Daniele Marton

Assessment of the environmental exposure of honeybees to particulate matter containing neonicotinoid insecticides coming from corn coated seeds.

Since seed coating with neonicotinoid insecticides was introduced in the late 1990s, European beekeepers have reported severe colony losses in the period of corn sowing (spring). As a consequence, seed-coating neonicotinoid insecticides that are used worldwide on corn crops have been blamed for honeybee decline. In view of the currently increasing crop production, and also of corn as a renewable energy source, the correct use of these insecticides within sustainable agriculture is a cause of concern. In this paper, a probable--but so far underestimated--route of environmental exposure of honeybees to and intoxication with neonicotinoid insecticides, namely, the atmospheric emission of particulate matter containing the insecticide by drilling machines, has been quantitatively studied. Using optimized analytical procedures, quantitative measurements of both the emitted particulate and the consequent direct contamination of single bees approaching the drilling machine during the foraging activity have been determined. Experimental results show that the environmental release of particles containing neonicotinoids can produce high exposure levels for bees, with lethal effects compatible with colony losses phenomena observed by beekeepers.

Fatal powdering of bees in flight with particulates of neonicotinoids seed coating and humidity implication

Losses of honeybees have been reported in Italy concurrent with the sowing of corn coated with neonicotinoids using a pneumatic drilling machine. Being unconvinced that solid particles containing systemic insecticide, falling on the vegetation surrounding the sown area, could poison bees foraging on contaminated nectar and pollen, the effect of direct aerial powdering was tested on foragers in free flight near the drilling machine. Bees were conditioned to visit a dispenser of sugar solution whilst a drilling machine was sowing corn along the flight path. Samples of bees were captured on the dispenser, caged and held in the laboratory. Chemical analysis showed some hundred nanograms of insecticide per bee. Nevertheless, caged bees, previously contaminated in flight, died only if kept in conditions of high humidity. After the sowing, an increase in bee mortality in front of the hives was also observed. Spring bee losses, which corresponded with the sowing of corn‐coated seed, seemed to be related to the casual encountering of drilling machine during foraging flight across the ploughed fields.

Allylstannation. II. A total "cis-preference" in the addition of n-Bu2ClSnCH(CH3)CHCH2 to aldehydes

Abstract 1-Buten-3-yldi-n-butylchlorotin, formed by redistribution of ( E Z )-2-butenyltri-n-butyltin and Bu2SnCl2, reacts readily with neat RCHO (R  C2H5, C2H5(CH3)CH, (CH3)2CH, (CH3)3C and C6H5) to give high yields (80–100%) of alcohols of the type RCH(OH)CH2CHCHCH3 only in the Z-configuration. This appears to be the first example of total “cis-preference” in the addition of Grignard-like reagents to carbonyl compounds. The results are discussed in terms of steric requirements around the tin centre which is probably five-coordinate in the transition state.

Aerial powdering of bees inside mobile cages and the extent of neonicotinoid cloud surrounding corn drillers

Sudden losses of bees have been observed in spring during maize sowing. The death of bees has been correlated with the use of neonicotinoid‐coated seed and the toxic particulates emitted by pneumatic drilling machines. The contamination of foragers in flight over the ploughed fields has been hypothesized. The airborne contamination has been proven, both with bees inside fixed cages around the field and in free flight near the driller. A new trial involving mobile cages has been established and consists of making rapid passes with single bees inside cages fixed to an aluminium bar. The bar was moved by two operators at different distances from the working drilling machine. A single pass was shown as sufficient to kill all the bees exposed to exhaust air on the emission side of the drill, when bees were subsequently held in high relative humidity. The extent of toxic cloud around driller was evaluated at the height of 0.5, 1.8 and 3.5 m and proved to be about 20 m in diameter, with an ellipsoidal shape. The shape may be influenced by working speed of the drill and environmental parameters, and is easily shown by adding talc powder to the seed in the machine hopper. A new driller equipment was evaluated consisting of two tubes inclined towards the soil that direct the exhaust air towards the ground. The survival rate of the bees was not substantially increased using the modified drill and was lower than 50%. Chemical analyses show up to 4000 ng of insecticide in single bees with an average content around 300 ng. Similar quantities were observed at increased distances from the modified or unmodified drillers. This new evaluation of bee mortality in the field is an innovative biological test to verify the hypothetical efficiency (or not) of driller modifications.

