Environmental Science and Pollution Research

, Volume 22, Issue 11, pp 8010–8021 | Cite as

Effects of realistic doses of atrazine, metolachlor, and glyphosate on lipid peroxidation and diet-derived antioxidants in caged honey bees (Apis mellifera)

  • Stephanie Hedrei Helmer
  • Anahi Kerbaol
  • Philippe Aras
  • Catherine Jumarie
  • Monique Boily
Crop protection: environment, human health, and biodiversity


The decline in the population of pollinators is a worrying phenomenon worldwide. In North America, the extensive use of herbicides in maize and soya crops may affect the health of nontarget organisms like the honey bee. In this study, caged honey bees were exposed to realistic doses of atrazine, metolachlor, and glyphosate for 10 days via contaminated syrup. Peroxidation of lipids was evaluated using the thiobarbituric acid reactive substance (TBARS) test, and diet-derived antioxidants—carotenoids, all-trans-retinol (at-ROH) and α-tocopherol—were detected and quantified using reversed-phase HPLC techniques. Significant increases in syrup consumption were observed in honey bees exposed to metolachlor, and a lower TBARS value was recorded for the highest dose. No relationship was observed between the peroxidation of lipids and the levels of antioxidants. However, β-carotene, which was found to be the most abundant carotenoid, and at-ROH (derived from β-carotene) both decreased with increasing doses of atrazine and glyphosate. In contrast, metolachlor increased levels of at-ROH without any effects on β-carotene. These results show that the honey bee carotenoid–retinoid system may be altered by sublethal field-realistic doses of herbicides.


Apis mellifera Carotenoids All-trans-retinol α-Tocopherol TBARS 



The authors would like to thank Maxime Gauthier for HPLC technical assistance. We also thank Dr. Diana Averill for generous advices in TBARS analysis. We are grateful to Dr. Philip Spear for providing access to his laboratory and TOXEN (Centre de recherche en toxicologie de l’environnement) for the use of analytical equipment. This study was supported by the Programme de soutien à l’innovation en agroalimentaire (PSIA) from Ministère de l’Agriculture, des Pêcheries et de l’Alimentation du Québec (MAPAQ), attributed to M. Boily (#811175).

Supplementary material

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ESM 1 (PDF 50 kb)


