Structure-activity relationship (SAR) studies of benzoxazinones, their degradation products, and analogues. Phytotoxicity on problematic weeds Avena fatua L. and Lolium rigidum Gaud

Avena fatua L. (wild oat) and Lolium rigidum Gaud. (rigid ryegrass) are highly problematic weeds affecting a wide variety of cereal crops worldwide. The fact that both of these weeds have developed resistance to several herbicide groups made them optimal candidates as target organisms for ongoing research about the potential application of allelochemicals and analogue compounds as natural herbicide models. Benzoxazinones, a family of natural allelochemicals present in corn, wheat, and rye, including 2,4-dihydroxy-(2H)-1,4-benzoxazin-3(4H)-one and 2,4-dihydroxy-7-methoxy-(2H)-1,4-benzoxazin-3(4H)-one, together with some degradation products, found in crop soils as well as in other systems, and some synthetic analogues of them were tested on wild oat and rigid ryegrass seeds; the results were statistically treated, and some structure-activity relationships, useful in further development of natural herbicide models, were elucidated. The most active compounds were the synthetic benzoxazinone 2-acetoxy-(2H)-1,4-benzoxazin-3(4H)-one and the degradation product 2-aminophenoxazin-3-one, with highly significant inhibition on the development of both weeds. The ecological role of these compounds is discussed by considering both degradability and phytotoxicity. The bioactivity of aminophenoxazines has been correlated by their aqueous solubility-lipophilicity predicted by means of computational methods.     

Structure-activity relationship studies of benzoxazinones and related compounds. Phytotoxicity on Echinochloa crus-galli (L.) P. Beauv

Echinochloa crus-galli (E. crus-galli; barnyardgrass) is a weed widely distributed. It constitutes a serious weed problem in 42 countries and has been found in at least 27 more. It is the world's main weed of rice affecting up to 36 crops worldwide. Several biotypes of this plant, with resistance to herbicides with different modes of action have evolved. In our ongoing studies regarding the potential application of benzoxazinones and their soil degradation products for weed control, a complete structure-activity relationships (SARs) study was made by using barnyardgrass as the target plant. Compounds used in this study were previously tested on a wide variety of standard target species (STS), and they include natural allelochemicals 2-O-beta-D-glucopyranosyl-4-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one (DIBOA-Glc), 2,4-dihydroxy-7-methoxy-(2H)-1,4-benzoxazin-3(4H)-one (DIMBOA), and 2,4-dihydroxy-(2H)-1,4-benzoxazin-3(4H)-one (DIBOA), together with some degradation derivatives found in wheat crop soil and some synthetic analogues. Their phytotoxicity on E. crus-galli is discussed and compared with the results obtained from previous screening. This work constitutes the next step in the search for natural herbicide models based on benzoxazinones and their degradation products. The most active compounds were the degradation product 2-aminophenol (APH) and the synthetic analogue 4-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one (D-DIBOA). Their activities confirm the direction proposed in our previous SAR study, which establishes D-DIBOA to be the best lead for natural herbicide model development with benzoxazinone structure.     

Optimization of benzoxazinones as natural herbicide models by lipophilicity enhancement

Benzoxazinones are plant allelochemicals well-known for their phytotoxic activity and for taking part in the defense strategies of Gramineae, Ranunculaceae, and Scrophulariceae plants. These properties, in addition to the recently optimized methodologies for their large-scale isolation and synthesis, have made some derivatives of natural products, 2,4-dihydroxy-(2H)-1,4-benzoxazin-3-(4H)-one (DIBOA) and its 7-methoxy analogue (DIMBOA), successful templates in the search for natural herbicide models. These new chemicals should be part of integrated methodologies for weed control. In ongoing research about the structure-activity relationships of benzoxazinones and the structural requirements for their phytotoxicity enhancement and after characterization of the optimal structural features, a new generation of chemicals with enhanced lipophilicity was developed. They were tested on selected standard target species and weeds in the search for the optimal aqueous solubility-lipophilicity rate for phytotoxicity. This physical parameter is known to be crucial in modern drug and agrochemical design strategies. The new compounds obtained in this way had interesting phytotoxicity profiles, empowering the phytotoxic effect of the starting benzoxazinone template in some cases. Quantitative structure-activity relationships were obtained by bioactivity-molecular parameters correlations. Because optimal lipophilicity values for phytotoxicity vary with the tested plant, these new derivatives constitute a more selective way to take advantage of benzoxazinone phytotoxic capabilities.     

