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The Effects of Exogenous Beta-Hydroxybutyrate Supplementation on Metrics of Safety and Health

  • The Applied Science and Performance Institute
  • Applied Science and Performance Institute
  • Applied Science and Performance Institute

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The ketogenic diet is a high-fat, very low-carbohydrate, moderate-protein diet that will induce a state of ketosis. Ketosis is a metabolic state characterized by elevated ketone body production in response to the absence of carbohydrates. Some drawbacks of the ketogenic diet are that it can be difficult to adhere to due to its restrictive nature, and it can also cause some undesirable side effects like gastrointestinal distress and increases in apoB-lipoproteins. In order to maximize the benefit of ketosis and to minimize side effects, supplementing with exogenous beta-hydroxybutyrate may induce a state of temporary ketosis without undesirable side effects. In the present study, 22 healthy male and female adults consumed 12.75 grams of beta-hydroxybutyrate salts or maltodextrin placebo twice daily for 90 days. Comprehensive blood safety analysis, body composition, bone densitometry, psychological and immune surveys, and blood pressure were administered at baseline, 30, 60, and 90 days. There were no significant differences in any measures collected, indicating that exogenous beta-hydroxybutyrate had no detrimental impact on fasting blood values such as electrolyte levels, glucose, hemoglobin A1c, complete blood count, body composition, bone density, psychological well-being, immune status, or blood pressure. We conclude that supplementing with exogenous beta-hydroxybutyrate is safe and well-tolerated by healthy adults.
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International Journal of Nutrition and Food Sciences
2020; 9(6): 154-162
doi: 10.11648/j.ijnfs.20200906.13
ISSN: 2327-2694 (Print); ISSN: 2327-2716 (Online)
The Effects of Exogenous Beta-Hydroxybutyrate
Supplementation on Metrics of Safety and Health
Matthew Stefan
, Matthew Sharp, Raad Gheith, Ryan Lowery, Jacob Wilson
Research Department, The Applied Science and Performance Institute, Tampa, Florida, USA
Email address:
Corresponding author
To cite this article:
Matthew Stefan, Matthew Sharp, Raad Gheith, Ryan Lowery, Jacob Wilson. The Effects of Exogenous Beta-Hydroxybutyrate Supplementation
on Metrics of Safety and Health. International Journal of Nutrition and Food Sciences. Vo l . 9 , N o . 6 , 2 0 2 0 , p p . 1 5 4 - 1 6 2 .
doi: 10.11648/j.ijnfs.20200906.13
Received: November 9, 2020; Accepted: November 24, 2020; Published: December 8, 2020
The ketogenic diet is a high-fat, very low-carbohydrate, moderate-protein diet that will induce a state of ketosis.
Ketosis is a metabolic state characterized by elevated ketone body production in response to the absence of carbohydrates. Some
drawbacks of the ketogenic diet are that it can be difficult to adhere to due to its restrictive nature, and it can also cause some
undesirable side effects like gastrointestinal distress and increases in apoB-lipoproteins. In order to maximize the benefit of
ketosis and to minimize side effects, supplementing with exogenous beta-hydroxybutyrate may induce a state of temporary
ketosis without undesirable side effects. In the present study, 22 healthy male and female adults consumed 12.75 grams of
beta-hydroxybutyrate salts or maltodextrin placebo twice daily for 90 days. Comprehensive blood safety analysis, body
composition, bone densitometry, psychological and immune surveys, and blood pressure were administered at baseline, 30, 60,
and 90 days. There were no significant differences in any measures collected, indicating that exogenous beta-hydroxybutyrate
had no detrimental impact on fasting blood values such as electrolyte levels, glucose, hemoglobin A1c, complete blood count,
body composition, bone density, psychological well-being, immune status, or blood pressure. We conclude that supplementing
with exogenous beta-hydroxybutyrate is safe and well-tolerated by healthy adults.
Beta-Hydroxybutyrate, Ketosis, Safety, Exogenous Ketones
1. Introduction
The ketogenic diet is categorized as a high-fat, very-low
carbohydrate, and moderate protein dietary strategy that is
meant to mimic a fasted state by restricting carbohydrate
intake. Research has commonly defined the intakes of a
ketogenic diet as less than 50 grams of carbohydrates per day,
or 5 to 10 percent of total caloric contribution coming from
carbohydrates, with fat contributing up to 90 percent of total
caloric intake [1, 2]. The goal of the ketogenic diet is to induce
ketosis a metabolic state characterized by increased ketone
body production in response to the absence of carbohydrates.
In order to reach a state of nutritional ketosis, blood ketone
concentration should be between 0.5 millimolar (mM) and 3.0
mM [3]. This rise in endogenous ketones is dependent on
macronutrient availability of glucose and fatty acids, and the
hormonal signaling of glucagon, insulin, and cortisol.
There are many benefits to human health for being in a state
of ketosis from consuming a ketogenic diet. Weight loss
occurs due to the reliance on fatty acid storage, and from the
mitochondria regaining their metabolic flexibility (countering
insulin resistance) [4, 5]. Young et al. demonstrated that as
ketone levels rose, fat loss rose as well [6]. In addition to
weight loss, research has shown that the ketogenic diet does
not have a negative impact on hunger hormones despite a
decline in total caloric intake [7]. To achieve an appetite
suppressive effect, ketones concentrations only need to reach
mild ketosis (greater than 0.5 mM) [8, 10]. Research has also
demonstrated that the application of the ketogenic diet can
have therapeutic benefits on diseases that impact metabolism
[9]; reduce the incidence of seizures in children with epilepsy
[11], improve outcomes of certain neurogenerative diseases
like Parkinson’s Disease [12], may help control glycolytic
phenotype of various cancers by limiting glucose availability
International Journal of Nutrition and Food Sciences 2020; 9(6): 154-162 155
[13], and lower glucose and hemoglobin A1c concentrations
in individuals with type 2 diabetes [14, 15]. Lastly, e le vate d
blood ketones could improve endurance performance and
further optimize substrate metabolism by providing an
alternative source for oxidative phosphorylation [16, 17].
In addition to being carbohydrate restrictive, adherence to
the ketogenic diet can be difficult due to some undesirable side
effects like gastrointestinal discomfort [18] and increases in
apoB-lipoproteins [19]. Therefore, temporary and rapid rises
in blood ketone concentrations with no dietary changes may
be of potential interest and benefit [20]; hence, the relevance
of exogenous ketones [21-23]. The safety of ketone esters has
been previously explored, however, there is a void in the
literature on the safety of ketone salts, which is what this study
investigated. One previous study on the safety of ketone salts
demonstrated that two servings of 7 grams of
beta-hydroxybutyrate (BHB) combined with erythritol,
L-Taurine, and L-Leucine was safe as demonstrated by no
changes in complete blood count (CBC) or biomarkers of a
comprehensive metabolic panel over 6 weeks [24]. Moreover,
markers of cardiovascular health, such as blood pressure,
improved while heart rate remained unchanged. The purpose
of this study is to extend this research to 90 days with a dosage
of 12.75 grams twice per day with additional metrics of safety
and health.
2. Methods
2.1. Subject Criteria
Twenty-two healthy male and female subjects aged 18 to 50
years old enrolled for study participation. Exclusion criteria
included: hypertension, obesity (body mass index [BMI] >30
), smoking or using smokeless tobacco, taking any
prescription medication, or having any underlying health
conditions (metabolic, heart disease, diabetes, kidney disease).
