Antimutagenic and antioxidant properties of the aqueous extracts of organic and conventional grapevine Vitis labrusca cv. Isabella leaves in V79 cells.

Grapes are one of the most commonly consumed fruit, in both fresh and processed forms; however, a significant amount is disposed of in the environment. Searching for a use of this waste, the antigenotoxic, antimutagenic, and antioxidant activities of aqueous extracts from organic and conventional Vitis labrusca leaves were determined using V79 cells as model. The antigenotoxic activity was analyzed by the alkaline comet assay using endonuclease III and formamidopyrimidine DNA glycosylase enzymes. The antimutagenic property was assessed through the micronucleus (MN) formation, and antioxidant activities were assessed using 2',7'-dichlorodihydrofluorescin diacetate (DCFH-DA) assay and 2,2-diphenyl-1-picrylhydrazyl (DPPH(●)) radical scavenging, as well as with superoxide dismutase (SOD) and catalase (CAT) activity assays. In addition, phenolic content and ascorbic acid levels of both extracts were determined. Data showed that both organic and conventional grapevine leaves extracts possessed antigenotoxic and antimutagenic properties. The extract of organic leaves significantly reduced intracellular reactive oxygen species (ROS) levels in V79 cells, and displayed greater ability for DPPH(●) scavenging and higher SOD and CAT activities than extract from conventional leaves. Further, the extract from organic leaves contained higher phenolic and ascorbic acid concentrations. In summary, extracts from organic and conventional grape leaves induced important in vitro biological effects.

Negative chemical ionization gas chromatography coupled to hybrid quadrupole time-of-flight mass spectrometry and automated accurate mass data processing for determination of pesticides in fruit and vegetables.

Gas chromatography coupled to high resolution hybrid quadrupole time-of-flight mass spectrometry (GC-QTOF MS), operating in negative chemical ionization (NCI) mode and combining full scan with MSMS experiments using accurate mass analysis, has been explored for the automated determination of pesticide residues in fruit and vegetables. Seventy compounds were included in this approach where 50 % of them are not approved by the EU legislation. A global 76 % of the analytes could be identified at 1 µg kg(-1). Recovery studies were developed at three concentration levels (1, 5, and 10 µg kg(-1)). Seventy-seven percent of the detected pesticides at the lowest level yielded recoveries within the 70 %-120 % range, whereas 94 % could be quantified at 5 µg kg(-1), and the 100 % were determined at 10 µg kg(-1). Good repeatability, expressed as relative standard deviation (RSD <20 %), was obtained for all compounds. The main drawback of the method was the limited dynamic range that was observed for some analytes that can be overcome either diluting the sample or lowering the injection volume. A home-made database was developed and applied to an automatic accurate mass data processing. Measured mass accuracies of the generated ions were mainly less than 5 ppm for at least one diagnostic ion. When only one ion was obtained in the single-stage NCI-MS, a representative product ion from MSMS experiments was used as identification criterion. A total of 30 real samples were analyzed and 67 % of the samples were positive for 12 different pesticides in the range 1.0-1321.3 µg kg(-1).

Liquid chromatography Orbitrap mass spectrometry with simultaneous full scan and tandem MS/MS for highly selective pesticide residue analysis.

