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Effect of Raspberry Ketone on Normal, Obese and Health-Compromised Obese Mice: A Preliminary Study

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  • NCNPR, University of Mississippi

Abstract and Figures

Raspberry ketone (RK)—an aromatic compound found mostly in red raspberries (Rubus idaeus) is widely used as an over the counter product for weight loss. The present study was conducted to determine adverse effects associated with RK in obese and health-compromised obese mice. Two sets of experiments were conducted on normal obese and health-compromised obese mice treated with RK for a duration of 10 days. Obese conditions were induced by feeding mice a high fat diet for 10 weeks, while the health compromised obese mouse model was developed by a single intraperitoneal injection of a nontoxic dose of lipopolysaccharide (LPS) (6 mg/kg) to obese mice. Results showed that RK (165, 330, and 500 mg/kg) under obese as well as health-compromised condition retarded the gain in body weights as compared to the control groups. RK at doses 330 and 500 mg/kg resulted in 67.6 and 50% mortality, respectively in normal obese mice and 70% mortality was observed in health-compromised obese mice treated with RK at 500 mg/kg. At higher doses deaths were observed earlier than those given lower doses of RK. Significant elevations in blood alanine transaminase (ALT) were also observed with RK treatment in obese mice. Blood glucose levels were significantly elevated in all groups of mice treated with RK. This study suggests that higher doses of RK may cause adverse effects in health compromised conditions. Under these conditions, prolonged use of RK, especially in high doses, may pose a health hazard.
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IJDS#1674996, VOL 0, ISS 0
Effect of Raspberry Ketone on Normal, Obese and
Health-Compromised Obese Mice: A Preliminary
Study
Tahir Maqbool Mir, Guoyi Ma, Zulfiqar Ali, Ikhlas A. Khan, and Mohammad
K. Ashfaq
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ARTICLE
Effect of Raspberry Ketone on Normal, Obese and
Health-Compromised Obese Mice: A Preliminary Study
Tahir Maqbool Mir
a
, Guoyi Ma
b
, Zulfiqar Ali
a
, Ikhlas A. Khan
a
, and
Mohammad K. Ashfaq
a
Q1
a
National Center for Natural Product Research, School of Pharmacy, University of Mississippi, University,
MS, USA;
b
Drug Discovery Division, Southern Research, Birmingham, AL, UK
ABSTRACT
Background: Raspberry ketone (RK)an aromatic compound found
mostly in red raspberries (Rubus idaeus) is widely used as an over
the counter product for weight loss. Aim: The present study was
conducted to determine adverse effects associated with RK in obese
and health-compromised obese mice. Methodology: Two sets of
experiments were conducted on normal obese and health-compro-
mised obese mice treated with RK for a duration of 10 days. Obese
conditions were induced by feeding mice a high fat diet for
10 weeks, while the health compromised obese mouse model was
developed by a single intraperitoneal injection of a nontoxic dose of
lipopolysaccharide (LPS) (6 mg/kg) to obese mice. Results: Results
showed that RK (165, 330, and 500 mg/kg) under obese as well as
health-compromised condition retarded the gain in body weights as
compared to the control groups. RK at doses 330 and 500mg/kg
resulted in 67.6 and 50% mortality, respectively in normal obese
mice and 70% mortality was observed in health-compromised obese
mice treated with RK at 500 mg/kg. At higher doses deaths were
observed earlier than those given lower doses of RK. Significant ele-
vations in blood alanine transaminase (ALT) were also observed with
RK treatment in obese mice. Blood glucose levels were significantly
elevated in all groups of mice treated with RK. Conclusion: This study
suggests that higher doses of RK may cause adverse effects in health
compromised conditions. Under these conditions, prolonged use of
RK, especially in high doses, may pose a health hazard.