This study was approved by an external institutional review
board (Integ Review IRB, Austin, TX, USA) and all
procedures were in agreement with institutional guidelines and
the Declaration of Helsinki. Prior to engagement in any study
procedures, subjects provided written informed content.
Tab l e 1. Descriptive Subject Characteristics.
Sample Size (n=11) (n=11)
Age (years) 44.45 ± 7.30 45.55 ± 9.05
Height (cm) 166.49 ± 9.80 169.03 ± 11.15
Wei g ht ( k g) 72.79 ± 14.67 78.81 ± 18.07
Body Mass Index (kg/m
) 26.32 ± 5.13 27.27 ± 3.66
2.2. Study Design
The study design was a randomized, double-blinded,
placebo-controlled trial. Subjects were stratified into quartiles
based on BMI and subjects from each quartile were randomly
assigned to conditions using a random number generator
(random. org). The conditions were sent to the primary
investigator in white packages labeled “A” or “B”. These were
administered as 12.75-gram servings of a R-Beta
Hydroxybutyrate (BHB) salt blend (KetoNAT™; Science
Backed Solutions, LLC; Melissa, TX, USA) or a similarly
flavored iso-energetic, iso-volumetric maltodextrin placebo
twice daily for 90 days for a total of 25.5-grams of the
respective condition, daily. Subjects underwent baseline
testing (PRE) which included: blood draw for safety measures
(complete blood count, comprehensive metabolic panel,
automated differential, and hemoglobin A1c), resting blood
pressure and heart rate, psychological mood assessment
(Profile of Mood States; [POMS]), immune status
questionnaire, body composition and bone densitometry.
Following PRE testing, subjects were given a 30-day supply
of either condition “A” or condition “B”. Subjects were
instructed to consume one serving in the morning and one
serving in the afternoon with at least three hours of separation
between servings. Subjects were also asked to track their
caloric intake 3 days every week for the duration of the study.
Lastly, subjects submitted VAS (visual analog scales) to report
subjective measures of satiety, hunger, and psychological
feelings (well-being, mental clarity, etc.). Testing was
repeated for all study procedures in an identical manner to
PRE at 30 days, 60 days, and 90 days following the original
PRE testing date, with the exception of the DXA which was
only performed at PRE and 90 days. Study procedures are
further described below.
2.3. Bone Densitometry and Body-Composition Analysis
Bone densitometry and body composition was determined
by a whole-body scan on a dual-energy x-ray absorptiometry
device (Horizon A DXA System, Hologic Inc, Marlborough,
MA, USA). Fat-free mass, fat mass, body fat percentage, bone
mineral content, and bone density was determined for the total
body with the subject lying in a supine position with knees and
elbows extended. Subjects were instructed not to move for the
entire duration of the scan (approximately 5 minutes). Results
from each scan were uploaded and accessed on computer that
was directly linked to the DXA device. Calibration of the
DXA device was done against a phantom provided by the
manufacturing company prior to testing.
2.4. Venous Blood Measures
Ve no us b l oo d w as e x tr a ct ed by v e ni pu n ct u re o f t he a ntecubital
vein using a 21-gauge syringe and collected into a 10mL EDTA
vacutainer tube (BD Vacutainer®, Becton, Dickinson and
Company, Franklin Lakes, NJ, USA) by a certified phlebotomist.
Afterward, blood samples were centrifuged at 2500 rpm for 10
minutes at 4°C. Resulting serum samples were then aliquoted and
stored at −80°C until further analysis. Samples were thawed once
and analyzed in duplicate in the same assay for each analysis to
avoid compounded inter-assay variance.
2.5. Blood Pressure and Heart Rate
Subjects rested in a supine position for 5 min in a quiet room
at 228°C before the baseline hemodynamic measurements were
obtained. Resting brachial blood pressure and heart rate were
measured on the right arm with an automated digital
156 Matthew Stefan et al.: The Effects of Exogenous Beta-Hydroxybutyrate Supplementation on Metrics of Safety and Health
oscillometric sphygmomanometer (Omron, Model HEM
705-CP; Omron Corporation, Shimogyo-ku, Kyoto, Japan).
Three readings separated by 1-min intervals were taken, and the
mean was used for the analysis.
2.6. Immune Status Questionnaire (ISQ) and Profile of
Mood States (POMS)
The Immune Status Questionnaire (ISQ) is a validated
self-assessment of subjective values of seven different
common symptoms associated with disease [25]. The ISQ was
scored on a 5-point Likert scale from 0 to 4 for how often the
subject has had the following symptoms in the past week;
Never, Sometimes, Regularly, Often, and Almost Always. The
values were summed up to equal a raw score. The raw score
was then converted into a final score between zero and ten,
with 0 being the poor immune status, and ten being excellent
immune status [26].
The Profile of Mood States is a validated self-assessment of
subjective values of forty different moods [25]. Those moods
then fall into seven categories: Tension, Anger, Fatigue,
Depression, Esteem - Related, Vigor, and Confusion. Subjects
were asked to assess each of the forty moods, and if they are
feeling that particular mood “right now”. Subjects assessed the
moods according to a 5-point Likert scale from 0 to 4: Not at All,
A Little, Moderately, Quite a Lot, Extremely. The following
formula was used to determine the overall POMS score:
(Tension + Depression + Anger + Fatigue + Confusion) –
(Vigor + Esteem-Related) + 100
A lo wer scor e indi cate d a be tter mo od , whi le a h igher score
indicated a poor mood [24].
2.7. Visual Analog Scales for Perceived Hunger and
Perceived Mental Clarity
The perceptual measures collected for the study were
perceived Hunger and perceived Mental Clarity. Hunger and
Mental Clarity scales consisted of a scalar representation
numbering from 0-10. On the Hunger Scale, visual descriptors
of “not hungry”, “adequately hungry” and “very hungry”
presented at numbers 0, 5, and 10, respectively. On the Mental
Clarity scale, visual descriptors of “Poor Mental Clarity”,
“Adequate Mental Clarity”, and “Very Mentally Clear” are
presented at numbers 0, 5, and 10, respectively.
2.8. Calorie and Macronutrient Reporting
Subjects were asked to record, and then report, their caloric
intake three times per week using a mobile tracking
application (MyFitnessPal, San Francisco, CA, USA).
2.9. Statistical Analysis
All statistical analyses were performed at the completion of
the study using GraphPad Prism (Version 8, San Diego, CA,
USA). Dependent variables were assessed for normality
(Shapiro-Wilk test) and homogeneity of variances (Levene’s
test). Two-way mixed model analysis of variance (ANOVA)
was performed assuming group and time as fixed factors and
subjects as a random factor. Whenever a significant F value
was obtained, a post hoc test with a Bonferroni adjustment
was used to for multiple comparisons purposes. The alpha
level was set a p ≤ 0.05. Data are reported as mean ± standard
3. Results
3.1. Complete Blood Count
There was no significant between or within group
differences in Complete Blood Count values (p > 0.05, Table
2). Mean and standard deviation are displayed in Table 2.
Tab l e 2. Complete Blood Count Results.