This paper describes the application of LC/Q-Orbitrap MS for the analysis of pesticide residues in fruit and vegetable commodities. LC/Q-Orbitrap MS working in full scan simultaneously with a single MS/MS scan was used to analyse 139 pesticide residues in QuEChERS extracts of tomato, pepper, orange and green tea. Full scan data were obtained at a resolution of 70,000 whereas MS/MS data were obtained at a resolution of 17,500. Quantitation and detection was carried out using full scan data while MS/MS data were used only for identification. MS/MS scans did not have a negative influence on quantitation under the applied conditions. Some peak area reproducibility problems were the consequence of the low sensitivity for some compounds (aldicarb, chlorpyriphos methyl, fenitrothion and fipronil) under the applied conditions. The relation between the operational parameters (viz. automatic gain control (AGC) target, maximum injection time (IT), underfill ratio, isolation window and apex trigger) and the number of automatically identified compounds was investigated. Mass error and minimal intensity of selected fragment ions were also studied. Various working modes were compared, such as full scan with single MS/MS scan and full scan with multiple MS/MS scans. In both cases, the number of automatically reported pesticides was the same. However full scan with single MS/MS scan ensured more points per peak in full scan mode and better peak area reproducibility. The evaluation of the identification and quantitation capabilities of the instrument was performed through the analysis of 100 real samples. The samples were also analysed by LC-QqQ MS/MS and the results of both analytical systems were compared. The comparison revealed that the two instruments were consistent with each other. They found the same pesticides and neither false positive nor false negatives were reported. Nevertheless the Q-Orbitrap MS allowed one to work in high resolution mass spectrometry, increasing the selectivity and, in full scan mode, permitting the retrospective analysis of the data feature that cannot be achieved with QqQ.

Viability of Cabralea canjerana extracts to control the South American fruit fly, Anastrepha fraterculus.

Several representatives of Meliaceae contain biologically active compounds that are toxic to insects with few negative effects on the environment and humans. Our study evaluated the activity of ethyl acetate and ethanol extracts from the fruit and seeds of Cabralea canjerana (Vellozo) Mart (Sapindales: Meliaceae) on Anastrepha fraterculus (Wiedemann) (Diptera: Tephritidae). Limonoids and triterpenes were detected in fruit and seed extracts. Each extract was added to an artificial diet at three concentrations and tested after 24, 48, and 72 hr of extract application. Ethyl acetate extracts were the most active ones and showed the effect of both dose and time elapses after application on the insects. The highest toxic effect on A. fraterculus adults was from ethyl acetate extracts from fruit, followed by extracts from seeds. These extracts showed antifeedant activities. Extract solutions sprinkled on fruits of Carica papaya (L.) (Brassicales: Caricaceae) caused oviposition repellency and negatively affected the biological development of A. fraterculus. Ethyl acetate extracts highly hampered oviposition, but seed extracts showed lesser oviposition deterrence. The fruit and seed extracts diminished pupal viability. Particularly, the ethyl acetate fruit extract caused malformed adults. The sex ratio was also affected, resulting in female predominance for the fruit extract, while the seed extract showed a dose-dependent effect. Low doses caused male abundance, but at higher concentrations the effect was reversed. These encouraging results showed that the C. canjerana extracts have great potential as new tools to be used in integrated pest management programs to protect fruits against A. fraterculus.

Influence of culture conditions on the biotransformation of (+)-limonene by Aspergillus niger.

The influence of the cultivation system and of the culture medium on the biotransformation of (+)-limonene by a strain of Aspergillus niger was investigated. Biooxidation products were obtained in all conditions tested. Using a laboratory bioreactor, six terpenes were identified in every medium, predominantly terpineols and carveols, whereas terpinen-4-ol and perillyl alcohol were the only terpenes found when flasks were used for culture. Perillyl alcohol and carveols predominated when the medium was tryptic soy broth (TSB), whereas the formation of terpineols was clearly favoured in malt broth (MB). Thus, there was a marked influence of the culture conditions on the results of the biotransformation. Changes in the conditions led to variations both in the type and relative amount of products obtained.

Multiple names for multi-scope analytical methods, more than a semantic issue.

The use of multiple names for multi-scope analytical methods is often confusing. More precise classification to better accommodate the scope of analytical validation and application is needed.

Holistic food safety evaluation of herbs: methods for the determination of organic and inorganic trace contaminants in Moringa stenopetala as a case study.

