Continuous hydrogen peroxide production by organic buffers in phytoplankton culture media

Abstract

We investigated the production of hydrogen peroxide (HOOH) in illuminated seawater media containing a variety of zwitterionic buffers. Production rates varied extensively among buffers, with 4-(2-hydroxyethyl)1-piperazineethanesulfonic acid (HEPES) highest and N-Tris(hydroxymethyl)methyl-3-aminopropanesulfonic acid (TAPS) among the lowest. The rate of HOOH accumulation was remarkably consistent over many days, and increased linearly with buffer concentration, natural seawater concentration, and light level. Concentrations of HEPES commonly used in culture media (1-10 mM) generated enough HOOH to kill the axenic Prochlorococcus strain VOL1 during growth in enriched seawater media at lower, environmentally realistic cell concentrations and/or under high light exposure. We also demonstrated that HEPES can be used experimentally to study the biological effects of chronic exposure to sublethal levels of HOOH such as may be experienced by light-exposed microorganisms.

Mimicking lichens: incorporation of yeast strains together with sucrose-secreting cyanobacteria improves survival, growth, ROS removal, and lipid production in a stable mutualistic co-culture production platform

“…Further, chemicals present in cultural systems are known to produce HOOH [40]. Previously, HEPES with 1-10 mM concentration in culture could generate enough HOOH to kill the axenic phytoplankter Prochlorococcus strain [40].…”

Flaming as part of aseptic technique increases CO2 (g) and decreases pH in freshwater culture media

“…4) may be due to the variability of CO 2 incorporation from flaming, as some cultures may have experienced larger-scale CO 2 inputs that resulted in pH decline if the CO 2 is not consumed by the culture. Previous studies have also found that up to 2 μmol L −1 of HOOH may be generated as a result of flaming (Morris and Zinser 2013). While it is recognized that mmol L −1 concentrations of HOOH are needed to cause cell death (Palenik et al 1991;Alam et al 2001), the 2 μmol L −1 of HOOH generated from flaming results in physiological changes during the upregulation of intracellular peroxidases (Price and Harrison 1988).…”

“…It has been demonstrated that light exposure causes photooxidation (and the inherent generation of reactive oxygen species, such as the weak acid HOOH) within a variety of media utilizing buffers such as HEPES, TAPS, Bicine, and TRIS (Morris and Zinser 2013). To determine the effects of photooxidation on media pH, 25 mL volumes of CT media were aliquoted into 50-mL glass culture tubes in triplicate for air-flame treatments.…”

“…S6). However, the approach of simply increasing buffer concentrations must be pursued with caution: for example, TRIS is toxic to cells in high concentrations and HEPES generates higher amounts of reactive oxygen species when present in higher concentrations (Morris and Zinser 2013).…”

Heterotrophic Bacteria Exhibit a Wide Range of Rates of Extracellular Production and Decay of Hydrogen Peroxide

“…As discussed for decay above, components originally present in the growth medium could have contributed to the production rates measured in the two supernatant experiments. Morris and Zinser (2013) showed that H 2 O 2 production could occur by exposing zwitterionic buffers such as those used in growth media to light. However, the bacteria used in this experiment were grown in a dark incubator, and syringes used for spiked batch incubations were covered between time points.…”