Every 7 days, animals were briefly fasted (4 h, water available) prior to intragastric gavage to standardize levels of blood metabolites prior to glucose and βHB measurements at baseline. Baseline (time 0) was immediately prior to gavage. Whole blood samples (10 μL) were taken from the saphenous vein for analysis of glucose and βHB levels with the commercially available glucose and ketone monitoring system Precision Xtra™ (Abbott Laboratories, Abbott Park, IL). Blood glucose and βHB were measured at 0, 0.5, 1, 4, 8, and 12 h after test substance administration, or until βHB returned to baseline levels. Food was returned to animals after blood analysis at time 0 and gavage. At baseline and week 4, whole blood samples (10 μL) were taken from the saphenous vein immediately prior to gavage (time 0) for analysis of total cholesterol, high-density lipoprotein (HDL), and triglycerides with the commercially available CardioChek™ blood lipid analyzer (Polymer Technology Systems, Inc., Indianapolis, IN). Low-density lipoprotein (LDL) cholesterol was calculated from the three measured lipid levels using the Friedewald equation: (LDL Cholesterol = Total Cholesterol - HDL - (Triglycerides/5)) [51, 52]. Animals were weighed once per week to track changes in body weight associated with hyperketonemia.


Firstly, in a randomized four-arm cross-over study, blood βHB concentrations were compared following ingestion of equal amounts of βHB as a KE or a KS at two doses by healthy volunteers at rest (Study 1; n = 15). Secondly, in a randomized five-arm cross-over study, inter- and intra-participant repeatability of ketosis was examined following ingestion of identical KE drinks, twice whilst fed and twice whilst fasted. As a control, participants also consumed one isocaloric (1.9 kCal.kg−1) dextrose drink (Study 2; n = 16). Finally, blood d-βHB was measured after equal amounts of KE were given as three drinks (n = 12) or a constant nasogastric (NG) infusion (n = 4) (Study 3; total n = 14) over 9 h.

Beta-Hydroxybutyrate (BHB) is a ketone body produced by the liver, from fat, for energy when glucose isn’t available. It ultimately becomes the body and brain’s primary source of energy. Since the liver naturally produces BHB during ketosis, the process can take quite some time, often resulting in symptoms of fatigue, hunger (cravings for sugar, a faster energy source), and mental cloudiness. That’s why supplementing BHB on a keto diet can have a profound positive impact.

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