Moringa stenopetala is considered a superfood due to the many bioactive compounds that it provides to the diet. However, like all edible plants, it is mandatory to guarantee food safety. Thus it is necessary to develop analytical methods that can rapidly and accurately determine hazardous pollutants, to evaluate compliance with food regulations. In this regard, two multi-component procedures were developed trying to cover some of the main organic and inorganic potential contaminants. A microwave-assisted digestion followed by electrothermal atomic absorption spectrometry was used for arsenic, cadmium, and lead determination, while a modification of the QuEChERS protocol followed by gas chromatography-tandem mass spectrometry was employed for the determination of 55 pesticides from different families. Both analytical methods were thoroughly validated according to international guidelines. The analyzed samples obtained from the Uruguayan market showed compliance with both, national and international, food regulations. The holistic approach employed in this research is not commonly presented in the literature, thus constituting a novel way to face food safety.

Development of a novel and useful methodology for the simultaneous analysis of multiclass contaminants in bovine fat.

The instrumental development of tandem mass spectrometers fosters the actual trend in the trace analysis of organic compounds to the development of methods that allow the analysis of contaminants of the most diverse origin in a single analytical sample. The multiclass methods are aligned with the Who's One Health initiative while accomplishing the concepts of green chemistry. However, there are few reports of wide scope multiclass methods for the analysis of contaminants in this matrix. In this work, a method for the simultaneous determination of 62 compounds in bovine fat, was developed and validated following DG-SANTE/11813/2021 guidance. Liquid nitrogen milled fat was extracted with toluene and acetonitrile. Then a clean-up in a cryogenic bath, followed by dispersive solid phase extraction was performed. Residue determination was done using liquid chromatography for 44 compounds and gas chromatography for 18 compounds, both coupled to tandem mass spectrometry in MRM mode. The method was first developed and validated for two ectoparasiticides, then the scope was expanded for the analysis of 13 veterinary drugs and 49 pesticides. Recovery percentages were in the range of 60-134%, high matrix effect was observed in 50% of the scope of the method. Most compounds presented limits of quantification of 10 µg kg-1 in compliance with international requirements. The method was applied to monitor 49 commercial samples to evaluate its performance. Eighty percent of samples contained ethion and 10% had fluazuron, both within MRLs, highlighting the need for proper withdrawal times.

Identification of novel unique manuka honey markers using high-resolution mass spectrometry-based metabolomics.

Manuka honey is a valuable commodity produced by bees foraging the flowers of Leptospermum scoparium, a bush native to New Zealand and Australia. Due to its high value and proven health benefits, authenticity fraud in the sale of this food is a significant risk, as recounted in the literature. Four compulsory natural products must be present at minimum concentrations to authenticate manuka honey (3-phenyllactic acid, 2'-methoxyacetophenone, 2-methoxybenzoic acid, and 4-hydroxyphenyllactic acid). However, spiking other kinds of honey with these compounds and/or the dilution of manuka honey with other varieties may result in fraud going undetected. In this work, liquid chromatography coupled with high-resolution mass spectrometry and a metabolomics-based strategy has allowed us to tentatively identify 19 natural products -putative manuka honey markers-, nine of which are reported for the first time. Chemometric models applied to these markers allowed the detection of both spiking and dilution fraud attempts of manuka honey, even at 75% manuka honey purity. Thus, the herein-reported methodology can be employed in the prevention and detection of manuka honey adulteration even at low levels, and the tentatively identified markers presented in this work proved valuable for manuka honey authentication procedures.

Tannat grape pomace as an ingredient for potential functional biscuits: bioactive compound identification, in vitro bioactivity, food safety, and sensory evaluation.