  1. Álvarez R, Vaz B, Gronemeyer H, de Lera AR (2013) Functions, therapeutic applications, and synthesis of retinoids and carotenoids. Chem Rev. doi: 10.1021/cr400126u Google Scholar
  2. ARLA (Agence de réglementation de la lutte antiparasitaire) (2007) Décision de réévaluation: atrazine (Évaluation environnementale). Santé Canada, Sécurité des produits de consommation, RVD2007-05Google Scholar
  3. Arshavsky VY (2009) Vision: the retinoid cycle in Drosophila. Curr Biol R96–8. doi: 10.1016/j.cub.2009.12.039
  4. Bahadorani S, Bahadorani P, Phillips JP, Hilliker AJ (2008) The effects of vitamin supplementation on Drosophila life span under normoxia and under oxidative stress. J Gerontol A Biol Sci Med Sci 63:35–42CrossRefGoogle Scholar
  5. Berry JA, Hood WM, Pietravalle S, Delaplane KS (2013) Field-level sublethal effects of approved bee hive chemicals on honey bees (Apis mellifera L). PLoS ONE 8(10):e76536. doi: 10.1371/journal.pone.0076536 CrossRefGoogle Scholar
  6. Boily M, Thibodeau J, Bisson M (2009) Retinoid metabolism (LRAT, REH) in the liver and plasma retinoids of bullfrog, Rana catesbeiana, in relation to agricultural contamination. Aquat Toxicol 91:118–125CrossRefGoogle Scholar
  7. Boily M, Sarrasin B, DeBlois C, Aras P, Chagnon M (2013) Acetylcholinesterase in honey bees (Apis mellifera) exposed to neonicotinoids, atrazine and glyphosate: laboratory and field experiments. Environ Sci Pollut Res 20:5603–5614CrossRefGoogle Scholar
  8. Bonnefont D, Legrand A, Peynet J, Emerit J, Delattre J, Galli A (1989) Distribution of thiobarbituric acid-reactive substances in lipoproteins and proteins in serum. Clin Chem 35:2054–2058Google Scholar
  9. Boucher C (2013) Rapport sur les mortalités de colonies d’abeilles observées à la suite de l’hivernage de 2012-2013. L’abeille 35:8–10Google Scholar
  10. Bradford MM (1976) A rapid and sensitive method for quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254CrossRefGoogle Scholar
  11. Casani R (2000) Vitamin E. Kirk-Othmer encyclopedia of chemical technology. doi: 10.1002/0471238961.2209200103011901.a01
  12. Chantarudee A, Phuwapraisirisan P, Kimura K, Okuyama M, Mori H, Kimura A, Chanchao C (2012) Chemical constituents and free radical scavenging activity of corn pollen collected from Apis mellifera hives compared to floral corn pollen of Nan, Thailand. BMC Complement Alter Med 12:45CrossRefGoogle Scholar
  13. Di Pasquale G, Salignon M, Le Conte Y, Belzunces LP, Decourtye A et al (2013) Influence of pollen nutrition on honey bee health: do pollen quality and diversity matter? PLoS ONE 8(8):e72016. doi: 10.1371/journal.pone.0072016 CrossRefGoogle Scholar
  14. Dobšíková R, Blahová J, Modrá H, Škorič M, Svobodová Z (2011) The effect of acute exposure to herbicide Gardoprim Plus Gold 500 SC on haematological and biochemical indicators and histopathological changes in common carp (Cyprinus carpio L.). Acta Vet BRNO 80:359–363CrossRefGoogle Scholar
  15. Farjan M, Dmitryjuk M, Lipiński Z, Biernat-Łopieńska E, Żółtowska K (2012) Supplementation of the honey bee diet with vitamin C: the effect on the antioxidative system of Apis mellifera carnica brood at different stages. J Api Res 51:263–270CrossRefGoogle Scholar
  16. García-Plazaola JI, Becerril JM (1999) A rapid high-performance liquid chromatography method to measure lipophilic antioxidants in stressed plants: simultaneous determination of carotenoids and tocopherols. Phytochem Anal 10:307–313CrossRefGoogle Scholar
  17. George DGM, Gatehouse AMR (2013) Oxidative stress enzymes in Busseola fusca. Int J Curr Microbiol App Sci 2:485–495Google Scholar
  18. Giovannucci DR, Stephenson RS (1999) Identification and distribution of dietary precursors of the Drosophila visual pigment chromophore: analysis of carotenoids in wild type and ninaD mutants by HPLC. Vis Res 39:219–229CrossRefGoogle Scholar
  19. Gorse I, Rivard L (2011) Bilan des ventes de pesticides au Québec pour l’année 2008, Québec, Ministère du Développement durable, de l’Environnement et des Parcs, ISBN (PDF) 978-2-550-61586-6, 85 pGoogle Scholar
  20. Grotto D, Santa Maria L, Valentini C, Paniz C, Schmitt G et al (2009) Importance of the lipid peroxidation biomarkers and methodological aspects for malondialdehyde quantification. Quim Nova 32:169–174CrossRefGoogle Scholar
  21. Hahn DA, Denlinger DL (2007) Meeting the energetic demands of insect diapause: nutrient storage and utilization. J Insect Physiol 53:760–773CrossRefGoogle Scholar