2,4-Dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA) and 6-methoxy-benzoxazolin-2-one (MBOA) levels in the wheat rhizosphere and their effect on the soil microbial community structure

Despite increasing knowledge of 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA) and 6-methoxy-benzoxazolin-2-one (MBOA) as allelochemicals involved in the defense of wheat against pests, relatively little is known about their levels in the rhizosphere and interactions with the soil microbial community. This study quantified DIMBOA and MBOA in the wheat rhizosphere and analyzed the soil microbial community structure. MBOA rather than DIMBAO was found in the wheat rhizosphere, and its concentration varied with cultivars, plant densities, and growth conditions. Wheat could detect the presence of competing weeds and respond by increased MBOA in the rhizosphere. There was a linear positive relationship between the MBOA level in the wheat rhizosphere and soil fungi/bacteria. When DIMBOA was applied to soil, yielding MBOA increased soil fungi. There were different phospholipid fatty acid (PLFA) patterns in soil incubated with DIMBOA and MBOA. These results suggested that DIMBOA and MBOA could affect the soil microbial community structure to their advantage through the change in fungi populations.     

New herbicide models from benzoxazinones: aromatic ring functionalization effects

The utility of benzoxazinones and some of their synthetic derivatives in the search for new leads for herbicide model development has been widely discussed. As the benzoxazinone skeleton contains three different potential areas for functionalization (C-2, N-4, and aromatic protons H-5, H-6, H-7, and H-8), and the first two have already been optimized, the main objective of this work was the substitution of aromatic protons for different substituent types and the study of the effects of the prepared chemicals on selected standard target species (STS) and weeds. Thus, different combinations of aromatic substituents, including methoxy, methoxycarbonyl, fluorine, chlorine, and trifluoromethyl, were introduced at different positions. Phytotoxicity results were successfully correlated with steric and electronic molecular parameters, the resulting molecular volume (V) and dipole moment (mu) being the most influential ones. Halogenations at C-6 and fluorination at C-7 were the most successful modifications. Compounds 6-fluoro-(2H)-1,4-benzoxazin-3(4H)-one (6F-D-DIBOA), 7-fluoro-(2H)-1,4-benzoxazin-3(4H)-one (7F-D-DIBOA), and 6-chloro-(2H)-1,4-benzoxazin-3(4H)-one (6Cl-D-DIBOA) had the highest phytotoxic activities. The dose-response profiles on wheat and two of its most common weeds (Lolium rigidum Gaud. and Avena fatua L.) were compared by means of a proposed selectivity index, which displayed 7F-D-DIBOA as the most selective of the tested benzoxazinones.     

Joint action of benzoxazinone derivatives and phenolic acids

The joint action of binary and ternary mixtures of benzoxazinone derivatives and phenolic acids was studied using the additive dose model (ADM) as reference model. The activity of fixed-ratio mixtures of phenolic acids [ferulic acid (FA), p-coumaric acid (CA), vanillic acid (VA), and p-hydroxybenzoic acid (HBA)] and benzoxazinone derivatives [2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA), 6-methoxybenzoxazolin-2-one (MBOA), benzoxazolin-2-one (BOA), 2-aminophenol (AP), and N-(2-hydroxyphenyl)acetamide (HPAA)] on Lolium perenne and Myosotis arvensis root growth was assessed in Petri dishes. Root length was recorded 6 days after seeding, and EC(50) and EC(90) values were estimated using nonlinear regression analyses. The benzoxazinone derivatives were found to be more phytotoxic than the phenolic acids, particularly on M. arvensis. Binary mixtures of phenolic acids responded predominantly additively on both plant species. Deviations from additivity were species-specific with antagonistic responses on L. perenne and synergistic responses on M. arvensis. Similarly, binary mixtures of benzoxazinone derivatives also followed the ADM, although synergistic responses were observed for BOA + AP and BOA + HPAA. Binary and ternary mixtures of benzoxazinone derivatives and phenolic acids responded primarily antagonistically; however, a significant synergistic performance was observed with DIMBOA + FA and DIMBOA + VA on L. perenne. These results do not support the assumption that allelopathic effects of wheat can be attributed to synergistic effects of otherwise weakly active allelopathic compounds, and it is suggested that future research be directed toward identifying and studying the effects of other potential allelochemicals including the degradation products of the most abundant wheat allelochemicals.     