PRE 30 DAY 60 DAY 90 DAYS p Value
WBC (K/uL)
BHB 5.13 ± 0.96 5.07 ± 0.95 5.38 ± 1.07 6.62 ± 2.49 0.1001
PLA 5.91 ± 1.03 5.91 ± 1.05 5.94 ± 1.03 6.19 ± 1.06
RBC (M/uL)
BHB 4.66 ± 0.41 4.61 ± 0.33 4.67 ± 0.34 4.70 ± 0.39 0.1084
PLA 4.36 ± 0.25 4.44 ± 0.34 4.37 ± 0.38 4.31 ± 0.40
Hemoglobin (g/dL)
BHB 14.43 ± 1.06 14.28 ± 0.85 14.37 ± 0.89 14.45 ± 1.01 0.1203
PLA 13.40 ± 1.25 13.64 ± 1.22 13.31 ± 1.29 13.15 ± 1.46
Hematocrit (%)
BHB 47.64 ± 3.07 46.82 ± 2.47 47.17 ± 2.76 46.92 ± 3.24 0.1233
PLA 44.44 ± 3.05 45.05 ± 3.23 45.05 ± 3.23 44.05 ± 3.45
MCV (fl)
BHB 102.36 ± 3.41 101.55 ± 3.53 101.00 ± 3.66 100.00 ± 3.29 0.8246
PLA 101.91 ± 6.69 101.64 ± 6.09 101.09 ± 6.27 100 ± 7.17
MCH (pg)
BHB 31.01 ± 1.01 31.00 ± 1.39 30.85 ± 1.11 30.85 ± 1.07 0.9716
PLA 30.73 ± 2.60 30.85 ± 2.76 30.60 ± 2.53 30.59 ± 2.81
MCHC (g/dL)
BHB 30.29 ± 0.46 30.50 ± 0.66 30.52 ± 0.79 30.85 ± 0.71 0.9812
PLA 30.10 ± 0.95 30.26 ± 1.08 30.24 ± 0.90 30.55 ± 1.06
RDW (%)
International Journal of Nutrition and Food Sciences 2020; 9(6): 154-162 157
PRE 30 DAY 60 DAY 90 DAYS p Value
BHB 13.15 ± 0.59 13.14 ± 0.57 12.94 ± 0.43 12.96 ± 0.47 0.4562
PLA 12.73 ±3.26 13.30 ± 1.06 13.30 ± 1.04 13.50 ± 0.42
Platelets (k/uL)
BHB 218.64±5.86 230.73±70.03 232.27±65.23 236.73±79.15 0.4181
PLA 218.09±34.31 217.5±33.87 217.64±40.66 222.00±45.73
Data reported in mean and standard deviation. P-value is from group by time interaction effect.
3.2. Automated Differential
There was no significant between or within group differences in any values of Automated Differential Cell Count (p > 0.05,
Tab le 3). Mean an d stand ard d ev iatio n are d ispla yed in Table 3.
Tab l e 3. Automated Differential Cell Count
PRE 30 DAY 60 DAY 90 DAYS p Value
Lymphocytes (%)
BHB 35.77 ± 8.38 35.34 ± 7.83 35.68 ± 8.83 30.99 ± 10.14 0.2397
PLA 35.86 ± 7.02 38.42 ± 7.16 37.58 ± 6.68 37.25 ± 6.28
Monocytes (%)
BHB 8.17 ± 5.42 6.01 ± 2.38 5.81 ± 2.45 6.70 ± 3.96 0.4749
PLA 6.35 ± 1.79 6.21 ± 2.10 5.90 ± 1.81 5.72 ± 1.73
Eosionophil (%)
BHB 2.02 ± 1.28 2.26 ± 1.32 2.35 ± 1.34 2.17 ± 1.35 0.6470
PLA 2.01 ± 1.21 2.02 ± 1.34 1.92 ± 1.16 2.05 ± 1.02
Basophil (%)
BHB 1.34 ± 0.27 1.22 ± 0.40 1.09 ± 0.32 1.05 ± 0.23 0.4171
PLA 1.15 ± 0.43 1.14 ± 0.46 1.03 ± 0.35 1.13 ± 0.74
Granulocytes (%)
BHB 54.76 ± 8.55 55.16 ± 8.82 54.15 ± 9.98 59.03 ± 12.18 0.4594
PLA 54.63 ± 8.75 52.25 ± 6.94 53.38 ± 7.07 53.98 ± 7.24
Lymphocytes (k/uL)
BHB 2.03 ± 0.62 1.76 ± 0.31 1.93 ± 0.37 1.86 ± 0.40 0.2579
PLA 2.13 ± 0.53 2.27 ± 0.58 2.23 ± 0.46 2.29 ± 0.53
Monocytes (k/uL)
BHB 0.33 ± 0.15 0.28 ± 0.11 0.30 ± 0.10 0.41 ± 0.25 0.0812
PLA 0.38 ± 0.08 0.35 ± 0.10 0.35 ± 0.12 0.34 ± 0.10
Eosionophil (k/uL)
BHB 0.09 ± 0.05 0.12 ± 0.06 0.12 ± 0.06 0.14 ± 0.08 0.1565
PLA 0.13 ± 0.05 0.13 ± 0.06 0.12 ± 0.08 0.13 ± 0.06
Basos (k/uL)
BHB 0.09 ± 0.03 0.07 ± 0.05 0.06 ± 0.05 0.08 ± 0.04 0.4746
PLA 0.07 ± 0.05 0.08 ± 0.04 0.07 ± 0.05 0.08 ± 0.06
Granulocytes (k/uL)
BHB 2.60 ± 0.99 2.85 ± 0.89 2.98 ± 0.94 4.15 ± 2.62 0.1234
PLA 3.26 ± 0.84 3.11 ± 0.77 3.21 ± 0.87 3.35 ± 0.81
Data reported in mean and standard deviation. P-value is from group by time interaction effect.
3.3. Comprehensive Metabolic Panel
There was no significant between or within group differences in any values of the comprehensive metabolic panel (p > 0.05,
Tab le 4). Mean an d stand ard d ev iatio n are d ispla yed in Table 4.
