Trichothecene relatives

The Fusarium spp. metabolic products sambucinol (81) and its derivatives diacetylsambucinol (82) and 3-deoxysambucinol (80) can be regarded as ll-epi-12-epitrichothecenes but are more conveniently classified with the trichothecene relatives (Table 8), as is gramilaurone (98). These compounds are excluded from Tables 1-4 and the total of trichothecenes, as is the isotrichothecin 233) whose C-nmr spectrum is indistinguishable from that of trichothecin. 

C15H2203 Sambucoin 69 F. sambucinum, F. sporotrichioides, F. poae F. culmonim, F. gramineanim, F. crookwellense 210, 65, 12 27, 43, 19 

C15H24 Trichodiene 49 T. roseum, F. sambucinum, F. sporotrichioides F. culmonim, Monascus purpureus, S. atra 216, 217, 218 219, 220, 221 

A West African shrub. The workers (61) could not exclude the possibility that this compound arose from fungal contamination of the plant material. The material tested negative for Fusarium spp. however, the pattern of the known metabolic products isolated included the diterpenoids rosenonolactone, 6/3-hydroxy-rosenonolactone, and rosololactone, and is typical of a 71 roseum strain. 

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While an entire chapter could be written about the phytopathogenesis, the selective activity of the trichothecenes relative to specific plant genera, species, and cultivars, it is time to turn our attention to the mode of action of these unique secondary metabolites. 

Scheme 1. Proven (-------) and hypothetical (—) pathways from trichodiene (49) to the trichothecene relatives listed in Table 8. (nn) Isolated...

Trichodiene (49 Scheme 1) has been detected as a volatile product of Monascus purpureus and of Stachybotrys atra, but the trichothecene relatives (Table 8), including, possibly, the loukacinols (see note c), are essentially products of Fusarium spp. and T. roseum. 

A cell-free enzyme system capable of epoxidizing the 12-ene of a trichodiene derivative has been obtained from F. culmorum (280). This system was also capable of effecting 3a-hydroxylation of the product, an interesting result since 3a-hydroxylation is commonly found amongst the trichothecene relatives. 

This account deals only with the 11-epimeric apotrichothecenes and with sambucinol (81). A more extended account covering all the trichothecene relatives derived from trichodiene is contained in Ref. 3,... 

Trichothecenes are stable at 120°C and relatively stable at 180°C, however they are destroyed at 210°C after 40 min (Kamimura, 1989). A reduction in deoxynivalenol levels of 44% in dough and baked products has been reported by Neira et al. (1997), with dough fermentation accounting for roughly 50% of this reduction. The combination of high-temperature and pressure, commonly utilized in extrusion or autoclaved food preparation, has proven far less effective, with the best results being a 12% reduction of deoxynivalenol in autoclaved cream corn (Wolf-Hall et al., 1999). 

Macromolecular Sites of Activity for the Trichothecenes. All the trichothecenes tested act, in eukaryotic systems, at the site of the 60s ribosome subunit and specifically inhibit peptidyl transferase. However, the trichothecenes are divided into two specific classes relative to this activity and are either initation inhibitors or elongation and/or termination inhibitors. The initiation inhibitors ares 15-acetoxyscirpen-diol 4-acetylnivalenol diacetoxyscirpenol HT-2 toxin nivalenol T-2 toxin scirpentriol verrucarin A (4-0, 11b). 

Negative ion chemical ionization mass spectrometry can be used to confirm trichothecenes and quantitate DON in naturally contaminated grains and feeds. The effects of interferences are eliminated by selectively scanning the relatively high molecular weight fragments and molecular ion of the trichothecene derivatives. Separation of a mixture of trichothecene derivatives is accomplished in 15 minutes on a packed GC column. 

The trichothecenes make up a family of closely related chemical compounds called sesquiterpenoids (Figure 34-1). The structures of close to 150 derivatives of the trichothecenes are described in the scientific literature.35,43 The specific side structures of the most abundant of the naturally occurring trichothecenes are shown in Table 34-1. Because of its availability and relatively high toxicity, T-2 toxin has been the most extensively studied trichothecene mycotoxin. 

The trichothecene mycotoxins are nonvolatile, low-molecular-weight (MW 250-550) compounds.43 This group of mycotoxins is relatively insoluble in water but highly soluble in acetone, ethyl acetate, chloroform, dimethyl sulfoxide (DMSO), ethanol, methanol, and propylene glycol.43 Purified trichothecenes generally have a low vapor pressure, but... 