Fate of benzoxazinone allelochemicals in soil after incorporation of wheat and rye sprouts

Growing cereals (especially rye), which are incorporated into the soil to increase soil fertility or organic matter content, is a common practice in crop rotation. The additional sanitizing effect of this incorporation has often been appreciated and is said to be due to leaching of benzoxazinones and subsequent formation of benzoxazolinones. In this study wheat (Stakado) and rye (Hacada) sprouts were incorporated into soil in amounts that simulated agricultural practice. By extraction and subsequent LC-MS analysis the disappearance and appearance of benzoxazinones, benzoxazolinones, and phenoxazinones in soil were followed. In the wheat experiments 6-methoxybenzoxazolin-2-one (MBOA) was detected as the main compound. 2-Hydroxy-7-methoxy-1,4-benzoxazin-3-one (HMBOA) and 2-hydroxy-1,4-benzoxazin-3-one (HBOA) were detected as well. No phenoxazinones were detected. For the rye experiment the picture was more complex. In the first 2 days of incubation MBOA and 2,4-dihydroxy-1,4-benzoxazin-3-one (DIBOA) were detected as the main allelochemicals along with HBOA, HMBOA, and benzoxazolin-2-one (BOA), in decreasing order. Later in the incubation period some 2-amino-3H-phenoxazin-3-one (APO) was detected and the amount of HBOA increased considerably and decreased again. The profiling of the benzoxazinone metabolites and their derivates in soil was dynamic and time-dependent. The highest concentrations of most of the compounds were seen at day 1 after incorporation. A maximum concentration was reached at day 4 for a few of the compounds. This study is the first of its kind that shows the dynamic pattern of biologically active benzoxazinone derivates in soil after incorporation of wheat and rye sprouts. Methods for organic synthesis of HBOA and HMBOA were developed as part of the study.     

Isolation and synthesis of allelochemicals from gramineae: benzoxazinones and related compounds

Compounds with a (2H)-1,4-benzoxazin-3(4H)-one skeleton have attracted the attention of phytochemistry researchers since 2,4-dihydroxy-(2H)-1,4-benzoxazin-3(4H)-one (DIBOA) and 2,4-dihydroxy-7-methoxy-(2H)-1,4-benzoxazin-3(4H)-one (DIMBOA) were isolated from plants belonging to the Poaceae family. These compounds exhibit interesting biological properties, such as phytotoxic, antimicrobial, antifeedant, antifungal, and insecticidal properties. These chemicals, in addition to a wide variety of related compounds involved in their metabolism, detoxification mechanisms, and degradation on crop soils and other systems, have high interest and in some cases potential agronomic utility. This paper presents a complete review of the methods employed for their synthetic obtention in addition to some of the authors' own contributions to their chemistry. The degradation and phytotoxicity experiments carried out in ongoing research into the potential agronomic utility of these compounds required large amounts of them, which were obtained from natural sources. This paper presents a modified methodology to access DIMBOA from Zea mays cv. Apache and to obtain 2-O-beta-D-glucopyranosyl-2,4-dihydroxy-(2H)-1,4-benzoxazin-3(4H)-one (DIBOA-Glc) and DIBOA from Secale cereale L. New synthetic methodologies were employed for the obtention of the lactams 2-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one and 2-hydroxy-7-methoxy-(2H)-1,4-benzoxazin-3(4H)-one and the malonamic acids N-(2-hydroxyphenyl)malonamic acid and N-(2-hydroxy-7-methoxyphenyl)malonamic acid. The aminophenoxazines 2-amino-7-methoxyphenoxazin-3-one and 2-acetamido-7-methoxyphenoxazin-3-one have been synthesized in the authors' laboratory by novel procedures. All of the methodologies employed allowed the desired compounds to be obtained in high yield and in an easy-to-scale manner.     