Q2
KEYWORDS
Health compromised
conditions; high fat diet;
mortality; obese;
Raspberry ketone
Introduction
Since ancient times, plants have served as the primary and invaluable sources of new
drugs. Renewed interests in plant-derived drugs over conventional medicines have
emerged, likely due to consumer perception pertaining to side effects associated with
conventional drugs. Till now, natural products in the form of herbal remedies, medi-
cinal plants, functional foods, and their constituents are effective in treating many
diseases including cancer (Kim et al. 2018). Tremendous research efforts on natural
products are being carried out to develop alternatives to synthetic drugs. As per
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CONTACT Mohammad K. Ashfaq mkashfaq@olemiss.edu National Center for Natural Product Research, School of
Pharmacy, University of Mississippi, 2047 TCRC, University, MS 38677.
Color versions of one or more of the figures in the article can be found online at www.tandfonline.com/ijds.
ß2019 Taylor & Francis Group, LLC
JOURNAL OF DIETARY SUPPLEMENTS
https://doi.org/10.1080/19390211.2019.1674996
Editorial in Lancet, Gastroenterology and hepatology (2018), the sales of natural supple-
ments in the United States alone, reached $37 billion in 2014, an increase from $7 bil-
lion in 1994. In Australia, spending on complementary medicines increased by more
than 100% between 1996 and 2004 (Bailey et al. 2013). Hundreds of natural products
are being sold in stores as over the counter herbal products or online with tremen-
dously conflated statements pertaining to their abilities to positively affect weight loss,
immune health, anti-diabetics, hair growth as well as dietary supplements. However, the
health benefits associated with natural products are substantiated with little clinical evi-
dence to prove their mechanistic aspects. Experimental studies have demonstrated that
several natural products have been associated with harmful side effects including liver
toxicity (Abdel-Bakky et al. 2010; Chan et al. 2010; Wang et al. 2015). The severity of
side effects associated with natural products depends on the dose, consumption duration
of the product and most importantly on the general health of the individual consumer.
The general health of the individual plays an important role in herbal product associ-
ated toxicities. It has been reported that several natural products can contribute to liver
toxicity when consumed during health compromised conditions (Maddox et al. 2010). It
has been found that a nontoxic dose of lipopolysaccharide (LPS) reduces the toxic
threshold of xenobiotics by inducing mild inflammatory conditions in the body (Ganey
et al. 2004; Deng et al. 2009; Tu et al. 2015). To this end, we have already established a
mouse model for hepatotoxicity, with co-administration of pyrrolizidine alkaloid
(monocrotaline) and a nontoxic dose of LPS (6 mg/kg) (Abdel-Bakky et al. 2010).
Similarly, our previous studies demonstrated that administration of epigallocatechin-3-
gallate (EGCG), a major constituent of green tea to mice sensitized with nontoxic dose
of LPS caused liver damage (Saleh et al. 2013). Raspberry ketone (4-(4-hydroxyphenyl)
butan-2-one, RK) is a major aromatic compound of red raspberries (Rubus idaeus)
(Schinz and Seidel 1957; Gallois 1982; Patel et al. 2004). Raspberry ketone is used as fla-
voring substance in a variety of processed foods such as soft drinks, puddings, yogurt,
baked goods, sweets and ice cream (Beekwilder et al. 2007; Crispim et al. 2010).
Initially, it was introduced as a fragrance and flavoring agent (Opdyke 1978). The
Flavor and Extract Manufacturers Association of the United States placed Raspberry
ketone on generally recognized as safe (GRAS) status (Opdyke 1978; Lin et al. 2011).
However, since 2012 the whole scenario about RK changed when claims of its anti-
obesity (weight reducing) property were made in the US media as No. 1miraculous
weight-loss supplement. This resulted in the dramatic increase in sales of dietary supple-
ments containing RK (Bootsman et al. 2014). RK is believed to affect fat metabolism by
enhancing lipolysis and fatty acid oxidation (Park 2010), resulting in its use in a wide
array of anti-obesity or weight loss supplements. In addition to its role as a weight
reducing natural supplement, RK is claimed to possess other pharmacological activities
including hepatoprotective, anti-androgenic activity (Ogawa et al. 2010), anti-inflamma-
tory activity by blocking the nuclear transcription factor-kB (NF-kB) activation (Jeong
and Jeong 2010), depigmenting activity (Lin et al. 2011), anti-obese activity (Park 2010;
Lopez et al. 2013), antihypertensive (Jia et al. 2011) and cardio-protective (Khan et al.