Tab l e 4. Comprehensive Metabolic Panel
PRE 30 DAY 60 DAY 90 DAY p Value
Total Protein (g/dL)
BHB 6.65 ± 0.35 6.48 ± 0.28 6.51 ± 0.38 6.48 ± 0.43 0.5227
PLA 6.58 ± 0.41 6.56 ± 0.44 6.47 ± 0.36 6.42 ± 0.38
Albumin (g/dL)
BHB 4.49 ± 0.22 4.28 ± 0.22 4.28 ± 0.22 4.44 ± 0.28 0.4284
158 Matthew Stefan et al.: The Effects of Exogenous Beta-Hydroxybutyrate Supplementation on Metrics of Safety and Health
PRE 30 DAY 60 DAY 90 DAY p Value
PLA 4.40 ± 0.20 4.29 ± 0.25 4.2 ± 0.20 4.29 ± 0.29
Globulin (g/dL)
BHB 2.15 ± 0.05 2.2 ± 0.18 2.25 ± 0.26 2.05 ± 0.42 0.8957
PLA 2.18 ± 0.27 2.27 ± 0.23 2.27 ± 0.21 2.13 ± 0.26
BHB 2.10 ± 0.24 1.97 ± 0.17 1.97 ± 0.17 1.91 ± 0.23 0.4320
PLA 2.05 ± 0.23 1.89 ± 0.16 1.88 ± 0.17 2.05 ± 0.30
Bilirubin (mg/dL)
BHB 0.66 ± 0.21 0.63 ± 0.11 0.65 ± 0.22 0.62 ± 0.17 0.8334
PLA 0.57 ± 0.21 0.55 ± 0.14 0.63 ± 0.21 0.54 ± 0.17
Alkaline Phosphate (U/L)
BHB 52.73 ± 11.42 50.09 ± 13.48 49.91 ± 13.23 50.09 ± 11.69 0.4123
PLA 52.64 ± 14.70 51.82 ± 15.09 52.45 ± 12.23 54.09 ± 12.93
BHB 23.27 ± 6.23 27.73 ± 19.89 24.82 ± 9.59 23.55 ± 10.47 0.3370
PLA 28.27 ± 16.87 23.64 ± 8.61 24.45 ± 7.53 22.64 ± 7.70
BHB 23.09 ± 9.60 26.36 ± 12.92 26.09 ± 16.16 25.64 ± 17.47 0.2445
PLA 29.00 ± 27.07 23.27 ± 9.71 25.73 ± 15.11 24.82 ± 15.68
BUN (mg/dL)
BHB 17.00 ± 4.27 14.73 ± 3.17 15.36 ± 4.11 16.09 ± 3.96 0.1065
PLA 15.91 ± 3.75 17.64 ± 5.41 17.00 ± 6.23 17.36 ± 6.34
Creatinine (mg/dL)
BHB 0.86 ± 0.22 0.90 ± 0.23 0.92 ± 0.22 0.89 ± 0.23 0.3388
PLA 0.85 ± 0.13 0.91 ± 0.13 0.89 ± 0.16 1.58 ± 2.17
BUN/Creatinine (mg/dL)
BHB 24.23 ± 4.84 21.30 ± 0.14 21.40 ± NA 21.40 ± N/A 0.0881
PLA 22.48 ± 0.86 24.73 ± 2.11 25.23 ± 1.27 23.30 ± N/A
eGFR (mL/min)
BHB 88.73 ± 15.41 84.55 ± 15.71 81.36 ± 11.93 84.82 ± 12.96 0.2965
PLA 86.64 ± 9.01 80.73 ± 10.07 82.36 ± 11.53 79.00 ± 9.34
Sodium (mg/dL)
BHB 141.45 ± 2.21 140.91 ± 1.64 140.18 ± 1.54 139.64 ± 2.06 0.6043
PLA 141.18 ± 1.83 140.45 ± 1.21 140.00 ± 1.73 140.18 ± 2.09
Potassium (mg/dL)
BHB 4.35 ± 0.35 4.27 ± 0.35 4.13 ± 0.23 4.26 ± 0.24 0.4547
PLA 4.35 ± 0.34 4.19 ± 0.18 4.08 ± 0.16 4.07 ± 0.18
Chloride (mg/dL)
BHB 103.55 ± 2.34 102 ± 1.95 102.91 ± 1.81 102.91 ± 2.26 0.9183
PLA 105.36 ± 2.46 103.82 ± 1.47 104.18 ± 1.78 104.55 ± 2.46
Carbon Dioxide (mL/min)
BHB 27.55 ± 2.25 29.36 ± 1.57 28.91 ± 1.58 29.36 ± 2.42 0.1070
PLA 26.55 ± 2.07 26.64 ± 2.11 27.18 ± 1.99 27.00 ± 2.10
Calcium (mg/dL)
BHB 9.23 ± 0.31 8.97 ± 0.35 9.21 ± 0.35 9.19 ± 0.28 0.1681
PLA 9.14 ± 0.34 8.95 ± 0.32 9.05 ± 0.36 8.92 ± 0.37
Glucose (mg/dL)
BHB 73.45 ± 8.00 87.64 ± 4.43 87.27 ± 4.98 93.36 ± 8.88 0.7419
PLA 76.36 ± 12.47 89.73 ± 10.25 92.09 ± 9.02 93.36 ± 10.53
Hemoglobin A1c
BHB 5.25 ± 0.27 5.10 ± 0.30 5.20 ± 0.31 5.38 ± 0.26 0.3925
PLA 5.32 ± 0.25 5.25 ± 0.29 5.38 ± 0.30 5.48 ± 0.30
Data reported in mean and standard deviation. P-value is from group by time interaction effect. (ALB:GLOB = Albumin:Globulin Ratio, AST =aspartate
aminotransferase, ALT = alanine transaminase, BUN = blood urea nitrogen, eGFR = estimated glomerular filtration rate.)
3.4. Blood Pressure and Heart Rate
There was no significant between or within group differences in resting blood pressure or heart rate (p > 0.05, Table 5). Mean
and standard deviation are displayed in Table 5.
International Journal of Nutrition and Food Sciences 2020; 9(6): 154-162 159
Tab l e 5. Blood Pressure and Cardiovascular Results.
PRE 30 DAY 60 DAY 90 DAYS p Value
Systolic BP (mmHg)
BHB 113.00±10.25 112.18±9.66 109.00±11.25 109.09 ± 9.27 0.6480
PLA 115.91±11.38 117.45±10.35 116.64±10.60 115.00 ± 8.56
Diastolic BP (mmHg)
BHB 68.27 ± 10.68 64.36 ± 7.12 63.55 ± 10.00 65.45 ± 7.61 0.5610
PLA 70.82 ± 10.25 68.55 ± 11.85 70.55 ± 11.10 68.36 ± 8.54
Heart Rate (bpm)
BHB 62.55 ± 10.68 64.55 ± 7.03 65.00 ± 10.00 63.82 ± 8.15 0.4459
PLA 61.64 ± 8.32 64.73 ± 8.68 61.64 ± 8.41 60.91 ± 8.60
Data reported in mean and standard deviation. P-value is from group by time interaction effect. (BP = blood pressure).
3.5. Profile of Mood States (POMS) & Immune Status Questionnaire (ISQ)
There was no significant between or within group differences for responses to the POMS questionnaire or the ISQ (p > 0.05,
Tab le 6). Mean an d stand ard d ev iatio n are d ispla yed in Table 6.
Tab l e 6. Survey Results
PRE 30 DAYS 60 DAYS 90 DAYS p Value
Total POMS Score (a.u.)
BHB 82.55 ± 17.06 77.36 ± 13.43 79.55 ± 12.13 86.09 ± 9.97 0.7240
PLA 73.64 ± 9.14 73.91 ± 11.22 74.45 ± 10.82 78.09 ± 7.83
ISQ Total Score (a.u. )
BHB 9.00 ± 1.10 8.73 ± 1.74 9.00 ± 1.48 8.73 ± 1.42 0.2195
PLA 9.27 ± 0.47 9.45 ± 0.52 9.36 ± 0.67 9.64 ± 0.50
Data reported in mean and standard deviation. P-value is from group by time interaction effect.
3.6. Body Composition & Bone Densitometry
There was no significant between or within group differences in any body composition or bone densitometry values (p > 0.05,
Tab le 7). Mean an d stand ard d ev iatio n are d ispla yed in Table 7.
Tab l e 7. Body Composition & Bone Densitometry Results.