Structure-Activity Relationships (SAR) studies of benzoxazinones, their degradation products and analogues. phytotoxicity on standard target species (STS)

Benzoxazinones 2,4-dihydroxy-7-methoxy-(2H)-1,4-benzoxazin-3(4H)-one (DIMBOA) and 2,4-dihydroxy-(2H)-1,4-benzoxazin-3(4H)-one (DIBOA) have been considered key compounds for understanding allelopathic phenomena in Gramineae crop plants such as corn (Zea mays L.), wheat (Triticum aestivum L.), and rye (Secale cereale L.). The degradation processes in the environment observed for these compounds, in which soil microbes are directly involved, could affect potential allelopathic activity of these plants. We present in this work a complete structure-activity relationships study based on the phytotoxic effects observed for DIMBOA, DIBOA, and their main degradation products, in addition to several synthetic analogues of them. Their effects were evaluated on standard target species (STS), which include Triticum aestivum L. (wheat) and Allium cepa L. (onion) as monocots and Lepidium sativum L. (cress), Lactuca sativa L. (lettuce), and Lycopersicon esculentum Will. (tomato) as dicots. This permitted us to elucidate their ecological role and to propose new herbicide models based on their structures. The best phytotoxicity results were shown by the degradation chemical 2-aminophenoxazin-3-one (APO) and several 2-deoxy derivatives of natural benzoxazinones, including 4-acetoxy-(2H)-1,4-benzoxazin-3(4H)-one (ABOA), 4-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one (D-DIBOA), and 4-hydroxy-7-methoxy-(2H)-1,4-benzoxazin-3(4H)-one (D-DIMBOA). They showed high inhibitory activity over almost all species growth. The fact that APO is a degradation product from DIBOA with high phytotoxicity and stability makes it possible to assign an important ecological role regarding plant defense mechanisms. 2-Deoxy derivatives of natural benzoxazinones display a wide range of activities that allow proposing them as new leads for natural herbicide models with a 1,4-benzoxazine skeleton.     

Herbicidal effects of soil-incorporated wheat

The hydroxamic acid 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA) and the benzoxazolinones benzoxazolin-2-one (BOA) and 6-methoxybenzoxazolin-2-one (MBOA) have been identified as important allelochemicals in wheat. This study examines the possibility of exploiting the allelopathic properties of wheat as a weed control strategy by cultivating wheat as a precrop and incorporating plant residues into the soil before the next crop is sown. Different wheat varieties were cultivated in field plots during two seasons in both conventional and organic farming systems. Plants were sampled at various growth stages, and their contents of DIMBOA, MBOA, and BOA were determined by chemical analyses. The wheat samples were incorporated into soil, and the effect on germination and growth of 12 different weed species was examined in pot experiments under controlled conditions. In some cases significant effects were obtained, but the results were inconsistent and the effects were not correlated to the content of DIMBOA, MBOA, and BOA in the incorporated wheat plants. ED50 doses of the pure compounds were estimated in dose-response experiments in Petri dishes, and these turned out to be much higher than the predicted maximum concentrations of DIMBOA, MBOA, and BOA in the soil water following incorporation. The study shows that a prerequisite for exploiting the incorporation of wheat residues as a weed control strategy is the development of wheat varieties with an increased content of allelochemicals.     

