was validated in a single laboratory using spiked samples at two levels; the recovery of the five toxins ranged from 91.1% to 113.8%. Enniatin B was frequently detected in imported wheat flour (81.8%) and domestic wheat flour (85.6%), and the highest concentration of enniatin B was present in a domestic wheat sample (633 μg kg−1). In corn grits, beauvericin was found in 34% of the samples, but enniatins were not detected at all. The maximum concentration of beauvericin in corn grits was 26.1 μg kg−1. Deoxynivalenol and nivalenol in the same samples were determined by a method using an immunoaffinity column. Co-contamination of deoxynivalenol and enniatins was observed in 61% of the imported wheat samples and in 58% of the domestic wheat samples. These results suggest the need for a risk assessment for cyclic depsipeptide mycotoxins in Japan and a study on the synergistic effect of deoxynivalenol and enniatins.

Introduction - Occupational exposures are characterized by being complex and associated to co-exposure to several contaminants by different exposure routes. Even if exposure occurs to only a chemical agent, it can have different exposure routes and can result in different health effects. The waste management setting is recognized by the presence of several chemical and biologic agents in the workplaces. Recently, it was reported occupational exposure to Aflatoxin B1 (AFB1) in one Portuguese waste management industry. However, data regarding to fungal burden showed that exposure to other mycotoxins should be expected. Aim of the study - The aim of the present work was to know if workers from this waste management industry were exposed to other mycotoxins besides AFB1.

Six new cyclohexadepsipeptides, beauvenniatins A–E ( 1 – 5 ), and beauvericin J ( 6 ), together with the known beauvericin ( 7 ) and enniatin B ( 8 ), were isolated from the fungus Acremonium sp. BCC 28424. The productions of minor derivatives 3 – 6 , possessing an N -methyl- l -tyrosine, were enhanced by feeding l -tyrosine in the liquid fermentation medium. Antimalarial, antitubercular, and cytotoxic activities of these cyclodepsipeptides were evaluated.

A LC-MS/MS method for the detection of beauvericin and the four enniatins A, A1, B, and B1 in maize and maize silage was developed. The method uses direct injection of maize extracts without any tedious and laborious cleanup procedures. The limit of quantification was determined at 13 ng g(-1) for beauvericin and at 17, 34, 24, and 26 ng g(-1) for enniatins A, A1, B, and B1, respectively. The method was used in surveys of the compounds in fresh maize samples collected at harvest in 2005 and 2006. All samples had the same distribution of the enniatins: B > B1 > A1 > A. Enniatin B was present in 90% of the samples in 2005 and in 100% in 2006 at levels up to 489 and 2598 ng g(-1), respectively. Beauvericin contamination was more frequently detected in 2006 than in 2005 (89 and 10%, respectively) and in higher amounts (988 and 71 ng g(-1), respectively). The occurrence of beauvericin and the four enniatins was examined in 3-month-old maize silage stacks from 20 different farms. As observed in fresh maize, enniatin B was the most abundant compound in ensiled maize and was found from 19 stacks at levels up to 218 ng g(-1). The stability of enniatin B in maize silage was assessed by analyzing samples from 10 of the silage stacks taken after 3, 7, and 11 months of ensiling. Enniatin B could be detected at all locations after 11 months and appeared to be stable during ensiling.

Reasons for mammalian cell toxicity observed in barley and spring wheat grains were sought. Streptomyces sp. isolates from wheat and barley produced heat-stable methanol-soluble substances which inhibited the motility of exposed porcine spermatozoa used as a toxicity indicator. Several barley isolates produced antimycin A (2 to 5 ng/mg wet wt of biomass), a macrolide antibiotic known to block oxygen utilization in mitochondria. The antimycinproducing isolates were members of the Streptomyces albidoflavus group. In in vitro assays with porcine kidney tubular epithelial cells, the specific toxicity of antimycin A towards mitochondria was higher than that of the mycotoxin enniatin B but lower than that of the mitochondriotoxins cereulide and paenilide, produced by food-related Bacillus cereus and Paenibacillus tundrae, respectively. The toxic wheat isolates, related to Streptomyces sedi, did not produce antimycin A and or any other known toxin. Our results suggest that the presence of toxin-producing streptomycetes in stored cereal grains may pose a thus far unrecognized threat for food and feed safety.