2018) activities.
Several reports revealed that the amount of RK sold in US markets in dietary supple-
ments is well above the maximum recommended concentration for food products.
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Further studies have been recommended to ensure its safety at such concentrations
(Bredsdorff et al. 2015; Lee 2016). As the weight reducing products are usually con-
sumed by overweight individuals, it is imperative that a safe dose be determined.
Considering that some of the consumers of such weight reduction products tend to
become over ambitious and may use higher doses than those prescribed by the manu-
facturer. Thus, studies need to be undertaken to define the safe levels of such products
under healthy and health-compromised conditions. To this end, we conducted a prelim-
inary study to determine the adverse effects of RK at different doses in obese and
health-compromised obese mice.
Materials and methods
Materials
Raspberry ketone and lipopolysaccharide (E. Coli O55:B5) were purchased from Sigma-
Aldrich (St. Louis, MO) Blood ALT level was measured using an automated VetScan
dry chemistry analyzer with V-DPP rotors (Abaxis, Union City, CA). Normal mouse
chow (TekLad 57001) and High Fat Diet (HFD) (TD. 06414) was purchased from
Envigo (Indianapolis, Indiana, USA).
Animals
Male ND-4 mice were obtained from Envigo (Indianapolis, Indiana, USA) at 5 weeks of
age and 1820 g body weight; housed in micro isolator cages with corn cob bedding
with 12 h light/12 h dark cycle at 72F (22 C) with 3550% relative humidity. Mice
were fed on laboratory chow and High Fat Diet (HFD) and water ad libitum as per the
protocol. All animal studies were approved by the Institutional Animal Care and Use
Committee (IACUC), University of Mississippi, USA.
Preparation of LPS and Raspberry ketone
Lipopolysaccharide (6 mg/kg in saline). RK was dissolved in propylene glycol and water
(1:1) to prepare doses of 165 mg/kg, 330 mg/kg and 500 mg/kg body weight.
Treatment protocol and study design
Experiment # 1
Mice were divided into five groups (n¼47) as shown in Table 1. Group I was
fed a normal diet and served as the control. Groups II to V were fed HFD for ten
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Table 1. Table showing experimental regimen for different dosing groups of HFD fed mice.
Mice ND4 Diet up to 10 weeks Treatments Day 1 onwards Day 10
th
after RK treatment
Group 1 (n55) Normal Diet Water Animals were sacrificed
Group 2 (n55) HFD Vehicle (Oral)
Group 3 (n56) HFD RK (500 mg/kg) (Oral)
Group 4 (n57) HFD RK (330 mg/kg) (Oral)
Group 5 (n54) HFD RK (165 mg/kg) (Oral)
JOURNAL OF DIETARY SUPPLEMENTS 3
weeks. HFD was continued throughout the experimental procedure as shown in
Table 1. Thereafter, group II was administered vehicle (propylene glycol and water
1:1) orally for 10 days and served as negative control (Obese group), Groups III, IV
and V were given different doses (500 mg/kg, 330 mg/kg, and 165 mg/kg, respect-
ively) of RK for 10 days. On day 10, one hour after RK administration, all mice
were sacrificed. Body weights before and after RK treatment were recorded. ALT
and fasting blood glucose levels were determined (Relion confirm glucometer) at the
end of the experiment. Mortality of mice throughout the dosing period
was recorded.
Experiment # 2
Mice (ND4) were divided into 4 groups (n¼35) (Table 2). Group I was fed on nor-
mal diet, and served as the control group. Mice in groups II, III, and IV were given
HFD for 10 weeks. Thereafter, group II mice were given vehicle only. Group III and
group IV mice were given single i.p injection of LPS (6 mg/kg) on day 0 (Hammad
et al. 2013). Group IV mice were given RK at a dose of 500 mg/kg body weight,
orally from day 1 to day 10. After 10
th
dose of RK, blood was collected by cardiac
puncture. Fasting blood glucose and ALT levels were determined at the end of the
experiment. Body weights and animal mortality were recorded throughout the dos-
ing period.
