Assessment of the effects of storage temperature on fatty acid analysis using dried blood spot cards from managed southern white rhinoceroses (Ceratotherium simum simum): implications for field collection and nutritional care
Authors: Wood Jordan , Minter Larry J. , Bibus Doug , Tollefson Troy N. , Ange-van Heugten Kimberly 
Published: February 2022

BackgroundSouthern white rhinoceroses (Ceratotherium simum simum) are an endangered species in decline due to poaching and negative habitat changes. Conservation of the species has become increasingly important and a focus on better human management has become prevalent. One area of management that impacts southern white rhinoceroses is nutritional health monitoring, which is often conducted through blood analysis. Blood analysis conducted during field research can be difficult due to temperature, distance, and limited technological resources, so new methods of fast, and relatively stable blood collection are being pursued. One method that has been used in humans for many years is beginning to make its way into wildlife studies: the use of dried blood spot (DBS) cards. These cards are used as a tool to store single drops of whole blood on specialized filter paper and, once dried, can be used for nutritional biomarker analysis. An area of interest for southern white rhinoceroses and nutrition is monitoring fatty acid percentages for cardiovascular, immune, and reproductive health. The time and temperature limitations for storing blood fractions or liquid whole blood when analyzing fatty acids have been investigated, but few studies have performed storage studies on DBS cards colder than −20 °C or in non-human species.MethodsIn order to better understand the limitations of DBS cards and the impact of temperature on fatty acid DBS samples in long-term storage, triplicate samples from seven adult southern white rhinoceroses at the North Carolina Zoo were collected and subjected to three storage treatments (immediate, room temperature (23 °C), or frozen (−80 °C) for 1 year).ResultsStearidonic (18:4w3) (Δ 0.3%), arachdic (20:0) (Δ 0.1%), eicosatetraenoic (20:4w3) (Δ 0.2%), and erucic acid (22:1w9) (Δ 0.1%) were in higher concentration in frozen than initial. Fatty acids in higher concentrations in the initial samples than frozen were myristic (14:0) (Δ 0.2%), mead (20:3w9) (Δ 0.1%), docosatetraenoic (22:4w6) (Δ 0.2%), nervonic (24:1) (Δ 0.1%), and total highly unsaturated fatty acids (HUFAs) (Δ 0.7%). Stearic (18:0) (Δ 2.2%), stearidonic (18:4w3) (Δ 0.3%), arachdic (20:0) (Δ 0.2%), paullinic (20:1w7) (Δ 0.4%), eicosatetraenoic (20:4w3) (Δ 0.1%), eicosapentaenoic (20:5w3) (Δ 0.1%), docosatetraenoic (22:4w6) (Δ 0.2%), nervonic acid (24:1) (Δ 0.2%), monoenes (Δ 1.9%), and total saturates (Δ 3.6%) had higher concentrations in room temperature than initial. Linoleic (18:2w6) (Δ 4.9%), mead acid (20:3w9) (Δ 0.1%), total polyunsaturated fatty acids (5.3%), and total omega-6 fatty acids (Δ 4.8%) had higher concentrations in initial compared to room temperature. Arachidonic (20:4w6) (Δ 0.4%) and omega-3 docosapentaenoic acid (22:5w3) (Δ 0.1%), had higher concentrations in frozen than in room temperature.DiscussionThe frozen samples had the fewest statistical differences compared to room temperature samples and essential omega-3 and -6 fatty acids were stable with freezing up to 1 year. While more research is still warranted, current results suggest that DBS samples are best utilized when immediate analysis or −80 °C storage is available.