Enniatin B (ENN B), deoxinivalenol (DON) and alternariol (AOH) are secondary metabolites of filamentous fungi. These mycotoxins are contaminants of vegetables and cereals. They are cytotoxic and their effects are enhanced by their mixtures. The objectives of this study were to compare the cytotoxicity of ENN B, DON and AOH alone or in combination in human adenocarcinoma (Caco-2) cells and to evaluate the type of interactions of mycotoxin mixtures by the isobologram analysis. Cells were treated with concentrations ranging from 1.85 to 90μM (AOH) and from 0.312 to 10μM (for ENN B and DON), individually and in combination of two and three mycotoxins (from 1.85 to 30μM for AOH and from 0.312 to 5μM for ENN B and DON). The relation ratios between the mixtures DON+ENN B was 1:1; AOH+DON and ENN B+AOH was 1:6, and for the tertiary combination DON, ENN B and AOH 1:1:6. The IC50 value of ENN B and DON were 3.87 and 5.54μM, respectively. No IC50 values were obtained for the AOH at any time tested in Caco-2 cells. With the isobologram the type of interaction between mycotoxin was evaluated. Synergistic, antagonistic and addictive effect was observed for the combination studied depending on the concentration affected. Mycotoxins combinations reduce cellular viability in the following increasing order: (DON+ENN B)>(ENN B+AOH)>(DON+AOH)>(DON+AOH+ENN B).

A nine-step (longest linear) batch total synthesis of the cyclic hexadepsipeptide (−)-enniatin B is described. The synthesis minimizes precipitation during reaction conditions for adaptability to flow synthesis. The route was used to prepare >100 mg of the natural product.

The crystal structure of the complex of enniatin B, cyclo[-(L-MeVal-D-Hyi)3-](C33H57N3O9), with KNCS is determined by X-ray diffraction [CuKαradiation, R = 0.0594 for 7925 reflections with I > 2 σ(I)]. The crystals belong to the space group P3, a = 24.448(5) A, c = 23.578(5) A, V = 12277(9) A3, and Z = 6. The unit cell contains 12 symmetrically independent molecules of the antibiotic, which are located on crystallographic threefold axes. The K+ ions are located on the threefold axes and are coordinated by the carbonyl oxygen atoms of the hydroxy acid or amino acid residues of the enniatin molecules to form prisms, twisted prisms, and antiprisms. All the independent enniatin molecules retain the principal conformational features revealed earlier in the structures of enniatin B and its complexes.

We evaluated Fusarium contamination and the levels of hexadepsipeptide mycotoxins in 13 wheat samples affected by head blight in Finland. Fusarium avenaceum was the dominant species (91%) isolated from all samples, but isolates of F. culmorum (4%), F. tricinctum (3%), and F. poae (2%) also were recovered. Beauvericin (0.64 to 3.5 μg/g) was detected in all 13 samples. Enniatin B (trace to 4.8 μg/g) was detected in 12 samples, enniatin B 1 (trace to 1.9 μg/g) was detected in 8 samples, and enniatin A 1 (trace to 6.9 μg/g) was detected in 10 samples. Ten of 13 strains of F. avenaceum and 2 strains of F. poae and F. tricinctum produced beauvericin in culture on rice (trace to 70, 9.4, and 33 μg/g, respectively). All strains also produced enniatins (trace to 2,700 μg/g). This is the first report on the natural cooccurence of beauvericin and enniatins in wheat infected predominantly by F. avenaceum .

Twenty-nine Fusarium isolates, representing three new taxa originated by Nirenberg from F. sambucinum Fuckel sensu lato, namely: F. sambucinum Fuckel sensu stricto, F. venenotum Nirenb., and F. torulosum (Berk.Curt.) Nirenb., were tested for in vitro production of toxic secondary metabolites on autoclaved corn kernels. F. sambucinum sensu stricto was able to produce type A trichothecenes and enniatin B (EB). In particular, amongst the 14 isolates tested, 5 produced only diacetoxyscirpenol (DAS) (up to 700 micrograms/g); 1 produced only neosolaniol (NEOS) (250 micrograms/g); 2 produced T-2 toxin (T-2) + NEOS (up to 175 and 150 micrograms/g, respectively); 1 produced NEOS + DAS (300 and 100 micrograms/g, respectively); and 5 produced DAS + EB (up to 500 and 140 micrograms/g, respectively). All six isolates of F. venenotum were able to produce only DAS (up to 100 micrograms/g). F. torulosum produced no trichothecenes, but four out of nine tested isolates were able to produce EB (up to 140 micrograms/g). Zearalenones and type B trichothecenes were not found. The toxicity of the culture extracts towards Artemia salina L. was correlated in general with the occurrence of the above toxins, except for some F. torulosum strains. However, the lack of correlation between the amounts of toxins recovered and toxic activity observed in the Geotrichum candidum Link ex Pers. and A. salina assays suggested the presence of unknown toxic compounds.