Histopathology
The liver tissues were fixed in 10% buffered formalin for at least 24 h, thereafter, the
specimens were dehydrated in ascending grades of ethanol, cleared in xylene, and
embedded in paraffin wax. Blocks were made and 5 lm thick sections were cut. The
paraffin embedded liver tissue sections were deparaffinized using xylene and ethanol.
The slides were washed with phosphate buffered saline (PBS). The sections were stained
with hematoxylin and eosin (H & E). These stained liver sections were observed under
light microscope at 40magnifications to investigate the histological architecture of
the liver.
Statistical analysis
The data from individual groups were presented as the mean ± SEM. Differences
between groups were analyzed using analysis of variance (ANOVA) followed by Tukeys
comparisons test and minimum criterion for statistical significance was set at p<0.05
for all comparisons.
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Table 2. Table showing dosing pattern of raspberry ketone and LPS in HFD fed mice.
Mice ND4 Diet up to 10 weeks Treatments Day 0 Day 1 onwards Day 10
th
After RK treatment
Group I (n55) Normal diet Saline (i.p) Water Animals were sacrificed
Group II (n55) HFD Saline (i.p) Vehicle (Oral)
Group III (n53) HFD LPS (6 mg/kg; i.p) Vehicle (Oral)
Group IV (n54) HFD LPS (6 mg/kg; i.p) RK (500 mg/kg) (Oral)
4 T. M. MIR ET AL.
Results
Experiment # 1
Effect of RK on body weights
Administration of RK (500 and 165 mg/kg) did not cause any appreciable reduction in
the body weight of obese mice. Treatment of HFD fed mouse with RK (330 mg/kg)
resulted in a reduced body weight as compared with the normal control and obese
mice. Normal control animals exhibited an almost linear increase in body weight over
time. (Figure 1). Values represent mean ± SEM
Effect of RK on daily mortality of animals
Figure 2 represents the daily mortality of animals treated with RK (500 mg/kg, 330 mg/
kg and 165 mg/kg). In the present study, it was observed that 5 out of 10 mice died
within 24 h after administration of the first dose of RK (500 mg/kg). After three doses,
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Figure 1. Effect of Raspberry ketone treatment on Body weight of Mice. Normal Control: Mice group
fed on normal diet. HFD: High Fat Diet. Vehicle: propylene glycol and water (1:1); RK 500, RK 330,
and RK 165: Raspberry ketone at doses 500 mg/kg, 330 mg/kg and 165 mg/kg, respectively. No signifi-
cant change in body weight gain were observed in mice treated with RK 500 and RK 330mg/kg. A
slight reduction in body weight gain was observed in mice group treated with RK 165 mg/kg.
Figure 2. Daily mortality observed in mice treated with Raspberry ketone at doses 500, 330, and
165 mg/kg orally for 10days. Normal Control: Mice group fed on normal diet. HFD: High Fat Diet.
Vehicle: propylene glycol and water (1:1); RK 500, RK 330, and RK 165: Raspberry ketone at doses
500 mg/kg, 330 mg/kg and 165 mg/kg, respectively.
JOURNAL OF DIETARY SUPPLEMENTS 5
another mortality occurred in this group. RK treatment of mice at a dose of 330 mg/kg,
caused mortality in 4 out of 10 mice after the first dose; and another mouse died after
administration of the third dose. No mortality was observed in any of the animals
treated with RK (165 mg/kg), normal control and vehicle control animals.
Effect of RK treatment on cumulative deaths of mice
Obese mice treated with RK at doses of 500 mg/kg and 330 mg/kg showed mortality of
67.6 and 50%, respectively. While as RK (165 mg/kg), normal control and vehicle con-
trol mice showed no mortality (Figure 3).
Effect of RK on blood glucose levels
Figure 4 shows the blood glucose levels of animals before and after RK (500 mg/kg,
330 mg/kg and 165 mg/kg) treatments. We observed that mice fed with HFD and treated
with RK (500 mg/kg (p>0.05), 330 mg/kg (p>0.001) and 165 mg/kg (p>0.01) showed
elevated levels of blood glucose levels after RK treatment. While mice in the normal
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Figure 3. Percent cumulative deaths observed in mice groups treated with Raspberry ketone at doses
500, 330, and 165 mg/kg orally for 10days. Normal Control: Mice group fed on normal diet. HFD:
High Fat Diet. Vehicle: propylene glycol and water (1:1); RK 500, RK 330, and RK 165: Raspberry
ketone at doses 500 mg/kg, 330 mg/kg and 165 mg/kg, respectively.
