Assessing toxicity of pesticide inert ingredients and spray adjuvant principal functioning agents to honey bees (Apis mellifera)

Authors: SHANNON, BRANDON - The Ohio State University; TARVER, LAUREN - The Ohio State University; Jeon, Hongyoung; JOHNSON, REED - The Ohio State University

Submitted to: Environmental Toxicology and Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/6/2025
Publication Date: 11/24/2025
Citation: Shannon, B., Tarver, L., Jeon, H., Johnson, R. 2025. Assessing toxicity of pesticide inert ingredients and spray adjuvant principal functioning agents to honey bees (Apis mellifera). Environmental Toxicology and Chemistry. 45(2):363-375. https://doi.org/10.1093/etojnl/vgaf283.
DOI: https://doi.org/10.1093/etojnl/vgaf283

Interpretive Summary: Honeybees pollinate over 100 crops in the US including a sizable portion of California almond trees. During this process, they are exposed to adjuvants, often labeled as "inert ingredients" in pesticide formulations or added separately to enhance pesticide application. Despite their widespread use, the adjuvant toxicity to honeybees is poorly understood, with limited data for adjuvant products used in almond orchards, even though evidence suggests they pose risks. This study investigated the 48-hour acute toxicity (LC50) of thirty-seven pesticide inert ingredients or adjuvant principal functioning agents using a Potter Spray Tower. Structural and physical properties of fifteen alcohol ethoxylates such as moles of ethoxylation, carbon chain length in the alcohol group, and Hydrophilic-Lipophilic Balance (HLB) were analyzed to determine the properties correlated with the bee toxicity. Additionally, a silicone-based adjuvant (Silwet Eco®) and a non-ionic surfactant (Makon 10®) were assessed for their toxicity on different body parts of adult honeybees. The results revealed that about 68% of the tested inert ingredients showed a significant dose-response within 48 hours. Approximately 41% of adjuvants had notable toxicity with LC50 below the maximum tested concentration. For alcohol ethoxylates, only HLB showed a significant correlation with toxicity levels. The body placement assay results highlighted that applying adjuvants to the dorsal head or ventral thorax generally caused the highest toxicity, though the two model adjuvants produced varying results. These findings underline the potential risks of adjuvants pose to honeybees and highlight the need for better toxicity data to guide safer choices in formulating pesticides and using adjuvants in agricultural pesticide applications for crops dependent on bee pollination.

Technical Abstract: Commercial beekeepers transport colonies across the U.S. to provide pollination services for over 100 different crops, especially for almond production in California. In these agricultural settings, honey bees are exposed to adjuvant compounds that are included as “inert ingredients” in pesticide formulations or are added as a separate adjuvant product to pesticides to improve application characteristics. Only 22 of the 459 unique adjuvant products that were applied on almond orchards during bloom from 2011-2021 have published bee toxicity data, despite evidence that they pose risks to bees. This study used a Potter Spray Tower to determine the 48-hour acute toxicities (LC50) of 37 different pesticide inert ingredients or adjuvant principal functioning agents. Additionally, toxicity trends of 15 alcohol ethoxylates were determined for structural and physical metrics, including moles of ethoxylation, number of carbons in the alcohol group, and Hydrophilic-Lipophilic Balance (HLB). Lastly, LD50 values of a model organo-silicone (Silwet Eco®) and a model non-ionic surfactant (Makon 10®) were determined when applied to different parts of the adult honey bee. Results demonstrate that 25 of the 37 inert ingredients tested demonstrated a significant 48-hour dose-response, and 15 of the 25 inert ingredients with a significant dose-response demonstrated an LC50 below the maximum concentration tested. For alcohol ethoxylates, moles of ethoxylation and number of carbons in the alcohol group were not significant predictors for toxicity, but HLB did demonstrate a significant trend with toxicity. The body placement assay indicated that application to the dorsal part of the head and the ventral thorax were generally the most toxic, but that the two adjuvants produced differing results. The findings from this study can inform choices of adjuvant and pesticide formulation components to reduce risks to honey bees.