The cyclic hexadepsipeptide framework of enniatin B was identified as a template matching the beta-turn tripeptide of tendamistat. The modified analog 1 was synthesized as a tendamistat mimic and compared to the acyclic derivative 2 and the tripeptide Ac-Try-Arg-Tyr-OMe. These compounds were assembled from the dimeric esters 3-5. As an inhibitor of alpha-amylase, 1 is twice as potent as 2 and comparable to the tripeptide. NMR studies of 1 reveal four conformers in equilibrium in a 50:25:15:10 ratio; the ring conformation of the major component is similar to that of the enniatin B template, with the cis geometry of the alpha-hydroxyisovaleryl-N-methylvaline amide linkage; the other conformers differ in the position or presence of the cis amide linkage.

Ten of 19 (53%) tested isolates of Fusarium acuminatum, from different geographical origins and sources, showed in vitro antagonistic activity (inhibition at distance) to mycelial growth of F. moniliforme. Moreover, when F. acuminatum ITEM-728 was tested against 25 different fungal species, an initial inhibition at a distance was observed which was followed by the spread of the F. acuminatum mycelium over the opposite fungal colony to various degrees. Most of the F. acuminatum isolates which showed antagonistic activity proved to be enniatin B (EB) producers, and some of them also formed moniliformin (MF). The toxic activity of the methanol extract of F. acuminatum ITEM-728 towards some test microorganisms was closely related to the EB concentration. In particular, Bacillus subtilis proved to be a very sensitive test microorganism. Zusammenfassung Zehnvon 19(53 %) gepruften Fusarium acuminatum-Isolaten, dieausunterschiedlichenSubstraten und Standorten herstammten, zeigten in vitro antagonistische Aktivitat (Bildung von Hemmhofen) gegen das Myzelwachstum von F. moniliforme. Auserdem konnte mit dem F. acuminatum-Sxamra ITEM-728 in einer Prufung gegen 25 Pilzarten gezeigt werden, das, nach einer anfanglichen, als Hemmhofbildung gekennzeichneten Aktivitat, das F. acuminatum-Myzti die andere Pilzkolonie zu unterschiedlichen Graden uberwuchs. Der Hauptanteii der F. acumtnatum-Isolate mit antagonistischer Aktivitat produzierte Enniatin B (EB). Einige dieser Isolate konnten auch Moniliformin (MF) produzieren. Die tojtische Aktivitat des Methanolextraktes von F. acuminatum HEM-728 gegen einige Prufmikroorganismer wurde hauptsachlich mit der EB-Konzentration verglichen. Bacillus subtilis zeigte sich als ein besonders empfindlicber Prufmikroorgamsmus.

Fusarium sambucinum Fuckel showed antifeedant activity towards larvae of Galleria mellonella L. when incorporated into insect diet. The activity appeared mostly due to the concentration of trichothecenes present in the fungal extracts. Diacetoxyscirpenol and neosolaniol showed similar levels of activity and were significant potent antifeedants against larvae at 50 and 100 ppm. On the contrary, enniatin B showed no activity up to 100 ppm.

The effects of crystal packing forces and of hydration on the conformations of the ion carrier enniatin B, cyclo-{O-CH[-CH(CH3)2]-C'O'-N(CH3)-CH[CH(CH3)2]-C'O'-}3, in two different crystal structures belonging to the trigonal space group R3 have been analyzed theoretically by the empirical forcefield method. In one structure the central cavity of the ion carrier shrinks owing to crystal packing forces, while the c axis of the equivalent hexagonal unit cell is extended due to the presence of water molecules in the interlayer space. In the other crystal structure, a water molecule occupies the cavity and counterbalances the crystal packing forces, and therefore the observed molecular conformation resembles more closely the calculated conformation of the isolated single molecule than is the case for the first structure.

The enniatins are cyclic depsihexapeptides with antibiotic properties and are produced by various strains of Fusarium [ 1,2] . Enniatin B, a lower analogue of valinomycin with a similar ionophoretic specificity for potassium ions [3], consists of three residues each of N-methyl-L-valine and 2-D-hydroxyisovaleric acid joined in an alternating fashion (fig.1). Many peptides antibiotics like gramicidin S [4] and tyrocidine [5] are formed from the constituent amino acids in a ribosomeand ribonucleic acid-independent mechanism. This principle seems also to apply to depsipeptides as shown in the case of valinomycin [6] . In this report we describe the enzymatic assembly of enniatin B from its primary precursors, i.e. amino acid and hydroxy acid, in an energy requiring step.

The X-ray crystal structure of enniatin B (trigonal (hexagonal axes), a = 14.626 A, c = 16.309 A, space group R3) is described and the conformation compared to results of other investigations.

The spatial conformation of the “polar” form of enniatin B has been established. A conformation of the N1 type has been proposed for the “nonpolar” form.

The X-ray crystal structure of enniatin B (trigonal (hexagonal axes), a = 14.626 A, c = 16.309 A, space group R3) is described and the conformation compared to results of other investigations.