Grape pomace, the main by-product of wine process, shows high potential for the development of functional foods, being a natural source of bioactive compounds and dietary fiber. Thus, the present study proposes the development of five potential functional biscuits. The five formulations were achieved by varying the Tannat grape pomace powder (TGP, 10-20% w/w total wet dough) and sweetener sucralose (2-4% w/w total wet dough) content through a factorial design with central points. TGP microbiological and pesticides analysis were performed as a food safety requirement. Identification of bioactive compounds by HPLC-DAD-MS, in vitro bioactivity (total phenol content, antioxidant by ABTS and ORAC-FL, antidiabetic and antiobesity by inhibition of α-glucosidase and pancreatic lipase, respectively) and sensory properties of the biscuits were evaluated. TGP microbiological and pesticides showed values within food safety criteria. Sensory profiles of TGP biscuits were obtained, showing biscuits with 20% TGP good sensory quality (7.3, scale 1-9) in a cluster of 37 out of 101 consumers. TGP addition in biscuits had a significant (p < 0.05) effect on total phenolic content (0.893-1.858 mg GAE/g biscuit) and bioactive properties when compared to controls: 11.467-50.491 and 4.342-50.912 µmol TE/g biscuit for ABTS and ORAC-FL, respectively; inhibition of α-glucosidase and pancreatic lipase, IC50 35.572-64.268 and 7.197-47.135 mg/mL, respectively. HPLC-DAD-MS results showed all the identified phenolic compounds in 20/4% biscuit (TGP/sucralose%) were degraded during baking. Malvidin-3-O-(6'-p-coumaroyl) glucoside, (+)-catechin, malvidin-3-O-glucoside, and (-)-epicatechin were the main phenolic compounds (in descendent order of content) found. The bioactive properties could be attributed to the remaining phenolic compounds in the biscuits. In conclusion, TGP biscuits seemed to be a promising functional food with potential for ameliorating oxidative stress, glucose and fatty acids levels with good sensory quality.

Multicompartmental monitoring of legacy and currently used pesticides in a subtropical lake used as a drinking water source (Laguna del Cisne, Uruguay).

A pilot annual monitoring survey (April 2018-March 2019) was conducted to investigate the presence of pesticides in superficial water and fish in Laguna del Cisne, one of the most critical drinking water sources in Uruguay. A total of 25 pesticide residues were detected in superficial water (89.3 % of the samples). Pesticide's temporal distribution was associated with crops and livestock practices, with higher occurrences in spring and summer than in autumn and winter. The most frequent compounds in superficial water were the insecticide chlorantraniliprole, and the herbicides glyphosate (including its metabolite AMPA) and metolachlor. The levels of Organochlorine pesticide, p,p'-DDT, was in some cases two order of magnitude above the international water quality guidelines for Ambient Water Criteria. In fishes, eight different pesticides were detected, at concentrations from 1000 to 453,000 ng·kg-1. The most frequent pesticides found were propiconazole, chlorpyrifos, and p,p'-DDE. The widespread occurrence of pesticides in fish suggests potential exposure effects on fish populations and the aquatic ecosystem. The sampling approach of this work allowed monitoring the continuous concentrations of several pesticides in surface waters and fishes to establish the influence from past and current agriculture practices in Laguna del Cisne basin. For safety measures, continuous monitoring programs must be performed in this system to prevent toxicity impacts on aquatic organisms and human health.

Effects of temperature and purity of magnesium sulfate during extraction of pesticide residues using the QuEChERS method.

Despite its many documented advantages, the QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) sample preparation approach has problems with a few unstable pesticides, partly due to the exothermic reaction generated by the use of anhydrous magnesium sulfate (anh. MgSO4) during extraction. These pesticides also tend to be difficult to analyze by GC/MS. The aim of this study was to evaluate the effect of temperature during the extraction process in a revised version of AOAC Official Method 2007.01 using anh. MgSO4 > or = 99% (fine powder) or > or = 97% (granular) purity, and the use of an ice bath for particular unstable pesticides of interest (chlorothalonil, captan, captafol, folpet, and the degradation products cis-1,2,3,6-tetrahydrophthalimide and phthalimide). Recoveries of 38 representative pesticides were measured in limes and broccoli at different extraction conditions by LC/MS/MS and low-pressure GC/MS/MS. Results showed that the difference in temperature when using > or = 99% versus > or = 97% purity anh. MgSO4 was 6-9 degrees C, which did not lead to significant differences in recoveries. The use of an ice bath aided recovery for some of the analytes in broccoli, but no significant differences were observed for limes, which already provided greater stability of the base-sensitive analytes due to acidity of the matrix.

