Translating PUFA omega 6:3 ratios from wild to captive hibernators (Urocitellus parryii) enhances sex-dependent mass-gain without increasing physiological stress indicators.

Omega 3 polyunsaturated fatty acids (PUFAs) are well-documented for their influence on health and weight loss. Recent studies indicate omega 3 PUFAs may exert a negative impact on cellular stress and physiology in some hibernators. We asked if physiological stress indicators, lipid peroxidation and mass gain in Arctic Ground Squirrels (AGS) were negatively influenced by naturally occurring dietary omega 3 PUFA levels compared to omega 3 PUFA levels found in common laboratory diets. We found plasma fatty acid profiles of free-ranging AGS to be high in omega 3 PUFAs with balanced omega 6:3 ratios, while standard laboratory diets and plasma of captive AGS are high in omega 6 and low in omega 3 PUFAs with higher omega 6:3 ratios. Subsequently, we designed a diet to mimick free-range AGS omega 6:3 ratios in captive AGS. Groups of wild-caught juvenile AGS were either fed: (1) Mazuri Rodent Chow (Standard Rodent chow, 4.95 omega 6:3 ratio), or (2) balanced omega 6:3 chow (Balanced Diet, 1.38 omega 6:3). AGS fed the Balanced Diet had plasma omega 6:3 ratios that mimicked plasma profiles of wild AGS. Balanced Diet increased female body mass before hibernation, but did not influence levels of cortisol in plasma or levels of the lipid peroxidation product 4-HNE in brown adipose tissue. Overall, as the mass gain is critical during pre-hibernation for obligate hibernators, the results show that mimicking a fatty acid profile of wild AGS facilitates sex-dependent mass accumulation without increasing stress indicators.

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.

BACKGROUND: Southern 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. METHODS: In 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). RESULTS: Stearidonic (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. DISCUSSION: The 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.

Implications of Nutritional Management on Fatty Acid Profiles of Southern White Rhinoceroses (Ceratotherium simum simum) Housed at Two Zoological Institutions.

Southern white rhinoceroses (Ceratotherium simum simum) are African megaherbivores that are considered near threatened by the International Union for the Conservation of Nature. The fatty acid circulating values of these animals have not been thoroughly investigated. Fatty acids are critical for immune, heart, skin, and reproductive health, and may have a significant impact on the management and conservation of this species. Published data on fatty acids in this species is limited to incomplete profiles with very few animals in managed environments. The objectives of this research were to provide novel fatty acid percentage profiles for managed healthy southern white rhinoceroses, as well as to provide comparisons between two zoological institutions with differences in diet and climate during two distinct pasture growth periods. Whole blood samples were collected as dried blood spots from six rhinoceroses at the North Carolina Zoo (NC Zoo) and five rhinoceroses at Busch Gardens Tampa (BGT) in the low growth period (February to April) of 2019 and during the high growth period (July to September) of 2020. Fatty acid results indicated numerous differences when comparing the institutions within the same growth period and when comparing the same institution between its two growth periods. Most noteworthy were the higher levels of α-linolenic acid (18:3w3) and total omega-3 fatty acids and the lower linoleic acid (18:2w6), total omega-6 fatty acids, and omega-6 to omega-3 ratio found in the BGT population in both growth periods. This study provides novel percentages of fatty acids in managed southern white rhinoceroses and data on how fatty acid profiles may be altered between two housing locations via dietary differences in hay type and quantity, pasture availability via season, and pellet inclusion levels.

Investigation of Dried Blood Spot Cards for Fatty Acid Analysis Using Porcine Blood.

Fatty acids, especially omega-3 and omega-6 fatty acids, are important for reproductive and cardiovascular health in animals. While monitoring fatty acids is traditionally conducted using frozen blood fractions such as serum and plasma, advancements in analytical technology have developed a method of collecting microsamples of dried whole blood on Ahlstrom 226 grade filter paper that can provide information on long-term fatty acid status of animals. Blood samples were collected from five male pigs in both the traditional frozen method and on dried blood spot cards (DBS). The DBS samples were collected with untreated syringes and tubes, and approximately 320 µL of blood was placed on each card with approximately 80 µL per spot (4 spots). Statistical analysis was performed to compare the two sample groups to each other using the Mann-Whitney U-test and determine if DBS samples were similar to traditional whole blood samples. Of the 30 fatty acids and fatty acid groups with measurable concentrations, only four individual fatty acids, behenic acid, omega-3 docosapentaenoic acid, nervonic acid, and adrenic acid, had statistical differences. Most of these differences were minor and could be due to analytical errors or contamination. Comparisons between sample types found similar concentrations of key omega fatty acids and PUFAs and support the use of DBS collection as a less invasive method of blood collection and fatty acid analysis.

Comparison of African savanna elephant (Loxodonta africana) fatty acid profiles in whole blood, whole blood dried on blood spot cards, serum, and plasma.