Figure 4. Effect of Raspberry ketone treatment on blood glucose levels. Normal Control: Mice group
fed on normal diet. HFD: High Fat Diet. Vehicle: propylene glycol and water (1:1); RK 500, RK 330,
and RK 165: Raspberry ketone at doses 500 mg/kg, 330 mg/kg, and 165 mg/kg, respectively.
6 T. M. MIR ET AL.
and vehicle control groups did not show any significant elevations in blood glu-
cose levels.
Effect of RK treatment on blood ALT levels
Mice treated with RK at doses of 500 and 330 mg/kg showed significant (p<0.01 and
p<0.05, respectively) elevation in blood ALT levels as compared to vehicle control
group. While the mice treated with RK (165 mg/kg) group showed ALT levels in the
normal range (Figure 5). Values are shown as Mean ± SEM.
Effect of RK on histopathological morphology of liver
Histopathological architecture of the liver samples from normal and HFD treated
groups exhibit fatty changes compared with each other. Liver micrographs from the
normal group mice showed normal cellular architecture and did not exhibit any vacu-
olar changes, whereas group II animals (HFD treated) showed occurrence of hepatocel-
lular vacuolar change and steatosis that was consistent with fatty liver alterations
(mostly microvesicular steatosis occasional with macrovesicular steatosis or lipidosis)
(Figure 6B). RK treatments showed apparent reduction in the size of fat vacuoles
(Figure 6).
Experiment # 2
Effect of RK on body weight of mice under health compromised condition
Under health compromised conditions, the 500 mg/kg dose of RK did not affect the
body weight of mice as compared to the vehicle control group. Both vehicle control
group and RK þLPS showed a similar trend in body weights of mice. Mice in the nor-
mal control group exhibited a linear increase in body weight over time (Figure 7).
Values are shown as Mean ± SEM.
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Figure 5. Effect of Raspberry ketone treatment on blood ALT levels. ALT: Alanine transaminase.
Normal Control: Mice group fed on normal diet. HFD: High Fat Diet. Vehicle: propylene glycol and
water (1:1); RK 500, RK 330, and RK 165: Raspberry ketone at doses 500 mg/kg, 330mg/kg, and
165 mg/kg, respectively.
JOURNAL OF DIETARY SUPPLEMENTS 7
Effect of RK on daily mortality of animals under health compromised conditions
No mortalities were observed in normal and vehicle control groups of mice. However,
in only LPS treated group (single i.p injection of LPS on day 0), one out of three mice
died on day 3 post LPS injection. Whereas, in HFD þLPS þRK(500 mg/kg) and
HFD þRK(500 mg/kg) groups two out of four mice and three out of six mice died on
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Figure 6. Effect of RK on HFD induced Liver histopathology changes. Representative photomicro-
graphs (magnification 40). Normal Control: Mice group fed on normal diet. HFD: High Fat Diet.
Vehicle: propylene glycol and water (1:1); RK 500, RK 330, and RK 165: Raspberry ketone at doses
500 mg/kg, 330 mg/kg, and 165mg/kg, respectively. (A) Normal diet (Group I), (B) HFD þVehicle
(Group II), (C) HFD þRK-500 (Group III), (D) HFD þRK-330 (Group IV), and (E) HFD þRK-165 (Group V).
(Black arrow: microvesicles; yellow arrow: macrovesicles).
Figure 7. Effect of RK on body weight of mice under health compromised condition. Normal control:
Mice group fed on normal diet. HFD: High fat diet. Vehicle: propylene glycol and water (1:1); LPS:
Lipopolysaccharide; RK 500: Raspberry ketone at dose 500 mg/kg.