Inclusion of 1-naphthylacetic acid and 2-(1-naphthyl)acetamide into three typical multiresidue methods for LC/MS/MS analysis of tomatoes and zucchini.

In spite of high plant growth regulator application rates, little has been reported in the literature on determination of their residues in fruits and vegetables. This would be useful in monitoring good manufacturing practices and overall safety through the enforcement of maximum residue levels (MRLs). The present work describes method validation for the determination of 1-naphthylacetic acid (NAA) and 2(1-naphthyl)acetamide (NAAm) in tomato and zucchini using the mini-Luke, ethyl acetate (EtOAc) and acetate-buffered quick, easy, cheap, effective, rugged, and safe (QuEChERS) methods. Samples were spiked at two different levels: 50 and 100 pg/kg for NAA and 20 and 100 pg/kg for NAAm. These compounds were analyzed within the same chromatographic run with LC coupled to triple quadrupole MS (LC/(QqQ)MS/MS) in positive and negative electrospray ionization [ESI(+) and ESI(-)] modes for NAAm and NAA, respectively. For analyte confirmation, LC/ESI(-)QTOF-MS was also investigated given that NAA has only one multiple reaction monitoring transition (185.1-*140.9 m/z). These three common methods were used to determine linearity, recoveries, precision (RSD), matrix effects, repeatability, and reproducibility (n = 5) for the selected matrixes. In terms of the Directorate-General for Health and Consumers (DG-SANCO) guidelines, only insignificant differences were found for the multiresidue methods tested, regardless of the commodity. Matrix-matched calibration was used, and LODs were below 10.1 pg/kg for NAA and 6.0 pg/kg for NAAm, which were lower than the MRLs established in current European Union legislation for these compounds. Obtained recoveries for NAA ranged from 87 to 107% with RSD values below 10% for mini-Luke, 83 to 107% with RSD <11% for EtOAc, and 76 to 85% with RSD <7% for QuEChERS. NAAm recoveries ranged from 74 to 102% with RSD 5 15% for mini-Luke, 76 to 97% with RSD <4% for EtOAc, and 76 to 93% with RSD < 5% for QuEChERS. The linearity of the response over two orders of magnitude was demonstrated (r2 > 0.996) for all methods used. Good repeatability and reproducibility in terms of RSD were obtained (11.3-17.8%) for these analytical methods. The results were acceptable within the studied ranges for both analytes in all methods. As these multiresidue methods are used at the present time in many laboratories, this survey indicates that these analytes should be included in their monitoring scope.

Cytotoxic, virucidal, and antiviral activity of South American plant and algae extracts.

Herpes simplex virus type 1 (HSV-1) infection has a prevalence of 70% in the human population. Treatment is based on acyclovir, valacyclovir, and foscarnet, three drugs that share the same mechanism of action and of which resistant strains have been isolated from patients. In this aspect, innovative drug therapies are required. Natural products offer unlimited opportunities for the discovery of antiviral compounds. In this study, 28 extracts corresponding to 24 plant species and 4 alga species were assayed in vitro to detect antiviral activity against HSV-1. Six of the methanolic extracts inactivated viral particles by direct interaction and 14 presented antiviral activity when incubated with cells already infected. Most interesting antiviral activity values obtained are those of Limonium brasiliense, Psidium guajava, and Phyllanthus niruri, which inhibit HSV-1 replication in vitro with 50% effective concentration (EC(50)) values of 185, 118, and 60 µg/mL, respectively. For these extracts toxicity values were calculated and therefore selectivity indexes (SI) obtained. Further characterization of the bioactive components of antiviral plants will pave the way for the discovery of new compounds against HSV-1.

