Oklahoma Fish Kill Study: Looking for a Toxic Needle in an Environmental Haystack

Overview
Purpose
To determine unknown contaminants in water samples during an active fish kill.

Methods
A combination of solid-phase extraction (SPE), LC-ion trap-MS/MS and high resolution LC-MS.

Results
An unknown contaminant was uniquely identified as chlorin-e6-trimethyl ester, using both LC-ion trap-MS/MS and high resolution LC-MS.

Introduction
On July 9, 2011, a major fish kill (fish kill I) was observed by the Oklahoma Department of Environmental Quality (OKDEQ) in the Red River, near Ketchum’s Bluff, Oklahoma. The Red River, with headwaters in the Texas panhandle, flows for 917 kilometers, between the borders of Oklahoma (OK) and Texas (TX), before emptying into the Mississippi River. During this fish kill, hundreds of large bottom feeder fish (i.e., catfish and buffalo) were observed as either dead, struggling, or actively dying. Nearly two months later, on September 14, 2011, another fish kill (fish kill II) occurred further south along the Red River, approximately 130 km downstream from Ketchum’s Bluff near Lake Texhoma. Again, it was observed that hundreds of only the large bottom feeder fish were affected by an unknown toxin(s). OKDEQ believed that the two fish kills were related, with the unknown toxicant(s) traveling further downstream from the first fish kill (July 9, 2011), but causing fish mortality 60 days later downstream. The following year, on June 13, 2012, another fish kill (fish kill III) occurred, again near the area of Ketchum’s Bluff and Red Creek confluence. And a final fish kill (fish kill IV) occurred on January 31, 2013, in the same watershed, near Red River and Beaver Creek confluence. Environmental samples (i.e., water, sediment, and fish) were collected, by OKDEQ and the United States Envi...

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...undinacea”, Phytochemistry Letters 4(2):79-85

Fig 1. Chlorin-e6-trimethyl ester
Figure 2a. CID MS/MS LC-ITMS: Chlorin-e6-trimethyl ester standard, m/z 639.3 (M+H)+
2b. CID MS/MS LC-ITMS Unknown m/z 639.3 (M+H)+
Fig. 3 Likely ammonolysis transformation product (SPE artifact)
Fig. 4 Relationship of mass m/z 639 Da vs timeline of fish kill
Acknowledgements: Slides of dead fish courtesy of OKDEQ. We would like to thank our students Trevor Nance Jr, and Matt Ward for their help in the laboratory sample preparations. We would also like to thank OKDEQ (Chris Armstrong) and EPA Region 6 (Rick McMillin) for their patience.
Notice: Although this work was reviewed and by EPA and approved for publication, it may not necessarily reflect official Agency policy. Mention of trade names or commercial products does not constitute endorsement or recommendation by EPA for use.