Revisiting chlorophyll extraction methods in biological soil crusts – methodology for determination of chlorophyll <i>a</i> and chlorophyll <i>a</i> + <i>b</i> as compared to previous methods

Abstract

Abstract. Chlorophyll concentrations of biological soil crust (biocrust) samples are commonly determined to quantify the relevance of photosynthetically active organisms within these surface soil communities. Whereas chlorophyll extraction methods for freshwater algae and leaf tissues of vascular plants are well established, there is still some uncertainty regarding the optimal extraction method for biocrusts, where organism composition is highly variable and samples comprise major amounts of soil. In this study we analyzed the efficiency of two different chlorophyll extraction solvents, the effect of grinding the soil samples prior to the extraction procedure, and the impact of shaking as an intermediate step during extraction. The analyses were conducted on four different types of biocrusts. Our results show that for all biocrust types chlorophyll contents obtained with ethanol were significantly lower than those obtained using dimethyl sulfoxide (DMSO) as a solvent. Grinding of biocrust samples prior to analysis caused a highly significant decrease in chlorophyll content for green algal lichen- and cyanolichen-dominated biocrusts, and a tendency towards lower values for moss- and algae-dominated biocrusts. Shaking of the samples after each extraction step had a significant positive effect on the chlorophyll content of green algal lichen- and cyanolichen-dominated biocrusts. Based on our results we confirm a DMSO-based chlorophyll extraction method without grinding pretreatment and suggest the addition of an intermediate shaking step for complete chlorophyll extraction (see Supplement S6 for detailed manual). Determination of a universal chlorophyll extraction method for biocrusts is essential for the inter-comparability of publications conducted across all continents.

Spectral Response Analysis: An Indirect and Non-Destructive Methodology for the Chlorophyll Quantification of Biocrusts

“…Chla provides valuable information about the photosynthetic capacity of biocrusts [27] and have been widely used as a surrogate for biocrust development and growth of both natural and artificially induced biocrust communities [15,28,58,70,71]. However, until now, most analyses were based on punctual measurements and required destructive sampling that hindered the analysis of seasonal dynamics and growth rates in these spatially heterogeneous communities during long term monitoring experiments.…”

“…Moreover, it is known that chlorophyll can be used to assess biocrust states in response to seasonal dynamics [17,18], anthropogenic disturbances [19,20], ongoing climate changes [21,22], and restoration activities [23,24]. For all these reasons, large efforts have been devoted during the past decades to obtain reliable measurements of biocrust chlorophyll concentrations [25][26][27]. In a similar way to wet chemical methods used for the extraction of chlorophyll a from plant tissue, most of these techniques require extraction in a solvent followed by spectrophotometric determinations of the chlorophyll-related absorbance peaks in the supernatant, and then conversion from absorbance to concentrations using standard published equations [25][26][27].…”

“…For all these reasons, large efforts have been devoted during the past decades to obtain reliable measurements of biocrust chlorophyll concentrations [25][26][27]. In a similar way to wet chemical methods used for the extraction of chlorophyll a from plant tissue, most of these techniques require extraction in a solvent followed by spectrophotometric determinations of the chlorophyll-related absorbance peaks in the supernatant, and then conversion from absorbance to concentrations using standard published equations [25][26][27]. Though accurate, these laboratory methods require destructive field sampling and are expensive and time consuming.…”

Effect of phosphate availability on biofilm formation in cooling towers

“…Quantification of chlorophyll a and b (chlorophyll a þ b) was conducted following the proposed protocol from Caesar et al (2018). In brief, each basin biofilm sample was dried at 60 C for 24 h before analysis.…”

“…The supernatants from the first and second extractions were combined and centrifuged for 10 min at 3,000 Â g. The absorbance was measured using a UV-Vis spectrophotometer (Lambda 45, Perkin-Elmer) at 648, 665 and 700 nm. The equations for the calculation of the amount of chlorophyll a and b are described elsewhere (Caesar et al 2018).…”