Predicting Mandarin Fruit Acceptability: From High-Field to Benchtop NMR Spectroscopy.

Recent advances in nuclear magnetic resonance (NMR) have led to the development of low-field benchtop NMR systems with improved sensitivity and resolution suitable for use in research and quality-control laboratories. Compared to their high-resolution counterparts, their lower purchase and running costs make them a good alternative for routine use. In this article, we show the adaptation of a method for predicting the consumer acceptability of mandarins, originally reported using a high-field 400 MHz NMR spectrometer, to benchtop 60 MHz NMR systems. Our findings reveal that both instruments yield comparable results regarding sugar and citric acid levels, leading to the development of virtually identical predictive linear models. However, the lower cost of benchtop NMR systems would allow cultivators to implement this chemometric-based method as an additional tool for the selection of new cultivars.

Multiresidue analysis of basic, neutral, and acidic pesticides in biobeds' biomixture.

Contain between 1 and 3 bullet points highlighting the customization rather than the steps of the procedure. An analytical methodology was adjusted and validated for the analysis of 15 selected pesticides currently employed in extensive agriculture. The main application of this methodology is studying the pesticides degradation behavior using biobeds as a friendly environmental tool for the treatment of wastewaters generated in fields. The scope of the method was selected based on the most used pesticides in soybean crops in Uruguay. The novelty of this work is the inclusion of neutral and acidic herbicides such as 2,4-D, clethodim, dicamba, together with fungicides and insecticides which are usually included in Multi Residue Methods. An acetonitrile extraction methodology without a clean-up step yielded acceptable results for all the analytes. The instrumental analysis was performed using HPLC-MS/MS. The selected methodology was validated according to the SANTE guidelines. The recoveries were between 65 and 130% with RSD < 20%. The instrumental LOQs were fixed at 1 µg/L for all the compounds except for clethodim, and the method LOQs were 1 mg/kg in biomixture dry basis. These LOQs values are acceptable for biodegradation studies in biobeds. A multiresidue methodology was: • Validated for 15 pesticides in biomixture. • Acidic herbicides were included in the scope. • The method was employed for the environmental monitoring of pesticide degradation in biobeds.

Influence of Cultivar on Nutritional Composition and Nutraceutical Potential of Pecan Growing in Uruguay.

Composition and antioxidant properties of sixteen pecan [Carya illinoinensis (Wangenh) K. Koch] cultivars grown simultaneously in a single experimental orchard in Uruguay were evaluated to characterize their nutritional properties and nutraceutical potential. The percentage of oil, moisture, ash, minerals, and proteins were determined and also the fatty acid profile. Total phenolic compounds [18-41 mg gallic acid equivalents (GAEs)/g], condensed tannin [2-12 mg catechin equivalents (CEs)/g], and tocopherols (110-163 µg/g) contents were estimated in nut kernels. Total phenolic compounds (32-117 mg GAE/g), condensed tannins (130-357 mg CE/g), and total anthocyanins (1-3 mg 3-glucoside cyanidin/g) were also determined for pecan shells. The antioxidant activity in shells [57.15-578.88 µmol Trolox equivalents (TEs)/g] was 5 times higher compared with the kernels (23.15-156.60 µmol TEs/g) measured with hydrophilic ORAC. Bioactive compounds concentrations present statistically significant genetic variability between cultivars studied (p < 0.05). The presence of phenolic compounds was related with high-antioxidant capacity in kernels and shells, and a strong correlation between content of total phenolic compounds and condensed tannins in pecan shells was found. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) show association between cultivars and the observed variables. The nutritional profile in the different cultivars showed the trends described in other countries, but this work shows some significant differences that could be attributed to the specific edaphoclimatic conditions of cultivation in Uruguay.