PRE DAY 90 p Value
Total Mass (kg)
BHB 72.66 ± 14.24 72.80 ± 14.44 0.9727
PLA 79.53 ± 17.98 79.60 ± 18.92
Fat Ma ss (kg)
BHB 23.21 ± 9.55 23.96 ± 10.48 0.8739
PLA 23.74 ± 6.87 25.31 ± 8.26
Fat Free Mass (kg)
BHB 49.45 ± 10.96 48.84 ± 9.69 0.5330
PLA 55.79 ± 14.28 54.28 ± 12.66
Body Fat %
BHB 31.66 ± 9.44 32.29 ± 9.63 0.3223
PLA 29.96 ± 5.94 31.45 ± 5.90
Bone Mineral Density (g/cm
BHB 1.13 ± 0.11 1.11 ± 0.09 0.1530
PLA 1.16 ± 0.13 1.18 ± 0.11
Bone Mineral Content (g)
BHB 2291.19 ± 369.46 2248.45 ± 245.24 0.2638
PLA 2510.98 ± 501.73 2534.13 ± 409.74
Data reported in mean and standard deviation. P-value is from group by time interaction effect.
4. Discussion
In this study, we demonstrated the safety of exogenous
BHB under uncontrolled conditions of daily living. The most
significant finding of this study was that sustained 25.5 grams
of daily exogenous ketone salt consumption for 90 days was
safe for healthy adults and that it had no adverse effect on any
blood health markers, hemoglobin A1c, psychological
well-being, or cardiovascular markers of health.
Comprehensive metabolic panel, complete blood count, and
automated differential cell count remained normal and
unaltered after supplementing twice daily with exogenous
160 Matthew Stefan et al.: The Effects of Exogenous Beta-Hydroxybutyrate Supplementation on Metrics of Safety and Health
BHB for 90 days. Furthermore, there were no significant
changes in the POMS or ISQ, resting blood pressure, or
resting heart rate. The findings in this study support, and
further build upon, a previous study by Holland et al. [24]
demonstrating that 6 weeks of exogenous ketone salts
supplementation did not negatively impact various markers of
human health and safety in adults.
There are many controversial views regarding the ketogenic
diet, ketosis, and exogenous ketones. Two such views are that
ketosis can increase the risk of complications in the liver [27],
and the kidneys [28]. However, in human studies, it was
demonstrated that the ketogenic diet may improve clinical
outcomes of nonalcoholic fatty liver disease [29, 31]. In the
present study, we found that markers of liver health; total
protein, albumin, globulin, ALB:GLOB ratio, bilirubin,
alkaline phosphate, AST, and ALT; were unaffected and no
different from placebo with exogenous ketone
supplementation in a healthy population over 90 days.
Depending on macronutrient distribution of caloric intake,
the ketogenic diet can be considered a high protein diet (≥ 20%
of caloric intake). It has been posited that diets higher in protein
could lower pH and increase the acidic load on the kidneys [28].
However, the previously referenced study used a high- protein,
low-carbohydrate diet, while a typical ketogenic diet consists of
low to moderate protein and very-low carbohydrate [30]. With a
typical ketogenic diet, Kossof et al. [32] demonstrated no
increased risk of kidney stones in children. Our study
demonstrated no changes in kidney function markers such as
BUN, creatinine, BUN/creatinine ratio, and eGFR in a healthy
population over 90 days. In addition, acid-base balance was
maintained as demonstrated by blood carbon dioxide and
chloride levels.
The ketogenic diet has shown to lead to calcium loss, and in
some cases, can increase the risk of bone loss. Previous
research has suggested that the ketogenic diet can increase
calcium excretion, which can lead to bone mass loss in
children and adolescents [33–35]. Our study demonstrated no
blood calcium or electrolyte loss, as all electrolyte levels were
unchanged over 90 days in a healthy adult population.
Moreover, we found no changes in bone mineral density or
bone mineral content as assessed by whole-body DXA scans
at PRE and Day 90. These results suggest that BHB does not
lead to bone density or bone mineral content loss.
Lastly, it has been postulated that electrolytes may alter
different markers of cardiovascular health such as blood
pressure and heart rate [36]. Exogenous ketone salts, like the
ones used in the present study, are bound to calcium and
magnesium in order to improve transport and absorption
across the gut-blood barrier. Therefore, it is reasonable to
investigate if altering dietary intake of electrolytes with the
consumption of exogenous ketone salts may have an effect on
blood pressure. However, in our present study, systolic blood
pressure, diastolic blood pressure, and heart rate were
unaffected in the resting state. In addition, blood
concentrations of sodium, potassium, chloride, and calcium all
remained unaltered.
A l im ita ti on o f t he p re sent st ud y i s th e me th od o f
supplementation, and the lack of exercise and dietary control.
Subjects were provided the supplement every 30 days when
they came into the laboratory for testing. They were asked to
return any unused supplements to the laboratory to help keep
subjects honest and to track adherence. Lastly, diet and
exercise were not controlled. However, to keep subjects
accountable, they were encouraged to track and report dietary
intake via a mobile application. Dietary records demonstrated
no differences in daily average of total calories consumed
(BHB: 1430.71 ± 370.84 vs PLA: 1560.04 ± 394.54 kcal/day,
p=0.4381). Future research may seek to directly compare the
effects of exogenous ketone esters and exogenous ketone salts.
5. Conclusion
Exogenous ketone salt supplementation (BHB) can be
considered safe and well tolerated. BHB showed no changes
in comprehensive metabolic panel, automated differential cell
count, complete blood count, hemoglobin A1c, resting blood
pressure and heart rate or psychological surveys over 90 days
of supplementation in a healthy population when compared to
PLA. This study established the safety of long term BHB
supplementation, but further investigation is needed to
examine the efficacy of exogenous ketone supplementation in
other areas of health, longevity, cognitive function, and other
chronic conditions.
Pruvit Ventures Inc. (Melissa, Texas USA) financially
supported the study and supplied the treatment and control
conditions used in this investigation.
Jacob Wilson and Ryan Lowery receive honorira from
Pruvit Ventures Inc. All other authors report no disclosures.
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... Cells were plated at 2.0 × 10 5 cells/mL and the final culture volume was 200 µL. The concentrations of TCN006 were 0.313, 0.625, 1.25, 2.5, 5, and 10 mM (corresponding to 55,111,223,445,890 and 1780 µg/mL). TCN006 was sourced from Baoray Chemical Limited (Hong Kong). ...
... The NOAEL for glycerol in a 50-day clinical study in humans of 2200 mg/kg/ bw/day [45] is 4 times lower than the NOAEL in rats mentioned above (8824 mg/kg bw/day). A clinical study showed that BHB is safe in healthy humans at up to 25.50 g/day for 90 days (estimated 364 mg/kg bw/day for a 70 kg individual), which is 5.5 times lower than the rat NOAEL of 2000 mg/kg bw/day [54,55]. The aforementioned clinical study showed a lack of an effect of BHB on markers of nutritional status of participants (i.e., electrolyte and glucose levels in blood, body weight and composition, and bone density), as well as complete blood count, blood pressure, hemoglobin A1c, immune status and psychological well-being. ...