8 T. M. MIR ET AL.
day 1 of the study, respectively. On days 2 and three of the study, one mouse from
each group (HFD þLPS þRK-500 mg/kg and HFD þRK-500 mg/kg) died (Figure 8).
Effect of RK treatment on cumulative deaths of mice under health compro-
mised conditions
Obese mice sensitized with LPS resulted in the 40% mortality, while mice treated with
HFD þLPS þRK-500 mg/kg and HFD þRK-500 mg/kg showed mortalities of 75% and
67%, respectively (Figure 9).
Effect of RK on blood glucose levels of mice under health compromised conditions
Figure 10, shows the blood glucose levels of mice treated with RK at a dose of 500 mg/
kg under health compromised condition. Increased blood glucose levels were observed
in the groups of mice given HFD þLPS or HFD þRK-500 mg/kg. As there was 75%,
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Figure 8. Daily mortality observed in obese mice treated with RK plus LPS. Normal control: Mice
group fed on normal diet. HFD: High fat diet. Vehicle: propylene glycol and water (1:1); LPS:
Lipopolysaccharide; RK 500: Raspberry ketone at dose 500 mg/kg.
Figure 9. Effect of RK treatment on cumulative deaths of mice under health compromised conditions.
High fat diet (HFD); Lipopolysaccharide (LPS); Raspberry ketone at dose 500mg/kg (RK 500).
JOURNAL OF DIETARY SUPPLEMENTS 9
mortality observed in group of mice treated with HFD þLPS þRK-500 mg/kg, blood
samples could not be collected from all animals, therefore glucose estimation was done
in the remaining mice in this group (Figure 10). Values are shown as Mean ± SEM
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Figure 10. Effect of RK on blood glucose levels of mice under health compromised conditions. High
fat diet (HFD); Lipopolysaccharide (LPS); Raspberry ketone at dose 500mg/kg (RK 500).
Figure 11. Effect of RK on HFD induced liver histopathology changes under health compromised con-
ditions. High fat diet (HFD); Lipopolysaccharide (LPS); Raspberry ketone at dose 500 mg/kg (RK 500).
Representative photomicrographs (magnification 40), CV: Central vein; micro and macro vesicles
(arrows). (A) Normal diet (Group I), (B) HFDþVehicle (Group II), (C) HFD þLPS (Group III), and (D)
HFD þLPS þRK-500 (Group IV).
10 T. M. MIR ET AL.
Effect of RK on histopathological morphology of liver under health compro-
mised conditions
Under health compromised conditions, it was observed that RK at high dose (500 mg/
kg) reduced the effects exhibited by HFD on liver (Figure 11A). Liver architecture of
mice treated with HFD showed accumulation of fat vesicles. The livers from mice
treated with HFD þLPS showed less degree of lipidosis (Figure 11B) as compared with
HFD group (Figure 11C).
Discussion
Fruits of Rubus idaeus L. commonly known as red raspberries are widely available
and possess both nutritional and medicinal properties. They possess several essential
micronutrients, dietary fibers, and phenolic components, especially raspberry ketone
ellagitannins, and anthocyanins. Scientific reports indicate that raspberries are being
used in the treatment of diabetes mellitus, cancer, cardiovascular disorders, obesity,
and neurodegeneration (Burton-Freeman et al. 2016; Kristo et al. 2016; Noratto et al.
2016). Raspberry ketone is an aromatic compound predominantly found in raspberries
and also in other fruits, including black raspberries and kiwifruit (Ulbricht et al.
2011). It is widely used as a fragrance in cosmetics and flavoring agent in foodstuff.
Several reports have demonstrated its lipolytic and anti-obesity activities (Morimoto
et al. 2005; Leu et al. 2018; Mehanna et al. 2018). In the present study, we induced
obesity in mice by feeding them for 10 weeks on a high-fat diet. Increases in body
weight and fat storage was noted in accordance with the previously published reports
(Morimoto et al. 2005; Mukai et al. 2016). In addition, we explored the possible
adverse effects of consumption of RK during normal obese and health compromised
obese condition in mice.