Behavior of amoxicillin in wastewater and river water: identification of its main transformation products by liquid chromatography/electrospray quadrupole time-of-flight mass spectrometry.

The identification of transformation products (TPs) of pharmaceuticals in the environment is essentially a challenging task due to the lack of standards and the instrumental capabilities required to detect compounds (sometimes unknowns) that are produced under environmental conditions. In this work, we report the use of liquid chromatography/electrospray quadrupole time-of-flight mass spectrometry (LC/QTOF-MS/MS) as a tool for the identification of amoxicillin (AMX) and its main TPs in wastewater and river water samples. Laboratory degradation experiments of AMX were performed in both alkaline and acidic media in order to confirm that the expected transformation pathway in the aquatic media is through the ß-lactam ring cleavage. A thorough study was carried out with both standards and real samples (wastewater and river water samples). Four compounds were identified as main TPs: both amoxicillin diketopiperacine-2',5' and amoxilloic acid diastereomers. Amoxilloic acid stereoisomers are reported for the first time in environmental matrices. The transformation product (5R)-amoxicillin diketopiperacine-2',5' was frequently detected in river waters. Besides, another AMX transformation product formed during analysis was also structurally elucidated for the first time (amoxicilloic acid methyl ester) via accurate mass measurements. Collected data show that although AMX is not present as such in environmental samples, different TPs occur. This study represent a valuable indicator of the potential of LC/QTOF-MS/MS for the identification and structural elucidation of TPs in the environment using accurate MS/MS experiments, enabling thus the recognition of the environmental transformation pathway.

Development and validation of a rapid method for microcystins in fish and comparing LC-MS/MS results with ELISA.

Microcystins (MCs) are the most common cyanotoxins found worldwide in freshwater, brackish, and marine environments. The rapid and accurate analysis of MCs and nodularin (Nod-R) in fish tissue is important for determining occurrence, following trends, and monitoring exposure for risk assessment and other purposes. The aim of this study was to develop a streamlined and reliable sample preparation method for eight MCs (MC-RR, MC-YR, MC-LR, MC-WR, MC-LA, MC-LY, MC-LW, and MC-LF) and Nod-R in fish, and conduct a validation of the new method using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for analysis and compare the results with a commercial enzyme-linked immunosorbent assay (ELISA) kit. Different sample preparation methods were compared, and a simple extraction protocol with acidified acetonitrile/water (3:1) followed by hexane partitioning cleanup was found to be most effective. Thorough validation of the final method was conducted, and 90-115% recoveries were achieved for all analytes except for MC-RR, which gave 130% average recovery (isotopically labeled internal standards were unavailable to correct for possible biases). The use of electrospray ionization in the negative mode gave few interferences and minimal matrix effects in the LC-MS/MS analysis overall. Precision was typically 10-20% RSD among multiple days in experiments, detection limits were <10 ng/g in the fish tissue (catfish, basa, and swai filets), and no false-positives or false-negatives occurred in blind analyses of many spiked samples. The ELISA was unable to distinguish between MCs but was found to correctly assess the presence or absence of MCs and Nod-R in the blind-fortified fish tissues. The capability of these approaches to measure covalently bound MCs was not assessed.

Antifeedant activity of citrus waste wax and its fractions against the dry wood termite, Cryptotermes brevis.

The wood protective action of citrus wax, a waste from the citrus industry that is a mixture of citrus fruit epicuticular waxes and essential oils, was evaluated against the termite Cryptotermes brevis Walker (Isoptera: Kalotermitidae). The antifeedant index (AI) of the total wax and fractions was calculated. The total citrus wax exhibited an AI50 value of 24.69 mg/cm³, the wax after hydrodistillation showed the strongest antifeedant property (AI50 11.68 mg/cm³). Fractionation of the wax and gas chromatography-mass spectrometric analysis allowed the identification of coumarins and furancoumarins as the active compounds. These results suggest the potential use of these industrial residues as a natural approach to termite control.