  22. Halme A, Cheng M, Hariharan IK (2010) Retinoids regulate a developmental checkpoint for tissue regeneration in Drosophila. Curr Biol 20:458–463CrossRefGoogle Scholar
  23. Hanganu D, Olah N, Vlase L, Marculedscu A, Pintea A (2012) Chemical research of carotenoids from Chenopodium bonus henricus L. (Chenopodiaceae). Farm 60:840–849Google Scholar
  24. Horváth G, Molnar P, Farkas A, Szabo LG, Turcsi E, Deli J (2010) Separation and identification of carotenoids in flowers of Chelidonium majus L. and inflorescences of Solidago canadensis L. Chromatographia 71:S103–S108CrossRefGoogle Scholar
  25. Hsu CY, Hsieh YS (2013) Oxidative stress decreases in the trophocytes and fat cells of worker honeybees during aging. Biogerontology. doi: 10.1007/s10522-013-9485-9 Google Scholar
  26. ISQ (Institut de la statistique du Québec), MAPAQ (Ministère de l’Agriculture, des Pêcheries et de l'Alimentation du Québec) (2013) Profil sectoriel de l’industrie bioalimentaire du Québec, édition 2013. Gouvernement du Québec, ISBN 978-2-550-70028-9 (PDF)Google Scholar
  27. Jasper R, Locatelli GO, Pilati C, Locatelli C (2012) Evaluation of biochemical, hematological and oxidative parameters in mice exposed to the herbicide glyphosate-Roundup®. Interdiscip Toxicol 5:133–140CrossRefGoogle Scholar
  28. Johnson RM, Dahlgren L, Siegfried BD, Ellis MD (2013) Acaricide, fungicide and drug interactions in honey bees (Apis mellifera). PLoS ONE 8(1):e54092. doi: 10.1371/journal.pone.0054092 CrossRefGoogle Scholar
  29. Johnson RM, Ellis MD, Mullin CA, Frazier M (2010) Pesticides and honey bee toxicity. Apidologie 41:312–332CrossRefGoogle Scholar
  30. Kane MA, Folias AE, Napoli JL (2008) HPLC/UV quantification of retinal, retinol, and retinyl esters in serum and tissues. Anal Biochem 378:71–79CrossRefGoogle Scholar
  31. Kayser H (1982) Carotenoids in insects. In: Britton G, Goodwin TW (eds) Carotenoid chemistry and biochemistry. Pergamon Press, Toronto, pp 195–210CrossRefGoogle Scholar
  32. Krupke CH, Hunt GJ, Eitzer BD, Andino G, Given K (2012) Multiple routes of pesticide exposure for honey bees living near agricultural fields. PLoS One 7:e29268. doi: 10.1371/journal.pone.0029268 CrossRefGoogle Scholar
  33. Lenkowski JR, McLaughlin KA (2010) Acute atrazine exposure disrupts matrix metalloproteinases and retinoid signaling during organ morphogenesis in Xenopus laevis. J Appl Toxicol 30:582–589CrossRefGoogle Scholar
  34. Mann RM, Hyne RV, Choung CB, Wilson SP (2009) Amphibians and agricultural chemicals: review of the risks in a complex environment. Environ Pollut 157:2903–2927CrossRefGoogle Scholar
  35. Maini S, Medrzycki P, Porrini C (2010) The puzzle of honey bee losses: a brief review. Bull Insectol 63:153–160Google Scholar
  36. Mueller L, Boehm V (2011) Antioxidant activity of β-carotene compounds in different in vitro assays. Molecules 16:1055–1069. doi: 10.3390/molecules16021055 CrossRefGoogle Scholar
  37. Mullin CA, Frazier M, Frazier JL, Ashcraft S, Simonds R, Vanengelsdorp D, Pettis JS (2010) High levels of miticides and agrochemicals in North American apiaries: implications for honey bee health. PLoS ONE 5(3):e9754. doi: 10.1371/journal.pone.0009754 CrossRefGoogle Scholar
  38. Murvoll KM, Skaare JU, Jensen H, Jenssen BM (2007) Associations between persistent organic pollutants and vitamin status in Brünnich’s guillemot and common eider hatchlings. Sci Total Environ 381:134–145CrossRefGoogle Scholar
  39. Nakamuraa A, Stiebler R, Fantappi MR, Fialho E, Masudaa H, Oliveira MF (2007) Effects of retinoids and juvenoids on moult and on phenoloxidase activity in the blood-sucking insect Rhodnius prolixus. Acta Trop 103:222–230CrossRefGoogle Scholar
  40. Ndayibagira A, Spear PA (1999) Esterification and hydrolysis of vitamin A in liver of brook trout (Salvelinus fontinalis) and the influence of a coplanar polychlorinated biphenyl. Comp Biochem Physiol C 122:317–325Google Scholar
  41. Nwani CD, Lakra WS, Nagpure NS, Kumar R, Kushwaha B, Srivastava SK (2010) Toxicity of the herbicide atrazine: effects on lipid peroxidation and activities of antioxidant enzymes in the freshwater fish Channa Punctatus (Bloch). Int J Environ Res Public Health 7:3298–3312CrossRefGoogle Scholar
  42. Ohkawa H, Ohishi N, Tagi K (1979) Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal Chem 95:351–358Google Scholar
  43. Otis GW, Wheeler DE, Buck N, Mattila HR (2004) Storage proteins in winter honey bees. Apiacata 38:352–357Google Scholar