Isolation and identification of insecticidal components from Citrus aurantium fruit peel extract

Three active components were identified by bioassay-guided fractionation of bitter orange ( Citrus aurantium L.) fruit peel petroleum ether extract. Silica gel fractionation of the extract yielded a fraction that inflicted up to 96% mortality to adults of the olive fruit fly Bactrocera oleae (Gmelin) three days post-treatment. Subsequent HPLC purification of the active fraction resulted in the isolation of three components, eluted in fractions F 222, F 224, and F 226, that induced adult mortality. Considering the data obtained from UV, FTIR, MS, and (1)H NMR spectra, they were identified as 7-methoxy-8-(3'-methyl-2'-butenyl)-2 H-1-benzopyran-2-one (osthol), 4-methoxy-7 H-furo[3,2- g]benzopyran-7-one (bergapten), and 4-(( E)-3'-methyl-5'-(3',3'-dimethyloxiran-2'-yl)pent-2'-enyloxy)-7 H-furo[3,2- g][1]benzopyran-7-one (6',7'-epoxybergamottin). Our results are in concordance with those reported in the literature and were further verified by direct comparison to authentic components. 6',7'-Epoxybergamottin was toxic when tested individually, while bergapten and osthol were found to act synergistically to 6',7'-epoxybergamottin.     

Isolation, characterization, and antioxidant activity of bromophenols of the marine red alga Rhodomela confervoides

A total of 19 naturally occurring bromophenols, with six new and 13 known structures, were isolated and identified from the methanolic extract of the marine red alga Rhodomela confervoides. The new compounds were identified by spectroscopic methods as 3,4-dibromo-5-((methylsulfonyl)methyl)benzene-1,2-diol (1), 3,4-dibromo-5-((2,3-dihydroxypropoxy)methyl)benzene-1,2-diol (2), 5-(aminomethyl)-3,4-dibromobenzene-1,2-diol (3), 2-(2,3-dibromo-4,5-dihydroxyphenyl)acetic acid (4), 2-methoxy-3-bromo-5-hydroxymethylphenol (5), and (E)-4-(2-bromo-4,5-dihydroxyphenyl)but-3-en-2-one (6). Each compound was evaluated for free radical scavenging activity against DPPH (α,α-diphenyl-β-dipicrylhydrazyl) and ABTS [2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt] radicals. Most of them exhibited potent activities stronger than or comparable to the positive controls butylated hydroxytoluene (BHT) and ascorbic acid. The results from this study suggest that R. confervoides is an excellent source of natural antioxidants, and inclusion of these antioxidant-rich algal components would likely help prevent the oxidative deterioration of food.     

Sesquiterpenoids isolated from the flower buds of Tussilago farfara L. inhibit diacylglycerol acyltransferase

Inhibition of acyl CoA:diacylglycerol acyltransferase (DGAT), which is a key enzyme in triglyceride synthesis in eukaryotic organisms, has been proposed as one of the drug targets for treating obesity, type II diabetes mellitus, and metabolic syndrome. Bioassay-guided fractionation of EtOH extract of the flower buds of Tussilago farfara , using an in vitro DGAT enzyme assay, resulted in the isolation of four known sesquiterpenoids, tussilagonone (1), tussilagone (2), 7beta-(3-ethyl-cis-crotonoyloxy)-1alpha-(2-methylbutyryloxy)-3,14-dehydro-Z-notonipetranone (3), and 8-angeloylxy-3,4-epoxy-bisabola-7(14),10-dien-2-one (4). DGAT1 inhibitory activity was studied by in vitro DGAT assay using rat liver microsomes and HepG2 cell microsomes. They showed DGAT1 inhibition with IC(50) values of 99.2 (1), 18.8 (2), 47.0 (3), and 211.1 (4) microM (for rat liver microsomes) and >1 mM (1), 49.1 (2), 160.7 (3), and 294.4 (4) microM (for HepG2 cell microsomes), respectively. Compound 2 showed the most potent inhibition against microsomal DGAT1 derived from rat liver and human hepatocellular carcinoma HepG2 cells and also significantly inhibited triglyceride synthesis by suppressing incorporation of [(14)C]acetate or [(14)C]glycerol into triglycerides in HepG2 cells. These findings suggest that tussilagone is a potential lead compound in the treatment of obesity and type 2 diabetes.     