We observed a slight trend of decreased gain in body weights in mice treated with
RK at 330 mg/kg; however, this may be attributed to reduced intake of feed as has been
hypothesized by others (Cotten et al. 2017). At other doses (500 and 165 mg/kg), no
appreciable change in body weight was observed. At the higher dose (500 mg/kg), RK
may be toxic and cause deaths within the 2 days post RK treatment, reducing the num-
ber of animals for statistical comparison of body weights at the termination of the
experiment. At the lower dose (165 m g/kg), RK may not be as effective in lowering the
body weight gain especially under this short duration (10 days) of RK treatment. A simi-
lar pattern of deaths and reduction in body weights was observed in the LPS sensitized
obese mice treated with RK at 500 mg/kg.
Dietary fat is digested by pancreatic lipase and absorbed from the small intestine
(Hernell et al. 1990). RK decreased the body weight and this may be due to inhibition
of pancreatic lipase enzyme activity and inhibition of intestinal absorption. Thus, the
weight lowering effect observed in this study conforms with other reports that demon-
strated weight reducing property of RK (Morimoto et al. 2005; Park 2010; Lopez et al.
2013). However, the retardation in weight gain during the 10-day study could also be
due to loss of appetite in the groups given LPS and RK. This point needs to be further
evaluated as in this study food intake was not measured.
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JOURNAL OF DIETARY SUPPLEMENTS 11
In the present study, our major objective was to assess the potential adverse effects
that may be associated with RK, if it was consumed at high doses or by individuals
with health compromised conditions. The LPS sensitized mouse (health compromised)
model used in this study has been used in other studies to demonstrate the hepatotoxic
potential of natural products (Abdel-Bakky et al. 2010; Saleh et al. 2013) as well as
pharmaceutical drugs (Shaw et al. 2007; Poulsen et al. 2014). The results of the current
preliminary study showed dose-related mortality in obese mice treated with RK. In add-
ition, LPS sensitized obese mice treated with high dose of RK showed higher mortality;
however, no mortality was observed in mice treated with RK at 165 mg/kg. These results
indicate the narrow spectrum of effective weight reducing dose of RK. Alanine amino-
transferase is a cytoplasmic enzyme of hepatocytes, and its elevation in blood is consid-
ered one of the key indicators of liver injury (McGill and Jaeschke 2013). During any
hepatotoxic insult, this enzyme is released from the hepatocytes into the blood stream
resulting in its higher levels in the blood (Kim et al. 2008). We have also observed sig-
nificant (p<0.05 and p<0.01 at 330 and 500 mg/kg, respectively) elevation of blood
ALT levels in mice treated with higher doses of RK as compared to those fed on normal
or high fat diet only. Our results are similar and support the ones reported earlier by
Joint FAO/WHO Expert Committee on Food Additives (JECFA) report. (study by
Hoffman 2004 cited from Joint FAO WHO Expert Committee on Food Additives,
World Health Organization 2011) (Hoffman 2004; Joint FAO WHO Expert Committee
on Food Additives, World Health Organization 2011). At lower doses, a rising trend in
the blood ALT levels was observed; however, it was not statistically significant, perhaps,
due to short duration of the study. Therefore, further studies of longer duration should
be undertaken to assess the toxic and safety potential of RK in both obese and health
compromised obese mice.
In this study, we observed a significant increase in blood glucose levels in mice
treated with different doses of RK compared to those of normal control and obese
mice. Even in health-compromised obese mice, elevated blood glucose levels were
observed. None of the published work on RK has earlier reported this observation. This
is the first report on elevated blood glucose level by RK treatment in obese and health
compromised obese mice. This increase in blood glucose levels implies RKs indirect
effect on glucose metabolism, as RK is involved in lipolysis and fatty acid oxidation in
the cells (Park 2010; Tsai et al. 2017), resulting in the production of ketone bodies that
are used as a source of fuel in the cells instead of glucose. This may result in the non-
utilization of glucose by cells resulting in higher glucose levels in blood (Collier
et al. 1993).