Full-text available
TCN006, a formulation of (R)-3-Hydroxybutyrate glycerides, is a promising ingredient for enhancing ketone intake of humans. Ketones have been shown to have beneficial effects on human health. To be used by humans, TCN006 must be determined safe in appropriately designed safety studies. The results of a bacterial reverse mutation assay, an in vitro mammalian micronucleus study, and 14-and 90-day repeat dose toxicity studies in rats are reported herein. In the 14- and 90-day studies, male and female Wistar rats had free access to drinking water containing 0, 75,000, 125,000 or 200,000 ppm TCN006 for 92 and 93 days, respectively. TCN006 tested negative for genotoxicity and the no observed adverse effect level (NOAEL) for toxicity in the 14- and 90-day studies was 200,000 ppm, the highest dose administered. In the longer term study, the mean overall daily intake of TCN006 in the 200,000 ppm groups was 14,027.9 mg/kg bw/day for males and 20,507.0 mg/kg bw/day for females. At this concentration, palatability of water was likely affected, which led to a decrease in water consumption in both males and females compared to respective controls. This had no effect on the health of the animals. Although the rats were administered very high levels of (R)-3-Hydroxybutyrate glycerides, there were no signs of ketoacidosis.
... As an alternative, or to supplement a well-formulated ketogenic diet, exogenous ketones have been shown to be a reliable method of rapidly increasing blood ketone levels and promoting a state of ketosis [18]. In addition, the safety of exogenous beta-hydroxybutyrate salts (BHB) has only been explored in a healthy adult population and was determined to be safe [19]. Although the ketogenic diet has appeared to be safe and effective in treating epilepsy and reducing the occurrence of seizures in adolescents [4,20], no studies have examined the safety of supplemental ketones (BHB) on a healthy, adolescent population. ...
... The primary finding of this study was that sustained 7.5 g of daily exogenous BHB salt consumption for 90 days was safe for healthy adolescents and had no adverse effect on hematological safety markers, bone densitometry, emotional intelligence and psychological well-being, or cardiovascular markers of health. The findings of this study support and further build upon previous research done in healthy adults [18,19], demonstrating that 90 days of exogenous BHB salt supplementation was considered safe. Secondly, a pilot study was performed to ensure that one serving (3.75 g of exogenous BHB salt) induced a metabolic state of ketosis (≥0.5 mmol/L). ...
Full-text available
The ketogenic diet is a high-fat, very low-carbohydrate, moderate-protein diet that will induce a state of ketosis, but because of its restrictive nature, it may be difficult to adhere to, especially in adolescents. Supplementing with exogenous beta-hydroxybutyrate salts may induce a state of temporary ketosis without any undesirable side effects, thereby promoting the benefits of ketosis and minimizing adherence requirements to a ketogenic diet. To date, beta-hydroxybutyrate supplementation in healthy adolescents has not been explored. Therefore, the purpose of this study was to examine the safety of exogenous beta-hydroxybutyrate salt supplementation in a healthy adolescent population. In the present study, 22 healthy male and female adolescents consumed 3.75 g of beta-hydroxybutyrate salts or maltodextrin placebo twice daily for 90 days. Comprehensive blood safety analysis, bone densitometry, happiness and emotional intelligence surveys, and blood pressure were assessed at Pre, Day 45, and Day 90. There were no significant differences detected in subjects supplementing with beta-hydroxybutyrate salts, indicating that exogenous beta-hydroxybutyrate salts had no detrimental impact on fasting blood safety metrics, bone density, happiness, emotional intelligence, or blood pressure. We conclude that supplementing with exogenous beta-hydroxybutyrate salts is safe and well-tolerated by healthy adolescents.
... In a further update in March 2021, the denomination was changed to a 'DL-beta hydroxybutyrate salt blend containing D and L salts at a 95:5 D:L ratio'. The Panel notes that the data were published in the literature in December 2020 (Stefan et al., 2020), indicating that the test material was a 'R-BHB salt blend', prior to the modifications made on the reporting of the identity and production process of the NF. ...
Full-text available
Following a request from the European Commission, the EFSA Panel on Nutrition, Novel Foods and Food Allergens (NDA) was asked to deliver an opinion on β-hydroxybutyrate (BHB) salts as a novel food (NF) pursuant to Regulation (EU) 2015/2283. The NF consists of sodium, magnesium and calcium BHB salts, and is proposed to be used by adults as a food ingredient in a number of food categories and as food supplement. The data provided by the applicant about the identity, the production process and the compositional data of the NF over the course of the risk assessment period were overall considered unsatisfactory. The Panel noted inconsistencies in the reporting of the test item used in the subchronic toxicity study and human studies provided by the applicant. Owing to these deficiencies, the Panel cannot establish a safe intake level of the NF. The Panel concludes that the safety of the NF has not been established.
... Ketone salts, MCTs, and at least one ketone ester have been widely studied in humans and shown to be safe and largely well tolerated [30][31][32]. Depending on the specific compound, formulation, dose and dosing context, there are some mild-moderate, transient side effects associated with exogenous ketone ingestion; these typically include nausea, dizziness and stomach cramping [33,34], not unlike the transient side-effects sometimes reported by individuals on a ketogenic diet [35]. However, associated side-effects have not been a barrier to the increasing use of exogenous ketones in research studies and in consumer products. ...
Full-text available
Nutritional ketosis is a state of mildly elevated blood ketone concentrations resulting from dietary changes (e.g., fasting or reduced carbohydrate intake) or exogenous ketone consumption. In this study, we determined the tolerability and safety of a novel exogenous ketone diester, bis-hexanoyl-(R)-1,3-butanediol (BH-BD), in a 28-day, randomized, double-blind, placebo-controlled, parallel trial (NCT04707989). Healthy adults (n = 59, mean (SD), age: 42.8 (13.4) y, body mass index: 27.8 (3.9) kg/m2) were randomized to consume a beverage containing 12.5 g (Days 0–7) and 25 g (Days 7–28) of BH-BD or a taste-matched placebo daily with breakfast. Tolerability, stimulation, and sedation were assessed daily by standardized questionnaires, and blood and urine samples were collected at Days 0, 7, 14, and 28 for safety assessment. There were no differences in at-home composite systemic and gastrointestinal tolerability scores between BH-BD and placebo at any time in the study, or in acute tolerability measured 1-h post-consumption in-clinic. Weekly at-home composite tolerability scores did not change when BH-BD servings were doubled. At-home scores for stimulation and sedation did not differ between groups. BH-BD significantly increased blood ketone concentrations 1-h post-consumption. No clinically meaningful changes in safety measures including vital signs and clinical laboratory measurements were detected within or between groups. These results support the overall tolerability and safety of consumption of up to 25 g/day BH-BD.
... 87,88], their effect on the inflammatory profile in vivo in humans remains unclear and will require further investigation. Recently, a controlled randomized trial in healthy humans found no deleterious effects on immune status and cardiometabolic markers after ingesting exogenous ketones precursors for 90 days [89]. Changes in IL8 and TNF-alpha in the whole group were correlated to changes in triglycerides (ρ = 0.625, P < 0.001 and ρ = 0.401, P = 0.015 respectively; Figure 1). ...