Quantitative model studies on the efficiency of precursors in the formation of cooling-active 1-pyrrolidinyl-2-cyclopenten-1-ones and bitter-tasting cyclopenta-[b]azepin-8(1H)-ones

The yields of the cooling-active compounds 3-methyl-2-(1-pyrrolidinyl)-2-cyclopenten-1-one (1) and 5-methyl-2-(1-pyrrolidinyl)-2-cyclopenten-1-one (2) as well as the bitter tastants 7-methyl-2,3,6,7-tetrahydrocyclopenta-[b]azepin-8(1H)-one (3) and 7-methyl-2,3,4,5,6,7-hexahydrocyclopenta-[b]azepin-8(1H)-one (4) obtained by heating mixtures of possible Maillard-type precursors in model systems varying in temperature, pH value, or water content were determined quantitatively. The results showed that hexose-derived cyclotene is the common precursor for all four tastants and that the formation of each individual tastant is strongly determined by the structure of the nitrogen-containing precursor, e.g., reaction of cyclotene with pyrrolidine formed by thermal decarboxylation of L-proline produced the cooling compounds 1 and 2 only, whereas in the presence of 1-pyrroline formed upon Strecker reactions of L-proline, the bitter tasting azepinone 3 was produced exclusively. In contrast, the structure of the secondary amino acid L-proline enabled the formation of compound 4, whereas the pyrrolidine and 1-pyrroline, respectively, do not generate this tastant. In addition, a nonvolatile, tasteless intermediate, (S)-3-methyl-2-[(2'-carboxy)-1-pyrrolidinyl]-2-cyclopenten-1-one (5), was isolated from the cyclotene/L-proline reaction mixture and could be confirmed as an efficient precursor for the cooling compound 1. The data, obtained by these studies, are the scientific basis to tailor the desired overall flavor of foods by means of a more controlled Maillard-type technology.     

Insecticidal activity of Paraherquamides, including paraherquamide H and paraherquamide I, two new alkaloids isolated from Penicillium cluniae

Paraherquamide H (1) and paraherquamide I (2), two new compounds of the paraherquamide (PHQ) family, together with the already known paraherquamide A (3), paraherquamide B (4), paraherquamide E (5), VM55596 (N-oxide paraherquamide) (6), paraherquamide VM55597 (7), and five known diketopiperazines (8-12) have been isolated from the culture broth of Penicillium cluniae Quintanilla. The structure of 1 and 2, on the basis of NMR and MS analysis, was established. It is worth noticing that, in both cases, an unusual oxidative substitution in C-16 was found, which had only previously been detected in PHQ 7. Isolated compounds were tested for insecticidal activity against the hemipteran Oncopeltus fasciatus Dallas. Mortality data have allowed preliminary structure activity relationships to be proposed. The most potent product was 5 with a LD(50) of 0.089 microg/nymph.     

Concentration of benzoxazinoids in roots of field-grown wheat (Triticum aestivum L.) varieties

Benzoxazinones are naturally occurring secondary metabolites of some Gramineae plants, responsible for their resistance to some pathogenic fungi and for their allelopathic action. Six varieties of winter wheat grown in fields under organic or conventional systems and 11 old accessions were tested for two consecutive seasons and three plant development stages for the concentration in their roots of cyclic hydroxamic acids and their degradation products. This is the first report of six benzoxazinones analyzed in plants grown in the field. An analytical technique employing LC-DAD was used for determination. It was shown that 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one, its degradation product 6-methoxybenzoxazolin-2-one, and the lactam 2-hydroxy-7-methoxy-1,4-benzoxazin-2-one were predominant compounds in all tested samples. Their concentrations significantly differed with plant development stage and season, but no significant differences were found between varieties and between plant cultivation systems. The concentrations of 2,4-dihydroxy-1,4-benzoxazin-3-one (DIBOA) and its degradation product benzoxazolin-2-one (BOA) were much lower, ranging from 60 to 430 mg/kg of dry matter, depending on accession, stage of development, and season. There was no significant difference found between plants grown in different cultivation systems, but there were significant differences between old and new varieties; concentrations of DIBOA and its derivatives were significantly lower in old accessions. It was concluded that the concentrations of DIBOA and BOA, which are precursors of highly fungicidal 2-aminophenol, 2-amino-3H-phenoxazin-3-one, and 2-acetylamino-3H-phenoxazin-3-one, are theoretically high enough to protect plants against some soilborne pathogens.     