RELATIVE ACUTE PARENTERAL TOXICITY OF THE MOST ABUNDANT TRICHOTHECENE MYCOTOXINS... 

When trichothecene mycotoxins of relatively low toxicity (crotocin and tricho-tecin) were applied to the volar surface of human forearm or to the human head, reddening and irritation occurred within a few hours of exposure, and was followed by inflammation or scrabbling that healed in 1 to 2 weeks.66... 

Screening tests for the trichothecene mycotoxins are generally simple and rapid but, with the exception of the immunochemical methods, are nonspecific. A number of bioassay systems have been used for the identification of trichothecene mycotoxins.73 Although most of these systems are very simple, they are not specific, their sensitivity is generally relatively low compared to other methods, and they require that the laboratory maintain vertebrates, invertebrates, plants, or cell cultures. Thin-layer chromatography (TLC) is one of the simplest and earliest analytical methods developed for myco-toxin analysis. Detection limits for trichothecene mycotoxins by TLC is 0.2 to 5 ppm (0.2 to 5 pg/ mL). Therefore, extracts from biomedical samples would have to be concentrated 10- to 1,000-fold to screen for trichothecene mycotoxins. 

USE SEPARATE ROOM FOR HANDLING STANDARDS WHERE SAMPLES ARE NEVER AUOWED USE STANDARD COCKTAIL OF TRICHOTHECENES AT KNOWN RELATIVE CONCENTRATION FOR ALL OUALITY CONTROL AND STANDARD SAMPLES. 

Some large scale and/or blocked fermentations with Trichothecium roseum or Fusarium spp. have yielded a number of metabolic products with structures closely related to the trichothecenes. These relatives, 49 in number (including 2 likely artefacts), an addition of six since 1995, are listed in Table 8 (structures Scheme 1), together with their sources. They consist mainly of... 

Trichodiene (49), the last hydrocarbon intermediate in the pathway, has been isolated from T. roseum, from Fusarium spp., and also from S. atra (Table 8). Specific labelling experiments have shown it to be a precursor of trichodiol (55) and 12,13-epoxy trichothecene (73), in addition to trichothecolone (101), in T. roseum (277) it is also a precursor of isotrichodiol (53) (219) and 3-acetylvomitoxin (104) in F. culmorum (278). However, relatively little work has been done on this stage of the pathway to simple trichothecenes in Myrothecium, Stachybotrys and Trichoderma spp. 

In Tables 4-7, the within-laboratory reproducibility standard deviation (sw), the reproducibility limit (Rw), and the relative standard deviation (RSDw), as well as CV derived from Horwitz equation are given for the contamination levels of 0.1 mg/kg, 0.3 mg/kg, 0.5 mg/kg, and 1.0 mg/kg. The results for sw, Rw and RSDw for each individual trichothecene were calculated from six experiments done in duplicates at the contamination level of 0.1 mg/kg and from ten experiments done in duplicates at the other three contamination levels except those for DON and nivalenol at the concentration levels of 0.3 mg/kg and 1.0 mg/kg which were calculated from nine experiments done in duplicates since one result at each of the two contamination levels was eliminated by the Cochran test. The experimental RSDw values were compared to the CV values derived from Horwitz equation. Majority of experimental RSDw values were lower than reference values, only a few exceeded it. However, they were much lower than upper limits for RSDr given in Regulation (EC) No 401/2006 (European Commission, 2006a) which were 40% for DON and 60% for T-2 and HT-2, thus the determined RSDw are considered acceptable. 

Deoxynivalenol (DON) is the most important trichothecene worldwide and is often detected in small cereal grains such as barley, oats and wheat. Due to relatively good thermal stability DON can be transmitted from contaminated barley into the final product (Schwarz et al., 1995). DON is frequently detected in barley and in commercial beer (discussed in Section 6.6.2). The occurrence of DON is largely dependent on weather conditions in the particular location and year. DON is predominantly produced by F. culmorum and F. graminearum species. 

GC detection systems for trichothecenes primarily featured classical detectors such as ECD, especially for fiuorinated derivatives however, mass spectrometers are nowadays routinely used for this purpose. The selectivity of a detector is an important issue as trichothecenes are present in relatively complicated matrices and create high background during the analysis. Three types of mass spectrometers have been used for trichothecene analysis by GC/MS, and quadrupole instruments are used most frequently. 

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