Introduction Mild cognitive impairment (MCI) is often accompanied by metabolic abnormalities and inflammation that might play a role in the development of cognitive impairment. The use of ketogenic medium-chain triglycerides (kMCT) to improve cognition in this population has shown promising results but remains controversial because of the potentially detrimental effect of elevated intake of saturated fatty acids on cardiovascular (CV) health and perhaps inflammatory processes. The primary aim of this secondary data analysis report is to describe changes in cardiometabolic markers and peripheral inflammation during a 6-month kMCT intervention in MCI. Methods Thirty-nine participants with MCI completed the intervention of 30 g/day of either a kMCT drink or calorie-matched placebo (high-oleic acid) for 6 months. Plasma concentrations of cardiometabolic and inflammatory markers were collected before (fasting state) and after the intervention (2 hours following the last drink). Results A mixed model ANOVA analysis revealed a time by group interaction for ketones (P < 0.001), plasma 8:0 and 10:0 acids (both P < 0.001) and IL-8 (P = 0.002) with follow up comparison revealing a significant increase in the kMCT group (+48%, P = 0.005), (+3,800 and +4,900%, both P < 0.001) and (+147%, P < 0.001) respectively. A main effect of time was observed for insulin (P = 0.004), triglycerides (P = 0.011) and non-esterified fatty acids (P = 0.036). Conclusion Under these study conditions, 30 g/d of kMCT taken for six months and up to 2-hour before post-intervention testing had minimal effect on an extensive profile of circulating cardiometabolic and inflammatory markers as compared to a placebo calorie-matched drink. Our results support the safety kMCT supplementation in individuals with MCI. The clinical significance of the observed increase in circulating IL-8 levels is presently unknown and awaits future studies.
Full-text available
Significance Ketogenic diet is an effective treatment for nonalcoholic fatty liver disease (NAFLD). Here, we present evidence that hepatic mitochondrial fluxes and redox state are markedly altered during ketogenic diet-induced reversal of NAFLD in humans. Ketogenic diet for 6 d markedly decreased liver fat content and hepatic insulin resistance. These changes were associated with increased net hydrolysis of liver triglycerides and decreased endogenous glucose production and serum insulin concentrations. Partitioning of fatty acids toward ketogenesis increased, which was associated with increased hepatic mitochondrial redox state and decreased hepatic citrate synthase flux. These data demonstrate heretofore undescribed adaptations underlying the reversal of NAFLD by ketogenic diet and highlight hepatic mitochondrial fluxes and redox state as potential treatment targets in NAFLD.
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The self-assessment of perceived immune status is important, as this subjective observation leads individuals to decide whether or not to seek medical help or adapt their lifestyle. In addition, it can be used in clinical settings and research. The aim of this series of studies was to develop and validate a short questionnaire to assess perceived immune functioning. Five surveys were conducted among Dutch and International young healthy adults (18–30 years old), and two others among older age groups with various health complaints. For the first study, an existing immune functioning scale was modified and elaborated resulting in 23 immune-health-related items, of which the occurrence was rated on a 5-point Likert scale. A student sample was surveyed, and the results were used to shorten the 23-item listing into a 7-item scale with a predictive validity of 85%. Items include “sudden high fever”, “diarrhea”, “headache”, “skin problems (e.g., acne and eczema)”, “muscle and joint pain”, “common cold” and “coughing”. The scale is named Immune Status Questionnaire (ISQ), and it aims to assess perceived immune status over the preceding year. The second study revealed that the ISQ score correlated significantly with a 1-item perceived immune functioning (r = 0.383, p < 0.0001). In the third study, the final Likert scale descriptors were determined (“never”, “sometimes”, “regularly”, “often” and “(almost) always)”. The fourth study showed that the test–retest reliability of the ISQ is acceptable (r = 0.80). The fifth study demonstrated the association of ISQ scores with various neuropsychological and health correlates in an international sample, including perceived health and immune fitness, as well as levels of stress, fatigue, depression and anxiety. Study 6 demonstrated significant associations between ISQ scores and experiencing irritable bowel syndrome (IBS) symptoms in a sample of insomnia patients. Study 7 compared the effect of a dietary intervention in participants reporting “poor health” versus “normal health”. It is shown that ISQ scores can differentiate between those with poor and normal health, and that an effective intervention is associated with a significant improvement in ISQ scores. Data from Study 7 were further used to determine an ISQ cut-off value for reduced immune functioning, and a direct comparison with 1-item perceived immune functioning scores enabled constructing the final scoring format of the ISQ. In conclusion, the ISQ has appropriate face, content, and construct validity and is a reliable, stable and valid method to assess the past 12 month’s perceived immune status.
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Nutrition is known to exert an undeniable impact on blood pressure with especially salt (sodium chloride), but also potassium, playing a prominent role. The aim of this review was to summarize meta-analyses studying the effect of different electrolytes on blood pressure or risk for hypertension, respectively. Overall, 32 meta-analyses evaluating the effect of sodium, potassium, calcium and magnesium on human blood pressure or hypertension risk were included after literature search. Most of the meta-analyses showed beneficial blood pressure lowering effects with the extent of systolic blood pressure reduction ranging between −0.7 (95% confidence interval: −2.6 to 1.2) to −8.9 (−14.1 to −3.7) mmHg for sodium/salt reduction, −3.5 (−5.2 to −1.8) to −9.5 (−10.8 to −8.1) mmHg for potassium, and −0.2 (−0.4 to −0.03) to −18.7 (−22.5 to −15.0) mmHg for magnesium. The range for diastolic blood pressure reduction was 0.03 (−0.4 to 0.4) to −5.9 (−9.7 to −2.1) mmHg for sodium/salt reduction, −2 (−3.1 to −0.9) to −6.4 (−7.3 to −5.6) mmHg for potassium, and −0.3 (−0.5 to −0.03) to −10.9 (−13.1 to −8.7) mmHg for magnesium. Moreover, sufficient calcium intake was found to reduce the risk of gestational hypertension.
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A crucial component of non‐alcoholic fatty liver disease (NAFLD) pathogenesis is lipid stress, which may contribute to hepatic inflammation and activation of innate immunity in the liver. However, little is known regarding how dietary lipids, including fat and cholesterol, may facilitate innate immune activation in vivo. We hypothesized that dietary fat and cholesterol drive NAFLD progression to steatohepatitis and hepatic fibrosis by altering the transcription and phenotype of hepatic macrophages This hypothesis was tested by using RNA‐seq methods to characterize and analyze sort‐purified hepatic macrophage populations that were isolated from mice fed diets with varying amounts of fat and cholesterol The addition of cholesterol to a high fat diet triggered hepatic pathology reminiscent of advanced non‐alcoholic steatohepatitis (NASH) in humans characterized by signs of cholesterol dysregulation, generation of oxidized LDL, increased recruitment of hepatic macrophages, and significant fibrosis. RNA‐seq analyses of hepatic macrophages in this model revealed that dietary cholesterol induced a tissue repair and regeneration phenotype in Kupffer cells and recruited infiltrating macrophages to a greater degree than fat. Furthermore, comparison of diseased Kupffer cells and infiltrating macrophages revealed that these two macrophage subsets are transcriptionally diverse. Finally, direct stimulation of murine and human macrophages with oxidized LDL recapitulated some of the transcriptional changes observed in the RNA‐seq study. These findings indicate that fat and cholesterol synergize to alter macrophage phenotype, and they also challenge the dogma that Kupffer cells are purely pro‐inflammatory in NASH This comprehensive view of macrophage populations in NASH indicates novel mechanisms by which cholesterol contribute to NASH progression and identifies potential therapeutic targets for this common disease. This article is protected by copyright. All rights reserved.