Effects of some benzoxazinoids on in vitro growth of Cephalosporium gramineum and other fungi pathogenic to cereals and on Cephalosporium stripe of winter wheat

The benzoxazolinones benzoxazolin-2(3H)-one (BOA) and 6-methoxybenzoxazolin-2(3H)-one (MBOA) and selected degradation products of these compounds were examined for their in vitro antifungal activity against Cephalosporium gramineum, Gaeumannomyces graminis var. graminis, and Fusarium culmorum. BOA was also applied to the soil-incorporated inoculum of C. gramineum to test its capability of reducing Cephalosporium stripe disease in winter wheat. MBOA reduced the mycelial growth of G. graminis var. tritici, C. gramineum, and F. culmorum by 50% (EC50) at the concentrations of 77, 134, and 271 microg/mL of corn meal agar, respectively, and the corresponding BOA EC50 values for the fungi were 11, 189, and 456 microg/mL. BOA degradation products 2-amino-3H-phenoxazin-3-one (APO), 2-acetylamino-3H-phenoxazin-3-one (AAPO), and o-aminophenol (o-AP) were much more inhibitory to the growth of C. gramineum and G. graminis var. tritici than the parent compounds. APO, AAPO, and o-AP EC50 values were found to be as low as 0.58, 4.57, and 1.4 microg/mL, respectively, for C. gramineum and 0.78, 2.18, and 0.80 microg/mL for G. graminis var. tritici. These compounds applied at the corresponding concentrations did not significantly affect the mycelial growth of F. culmorum. The treatment of C. gramineum inoculum with a 1% water solution of BOA resulted in a significant reduction infection of winter wheat with C. gramineum as compared to the control with the untreated inoculum,but this treatment was not as effective as the application of a commercial fungicide.     

Search for a standard phytotoxic bioassay for allelochemicals. Selection of standard target species

In the search for a standard bioassay of phytotoxicity for allelochemicals, 22 commercial varieties of eight plant species [four dicotyledons: lettuce (Compositae), carrot (Umbelliferae), cress (Cruciferae), tomato (Solanaceae); and four monocotyledons: onion (Liliaceae), barley, wheat, and corn (Gramineae)] proposed as models for the most common weed families have been tested at different pH and solution volumes per set conditions. Nine commercial varieties selected as standard target species (STS) were tested with standard commercial herbicides to ensure their sensitivity to phytotoxic compounds. Results are discussed to establish the proper growth requirements, and sensitivity of commercial seeds of STS and to find the most suitable commercial herbicides that allow comparison with an internal standard to validate the response of potential allelochemicals.     

Antioxidative compounds from the outer scales of onion

Antioxidative compounds were isolated from the methanol extract of dry outer scales of onion (Allium cepa L.). Nine phenolic compounds (1-9) were finally obtained by reversed-phase high-performance liquid chromatography, and their structures were elucidated by NMR and mass spectrometry analyses. They were the six known compounds, protocatechuic acid (1), 2-(3,4-dihydroxybenzoyl)-2,4,6-trihydroxy-3(2H)-benzofuranone (2), quercetin 4'-O-beta-D-glucopyranoside (3), quercetin (5), 4'-O-beta-d-glucopyranoside of quercetin dimer (7), and quercetin dimer (8), and three novel compounds, condensation products of quercetin with protocatechuic acid (4), adduct of quercetin with quercetin 4'-O-beta-D-glucopyranoside (6), and quercetin trimer (9). These phenolic compounds were tested for their antioxidant properties using autoxidation of methyl linoleate in bulk phase or free radical initiated peroxidation of soybean phosphatidylcholine in liposomes. The flavonoid compounds having o-dihydroxy substituent in the B-ring were shown to be effective antioxidants against nonenzymic lipid peroxidation.