Allylstannation : I. stereochemistry of the addition of trans/cis-2-butenyl-chloro-di-n-butyltin to aldehydes

2-Butenyl-chloro-di-n-butyltin, in various trans/cis ratios, reacts readily with neat RCHO (R = CH3, C2H5, C2H5(CH3)CH, (CH3)2CH, C6H5) at 25°C to give mixtures of threo/erythro-α-methylallylcarbinols in high yields. The same mixtures are obtained from the equilibrated mixtures obtained by redistribution between Bu3SnC4H7 (C4H7 = trans-, cis-crotyl and α-methylallyl group) and Bu2SnCl2. The reactions are characterized by a high degree of stereoselectivity, especially when bulky R groups are present. The complete allylic rearrangement and the stereoselectivity indicate that an exacyclic transition state is involved. Two stereochemically different transition states lead to two diastereoisomers, threo- and ertythro-α-methylallylcarbinol in the enantioforms RS, SR and RR, SS, respectively.

Reversible crotylstannation of carbonyl compounds. Crotylstannation ability of Bu3-nClnSnCH2CHCHCH3 (n = 0, 1, 2) compounds towards acetone and benzaldehyde and 13C NMR characterization

Abstract The crotylstannation reaction: has been found to be reversible. The compounds trans/cis- Bu 3- n Cl n SnCH 2 CH= CHCH 3 ( n = 0, 1, 2) have been prepared by elimination reactions of organostannoxy compounds,Bu 3− n Cl n Sn—O—C(Me)(i-Pr)CH(Me)CHCH2, which were synthesized by means of transalkoxylation between Bu 3- n CI n Sn(OMe) compounds and threo/erithro -2,3,4-trimethyl-5-hexen-3-ol. Under the conditions used the elimination occurs stereospecifically and with complete allylic rearrangement. The ability of the organostannoxy compounds to yield crotyl butylchlorotins via elimination increases in the order, Bu 3 Sn—O—C 2 ClSn—O—C 2 Sn—O—C. In the addition reactions, the sequence of increasing reactivity is Bu 3 SnCrot 2 SnCrot 2 BuSnCrot (Crot = crotyl). The 13 C NMR spectra of the compounds made reveal that the chemical shifts of the allylic carbon atoms are related to the inductive effects of the chloro-substituents.

A proposal for a new mechanism of interaction of trialkyltin (TAT) compounds with mitochondria

The interactions of trialkyltin (TAT) compounds with the mitochondria were largely studied. The current view of this phenomenon is that these compounds, by exploiting the Cl− and OH− gradient in energised mitochondria, behave as electroneutral OH−/Cl− exchangers. Experiments performed with triethyltin-, tripropyltin-, and tributyltin-chloride, allow us to propose an alternative mechanism. The crucial point of this new mechanism is that TATs enter the mitochondria as lipophilic cations (alkyl)3Sn+ and not as electroneutral compounds. The influx is followed by extrusion of the trialkyltin compounds as electroneutral hydroxi compounds (alkyl)3SnOH. Measurements of membrane potential show that the uptake of the TAT cation (alkyl)3Sn+ is followed by membrane depolarisation measured as the release of the trimethylphenylphoshonium probe. The depolarization occurs in the order: (Et)3SnCl < (Pro)3SnCl < (Bu)3SnCl in agreement with the increasing lipophilicity of the tested compounds from ethyltin toward butylin derivative. The uptake of TAT induces the opening of a selective anionic channel which could explain the swelling of mitochondria observed in a chloride medium.