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The low-carbohydrate high-fat diet (LCHFD), also known as the ketogenic diet, has cycled in and out of popularity for decades as a therapeutic program to treat metabolic syndrome, weight mismanagement, and drug-resistant disorders as complex as epilepsy, cancer, dementia, and depression. Despite the benefits of this diet, health care professionals still question its safety due to the elevated serum ketones it induces and the limited dietary fiber. To compound the controversy, patient compliance with the program is poor due to the restrictive nature of the diet and symptoms related to energy deficit and gastrointestinal adversity during the introductory and energy substrate transition phase of the diet. The studies presented here demonstrate safety and efficacy of the diet including the scientific support and rationale for the administration of exogenous ketone bodies and ketone sources as a complement to the restrictive dietary protocol or as an alternative to the diet. This review also highlights the synergy provided by exogenous ketone, β -hydroxybutyrate (BHB), accompanied by the short chain fatty acid, butyrate (BA) in the context of cellular and physiological outcomes. More work is needed to unveil the molecular mechanisms by which this program provides health benefits.
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Background Adaptation to a ketogenic diet (keto-induction) can cause unpleasant symptoms, and this can reduce tolerability of the diet. Several methods have been suggested as useful for encouraging entry into nutritional ketosis (NK) and reducing symptoms of keto-induction. This paper reviews the scientific literature on the effects of these methods on time-to-NK and on symptoms during the keto-induction phase. Methods PubMed, Science Direct, CINAHL, MEDLINE, Alt Health Watch, Food Science Source and EBSCO Psychology and Behavioural Sciences Collection electronic databases were searched online. Various purported ketogenic supplements were searched along with the terms “ketogenic diet”, “ketogenic”, “ketosis” and ketonaemia (/ ketonemia). Additionally, author names and reference lists were used for further search of the selected papers for related references. Results Evidence, from one mouse study, suggests that leucine doesn’t significantly increase beta-hydroxybutyrate (BOHB) but the addition of leucine to a ketogenic diet in humans, while increasing the protein-to-fat ratio of the diet, doesn’t reduce ketosis. Animal studies indicate that the short chain fatty acids acetic acid and butyric acid, increase ketone body concentrations. However, only one study has been performed in humans. This demonstrated that butyric acid is more ketogenic than either leucine or an 8-chain monoglyceride. Medium-chain triglycerides (MCTs) increase BOHB in a linear, dose-dependent manner, and promote both ketonaemia and ketogenesis. Exogenous ketones promote ketonaemia but may inhibit ketogenesis. Conclusions There is a clear ketogenic effect of supplemental MCTs; however, it is unclear whether they independently improve time to NK and reduce symptoms of keto-induction. There is limited research on the potential for other supplements to improve time to NK and reduce symptoms of keto-induction. Few studies have specifically evaluated symptoms and adverse effects of a ketogenic diet during the induction phase. Those that have typically were not designed to evaluate these variables as primary outcomes, and thus, more research is required to elucidate the role that supplementation might play in encouraging ketogenesis, improve time to NK, and reduce symptoms associated with keto-induction.
The ketogenic diet (KD) is a high fat, adequate protein, low carbohydrate diet that is used clinically for refractory pediatric epilepsy, and is under investigation as a treatment for a wide variety of disorders, including many neurological diseases, cancer, obesity, and diabetes. Studies suggest that many of the beneficial effects associated with the KD are mechanistically attributable to the ketone bodies, leading our lab and others to develop and test exogenous ketone supplements (KS). KS elevate blood ketones without the need for dietary restriction of carbohydrate. In order to further our understanding of this new technology, we sought to characterize the metabolic effects, safety, and toxicity of KS in healthy rats. KS tested include the R,S‐1,3‐butanediol acetoacetate diester, 1,3‐butanediol, medium chain triglyceride oil, caprylic acid, and beta‐hydroxybutyrate mineral salts, alone and in various combinations. Healthy rats were administered KS by oral gavage or in their food in an acute (once, or 7 days), sub‐chronic (1 month) or chronic (4 months) fashion and analyzed for a variety of metabolic and physiologic biomarkers. KS rapidly elevate blood ketones to therapeutic levels (2–5mM), and simultaneously decrease blood glucose. Some types of KS induce a mild calorie restriction effect, and preliminary evidence suggests ketone ester treatment during CR may preserve lean muscle mass in this catabolic state via a non‐insulin dependent mechanism. Satiety may be increased by KS as demonstrated by an elevation in leptin following KE administration, even when animals were self calorie‐restricting. KS significantly alters the global metabolome, indicating a shift from carbohydrate‐based to fat‐based metabolism. Some markers of this were an elevation in medium chain fatty acids, an increase in acetyl coA and acetylcarnitine suggestive of enhanced mitochondrial LCFA oxidation, and an increase in Kreb's cycle intermediates such as citrate, succinate, and fumarate. Inflammatory profiling following chronic, but not acute, KS administration revealed decreases in many pro‐inflammatory cytokines, including IL‐1β, IL‐6, IFN‐γ, MCP‐1, and RANTES. KS appears safe as there were no signs of toxicity or adverse changes in total cholesterol, HDL, LDL, triglycerides, or markers of liver/kidney function over the chronic treatment protocol. KS may be a useful alternative or adjuvant to the KD for disorders where nutritional ketosis is therapeutic. Support or Funding Information Office of Naval Research, Scivation Inc, Disruptive Nutrition, Epigenix Foundation This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
Athletes, clinicians, and practitioners are increasingly interested in the proposed performance and therapeutic benefits of nutritional ketosis (NK). NK is best operationally defined as a nutritionally induced metabolic state resulting in blood β-hydroxybutyrate concentrations of ≥0.5 mM. Most tissues readily metabolize ketone bodies (KBs), and KBs in turn regulate metabolism and signaling in both a systemic and tissue-specific manner. During fasting, starvation, or ketogenic diets, endogenous synthesis of KBs is amplified resulting in a state of NK. Orally administered exogenous ketone supplements rapidly elevate circulating KBs and produce a similar, but far from identical, metabolic state. NK results in a number of convergent features regardless of endogenous or exogenous induction; however, important differences also are observed. The implications of NK across health, disease, and performance is rapidly becoming more evident, thus acknowledging the convergent and divergent features of NK is critical for fully understanding the potential utility of this metabolic state.
Throughout history, the only way humans could raise their blood ketone levels was by several days of fasting or by following a strict low-carb, high-fat diet. A recently developed, dietary source of ketones, a ketone monoester, elevates d-β-hydroxybutyrate (βHB) to similar concentrations within minutes, with βHB remaining raised for several hours. To date, the longest human safety study of the exogenous ketone ester was for 5 days, but longer consumption times may be desired. Here we report results for 24 healthy adults, aged 18-70 years, who drank 25 ml (26.8 g) of the ketone monoester, (R)-3-hydroxybutyl (R)-3-hydroxybutyrate, three times a day for 28 days (a total of 2.1 L). Anthropomorphic measurements, plus fasting blood and urine analyses were made weekly. It was found that elevating blood βHB concentrations from 0.1 to 4.1 (±1.1) mM three times a day for 28 days had no effect on body weights or composition, fasting blood glucose, cholesterol, triglyceride or electrolyte concentrations, nor blood gases or kidney function, which were invariably normal. Mild nausea was reported following 6 of the 2,016 drinks consumed. We conclude that sustained exogenous ketosis using a ketone monoester is safe and well-tolerated by healthy adults.