BACKGROUND: African elephants in managed care have presented differences in the balance between omega-3 and omega-6 fatty acids, a situation primarily thought to be due to dietary differences between the managed animals and their free-ranging counterparts. Because of this, circulating fatty acid status is included in routine monitoring of elephant health. A method of blood collection that requires only a few drops of whole blood, dried on filter paper (DBS) and can be used for analyzing full fatty acid profiles offers advantages in clinical application. METHODS: This study compared the use of whole blood, and whole blood DBS, serum or plasma for use in evaluating circulating fatty acid composition in African savannah elephants. Samples from six African elephants (two males and four females) were collected during the same week at the NC Zoo, Asheboro, NC. RESULTS: Results found only 2 of 36 individual fatty acids and none of the 10 fatty acid groupings were different when comparing the four blood fraction sample types to each other with Mann-Whitney U-Test pairwise comparisons. Myristic acid (14:0) was lower in the DBS samples than in whole blood, serum, and plasma and pentadecaenoic acid (15:1) was slightly more concentrated in DBS and whole blood. DISCUSSION: Results indicate that fatty acid profile of serum, plasma, whole blood, and DBS are comparable in African elephants. The DBS method offers advantages in acquisition and handling and may be preferable to other methods in both routine health assessment of captive animals and field research on free ranging animals.

Balancing proportions of competing omega-3 and omega-6 highly unsaturated fatty acids (HUFA) in tissue lipids.

People eating different balances of omega-3 and omega-6 nutrients develop predictably different proportions of competing highly unsaturated fatty acids (HUFA) in their tissue lipids. While epidemiological studies have associated wide differences in HUFA balance with disease severity, some clinical studies that did not examine wide differences failed to confirm the association. We examined the degree to which the relative amount of arachidonic acid, the major precursor of omega-6 eicosanoids, differs among people who have widely different dietary intakes of omega-3 and omega-6 nutrients. Gas chromatographic analyses of human blood samples describe the balance among n-3 and n-6 HUFA for different individuals. The proportion of the omega-6 arachidonic acid, from which potent eicosanoids are formed, is not constant. It ranges from 30% to 70% of HUFA while the competing n-3 HUFA range from 60% to 10% of HUFA. Significant differences in clinical outcomes between control and intervention groups have been seen when using dietary interventions that shift the balance of n-3 and n-6 nutrients far enough to create a biologically significant difference in the HUFA balance.

Carbohydrate restriction has a more favorable impact on the metabolic syndrome than a low fat diet.

We recently proposed that the biological markers improved by carbohydrate restriction were precisely those that define the metabolic syndrome (MetS), and that the common thread was regulation of insulin as a control element. We specifically tested the idea with a 12-week study comparing two hypocaloric diets (approximately 1,500 kcal): a carbohydrate-restricted diet (CRD) (%carbohydrate:fat:protein = 12:59:28) and a low-fat diet (LFD) (56:24:20) in 40 subjects with atherogenic dyslipidemia. Both interventions led to improvements in several metabolic markers, but subjects following the CRD had consistently reduced glucose (-12%) and insulin (-50%) concentrations, insulin sensitivity (-55%), weight loss (-10%), decreased adiposity (-14%), and more favorable triacylglycerol (TAG) (-51%), HDL-C (13%) and total cholesterol/HDL-C ratio (-14%) responses. In addition to these markers for MetS, the CRD subjects showed more favorable responses to alternative indicators of cardiovascular risk: postprandial lipemia (-47%), the Apo B/Apo A-1 ratio (-16%), and LDL particle distribution. Despite a threefold higher intake of dietary saturated fat during the CRD, saturated fatty acids in TAG and cholesteryl ester were significantly decreased, as was palmitoleic acid (16:1n-7), an endogenous marker of lipogenesis, compared to subjects consuming the LFD. Serum retinol binding protein 4 has been linked to insulin-resistant states, and only the CRD decreased this marker (-20%). The findings provide support for unifying the disparate markers of MetS and for the proposed intimate connection with dietary carbohydrate. The results support the use of dietary carbohydrate restriction as an effective approach to improve features of MetS and cardiovascular risk.

Comparison of low fat and low carbohydrate diets on circulating fatty acid composition and markers of inflammation.

Abnormal distribution of plasma fatty acids and increased inflammation are prominent features of metabolic syndrome. We tested whether these components of metabolic syndrome, like dyslipidemia and glycemia, are responsive to carbohydrate restriction. Overweight men and women with atherogenic dyslipidemia consumed ad libitum diets very low in carbohydrate (VLCKD) (1504 kcal:%CHO:fat:protein = 12:59:28) or low in fat (LFD) (1478 kcal:%CHO:fat:protein = 56:24:20) for 12 weeks. In comparison to the LFD, the VLCKD resulted in an increased proportion of serum total n-6 PUFA, mainly attributed to a marked increase in arachidonate (20:4n-6), while its biosynthetic metabolic intermediates were decreased. The n-6/n-3 and arachidonic/eicosapentaenoic acid ratio also increased sharply. Total saturated fatty acids and 16:1n-7 were consistently decreased following the VLCKD. Both diets significantly decreased the concentration of several serum inflammatory markers, but there was an overall greater anti-inflammatory effect associated with the VLCKD, as evidenced by greater decreases in TNF-alpha, IL-6, IL-8, MCP-1, E-selectin, I-CAM, and PAI-1. Increased 20:4n-6 and the ratios of 20:4n-6/20:5n-3 and n-6/n-3 are commonly viewed as pro-inflammatory, but unexpectedly were consistently inversely associated with responses in inflammatory proteins. In summary, a very low carbohydrate diet resulted in profound alterations in fatty acid composition and reduced inflammation compared to a low fat diet.