  44. Paganelli A, Gnazzo V, Acosta H, López SL, Carrasco AE (2010) Glyphosate-based herbicides produce teratogenic effects on vertebrates by impairing retinoic acid signaling. Chem Res Toxicol 23:1586–1595CrossRefGoogle Scholar
  45. Potts SG, Roberts SPM, Dean R, Marris G, Brown MA, Jones R, Neumann P, Settele J (2010) Declines of managed honey bees and beekeepers in Europe. J Api Res 49:15–22CrossRefGoogle Scholar
  46. Rael LT, Thomas GW, Craun ML, Curtis CG, Bar-Or R, Bar-Or D (2004) Lipid peroxidation and the thiobarbituric acid assay: standardization of the assay when using saturated and unsaturated fatty acids. J Biochem Mol Biol 37:749–752CrossRefGoogle Scholar
  47. Rehman S, Rehman S, Waliullah MIS (2012) Chlorpyrifos-induced neuro-oxidative damage in bee. Toxicol Environ Health Sci 4:30–36. doi: 10.1007/s13530-012-0114-9 CrossRefGoogle Scholar
  48. Rousse P, Gourdon F, Roubaud M, Chiroleu F, Quilici S (2009) Biotic and abiotic factors affecting the flight activity of Fopius arisanus, an egg-pupal parasitoid of fruit fly pests. Environ Entomol 38:896–903CrossRefGoogle Scholar
  49. Shete V, Quadro L (2013) Mammalian metabolism of β-Carotene: gaps in knowledge. Nutrients 5:4849–4868. doi: 10.3390/nu5124849 CrossRefGoogle Scholar
  50. Singh M, Sandhir R, Kiron R (2010) Alterations in Ca2+ homeostasis in rat erythrocytes with atrazine treatment: positive modulation by vitamin E. Mol Cell Biochem 340:231–238CrossRefGoogle Scholar
  51. Široká Z, Drastichová J (2004) Biochemical markers of aquatic environment contamination-cytochrome P450 in fish. A review. Acta Vet Brno 73:123–132Google Scholar
  52. Sleeman JM, Brown J, Steffen D, Jones D, Roberston J, Holladay S (2008) Relationships among aural abscesses, organochlorine compounds, and vitamin A in free-ranging eastern Box turtles (Terrapene Carolina carolina). J Wild Dis 44:922–929CrossRefGoogle Scholar
  53. Smith WC, Goldsmith TH (1991) The role of retinal photoisomerase in the visual cycle of the honeybee. J Gen Physiol 97:143–165CrossRefGoogle Scholar
  54. Spear PA, Boily M, Giroux I, DeBlois C, Leclair MH, Levasseur M, Leclair R (2009) Study design, water quality, morphometrics and age of the bullfrog, Rana catesbeiana, in sub-watersheds of the Yamaska River drainage basin, Québec, Canada. Aquat Toxicol 91:110–117CrossRefGoogle Scholar
  55. Tapparo A, Giorio C, Marzaro M, Marton D, Sold L, Girolami V (2011) Rapid analysis of neonicotinoid insecticides in guttation drops of corn seedlings obtained from coated seeds. J Environ Monit 13:1564–1568CrossRefGoogle Scholar
  56. Thornton BJ, Elthon TE, Cerny RL, Siegfried BD (2010) Proteomic analysis of atrazine exposure in Drosophila melanogaster (Diptera: Drosophilidae). Chemosphere 81:235–241CrossRefGoogle Scholar
  57. von Lintig J (2012) Metabolism of carotenoids and retinoids related to vision. J Biol Chem 287:1627–1634. doi: 10.1074/jbc.R111.303990 CrossRefGoogle Scholar
  58. von Lintig J, Dreher A, Kiefer C, Wernet M, Vogt K (2001) Analysis of the blind Drosophila mutant ninaB identifies the gene encoding the key enzyme for vitamin A formation in vivo. PNAS 98:1130–1135. doi: 10.1073/pnas.031576398 Google Scholar
  59. Wang T, Jiao Y, Montell C (2007) Dissection of the pathway required for génération of vitamin A and for Drosophila phototransduction. J Cell Biol 177:305–316CrossRefGoogle Scholar
  60. Waris G, Ahsan H (2006) Reactive oxygen species: role in the development of cancer and various chronic conditions. J Carcinogen 5:14CrossRefGoogle Scholar
  61. Weirich GF, Collins AM, Williams VP (2002) Antioxidant enzymes in the honey bee, Apis mellifera. Apidologie 33:3–14CrossRefGoogle Scholar
  62. Wu JY, Anelli CM, Sheppard WS (2011) Sub-lethal effects of pesticide residues in brood comb on worker honey bee (Apis mellifera) development and longevity. PLoS ONE 6:e14720. doi: 10.1371/journal.pone.0014720 CrossRefGoogle Scholar
  63. Yang EC, Chuang YC, Chen YL, Chang LH (2008) Abnormal foraging behavior induced by sublethal dosage of imidacloprid in the honey bee (Hymenoptera: Apidae). J Econ Entomol 101:1743–1748CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Stephanie Hedrei Helmer
    • 1
  • Anahi Kerbaol
    • 2
  • Philippe Aras
    • 1
  • Catherine Jumarie
    • 1
  • Monique Boily
    • 1
  1. 1.Département des Sciences BiologiquesUniversité du Québec à MontréalMontréalCanada
  2. 2.Département des Sciences de l’EnvironnementUniversité du Québec à MontréalMontréalCanada

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