Hydrated σ-bonded organometallic cations in organic synthesis: I. Allyl-, crotyl-, 1-methylallyl-, cyclohex-2-enyl-, and cinnamyl-stannation of carbonyl compounds in water

Homoallylic alcohols can be prepared in water by allyl-, crotyl-, 1-methylallyl-, cyclohex-2-enyl-, or cinnamyl-stannation of such carbonyl compounds as aldehydes, dialdehydes, and ketones, as well as acetals. The procedure is based on: Bu2RSnCl + R′COR″ + 12H2O → R(HO)CR′R″ + 12(Bu2SnCl)2O ↓ where R = allyl, crotyl, 1-methylallyl, cyclohex-2-enyl, or cinnamyl group, R′ = H or alkyl group, R″ ≠ R′ = alkyl group. In most cases, the reaction products are obtained rapidly in high yields (80–100%). Hydrated organometallic cations Bu2RSn+(aq) are partly involved. These results, together with those already published on 2-propynyl- and allenyl-stannation, indicate the value of this procedure.

Allylstannation: VI. Allylation and allenylation of aldehydes and ketones by allyl- and allenyl-tin chlorides in the presence of water

Abstract Allyl- and propargyl-carbinols can be readily prepared in one pot reaction, in the presence of water, by addition of allyl- or allenyl-tin chlorides to carbonyl compounds. The reactions of Bu3-n(CH2CHCH2)SnCln, Bu3-n(CH3CHCHCH2)SnCln (n = 1, 2) and Bu2(CH2CCH)SnCl and the carbonyl compounds RCHO (R = H, C2H5, (CH3)2CH, (CH3)3C, (E)-CH3CHCH, C6H5) and RCOR′ (R = R′ = CH3 and R = CH3 and R′ = (CH3)2CH) have been examined. The monochloroorganotin derivatives undergo the following quantitatvive reactions: Bu2(CH2CHCH2)SnCl + RCOR′ + 1 2 H2O → R(HO)C(R′)CH2CHCH2 + 1 2 [Bu2SnCl]2O↓ (1) Bu2(CH2CHCH2)SnCl + RCOR′ + 1 2 H2O → R(HO)C(R′)CH2CHCH + 1 2 [Bu2SnCl]2O↓ (2) Reaction (2), which involves the allenic substrate, is characterized by rearrangement of the unsaturated chain, the propargylic carbinol being isolated as the major product (∼ 90%).

UHPLC-DAD method for the determination of neonicotinoid insecticides in single bees and its relevance in honeybee colony loss investigations.

In the understanding of colony loss phenomena, a worldwide crisis of honeybee colonies which has serious consequences for both apiculture and bee-pollination-dependent farm production, analytical chemistry can play an important role. For instance, rapid and accurate analytical procedures are currently required to better assess the effects of neonicotinoid insecticides on honeybee health. Since their introduction in agriculture, neonicotinoid insecticides have been blamed for being highly toxic to honeybees, possibly at the nanogram per bee level or lower. As a consequence, most of the analytical methods recently optimized have focused on the analysis of ultratraces of neonicotinoids using liquid chromatography–mass spectrometry techniques to study the effects of sublethal doses. However, recent evidences on two novel routes—seedling guttations and seed coating particulate, both associated with corn crops—that may expose honeybees to huge amounts of neonicotinoids in the field, with instantly lethal effects, suggest that selected procedures need optimizing. In the present work, a simplified ultra-high-performance liquid chromatography–diode-array detection method for the determination of neonicotinoids in single bees has been optimized and validated. The method ensures good selectivity, good accuracy, and adequate detection limits, which make it suitable for the purpose, while maintaining its ability to evaluate exposure variability of individual bees. It has been successfully applied to the analysis of bees in free flight over an experimental sowing field, with the bees therefore being exposed to seed coating particulate released by the pneumatic drilling machine.