The extent of lipid accumulation in liver parenchymal cells is the direct indices of
liver steatosis (Mezey 1999). Several reports indicate HFD administration to mice over a
period, resulted in fat accumulation in the liver (Choi et al. 2018; Kim et al. 2016) lead-
ing to steatosis. To assess the effect of RK on hepatic steatosis, we performed H & E
staining. Histology revealed that liver from mice fed on HFD exhibited hepatic lipid
vesicles compared to none seen in mice fed on normal diet. Although, RK treatments
resulted in the reduction in the size of lipid vesicles in liver tissue, but it did not com-
pletely eliminate the effects of HFD in this short period of study. Similar results were
observed under health-compromised conditions.
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Conclusions
In conclusion, the results of the present study showed that while RK may have weight-
reducing properties at higher doses and longer durations, it also has the potential to be
hepatotoxic. Weight-reducing agents are often consumed for long durations to achieve
the desired level of effect. Therefore, long-term experiments are essential at more
physiologically relevant doses to determine a safe dose of RK, under obese and health
compromised-obese conditions.
Acknowledgments
The authors thank Dr. Jon F. Parcher for editing the manuscript for English language. The
authors sincerely thank Ms. Penny Bolton for assistance in maintenance of animals used in this
study and biological work.
Conflicts of interest
The authors declare no conflicts of interest.
Funding
This work was supported in part by the United States Department of Agriculture, Agricultural
Research Service, Specific Cooperative agreement 58-6408-1-603-04 and US Food and Drug
Administration 1U01FD004246-04.
Q3
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16 T. M. MIR ET AL.
... All of these functions likely contribute to the protective effect of metabolic disease. While, Hao et al. (2020) reported that RK could be toxicity to mice, because approximate of 43% mortality rate appeared after a single oral garage of 640 mg/kg RK. Mir et al. (2021) found that RK at doses 330 and 500 mg/kg resulted in 67.6 and 50% mortality, respectively in normal obese mice. Although this dose is much higher than the daily recommended dose (between 100 and 1400 mg per day) of commercial RK supplement, the safety of RK is still worthy of attention in consideration of previous reports on its appetite suppression. ...
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Novel food and food compounds interventions have attracted a lot of attention nowadays for the prevention and treatment of metabolic diseases. Raspberry ketone (RK) is aromatic compound found within red fruits and berries, has been used as an over-the-counter product for weight loss. However, actually, the effect of RK on weight loss is still controversial, and the mechanism is largely unknown. Besides, in vivo and in vitro studies have demonstrated the beneficial effect of RK on the development of other metabolic diseases. In this review, we comprehensively highlighted the synthesis, bioavailability, and metabolism of RK, and summarized the progress made in our understanding of the potential biological activities of RK, including antiobesity, antidiabetes, cardioprotection, and hepatoprotection, as well as their underlying mechanisms. This paper provides a critical overview about the current findings and proposes the future studies in the area of RK on human health. Practical applications Raspberry ketone (RK) has been used for weight control for years, but this effect is controversial considering food intake. Additionally, RK is beneficial for T2DM, liver and heart injury. The underlying mechanisms of the protective effect of RK including accelerating fatty acid oxidation, balancing serum glucose level, anti-inflammation, antioxidant process, and so on. In this context, we provide a comprehensive analysis of the benefits of RK against many metabolic diseases and discuss the underlying molecular mechanisms. We hope our work will be helpful for further researches on RK and improve its public recognition.
... Higher doses of RK at 500 mg/kg indicated high levels of blood glucose and ALT. This study even indicated that RK can cause adverse effects in health compromised conditions and high doses may pose a potential risk [56]. ...
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Raspberry ketone (RK) is an aromatic phenolic compound naturally occurring in red raspberries, kiwifruit, peaches, and apples and reported for its potential therapeutic and nutraceutical properties. Studies in cells and rodents have suggested an important role for RK in hepatic/cardio/gastric protection and as an anti-hyperlipidemic, anti-obesity, depigmentation, and sexual maturation agent. Raspberry ketone-mediated activation of peroxisome proliferator-activated receptor-α (PPAR-α) stands out as one of its main modes of action. Although rodent studies have demonstrated the efficacious effects of RK, its mechanism remains largely unknown. In spite of a lack of reliable human research, RK is marketed as a health supplement, at very high doses. In this review, we provide a compilation of scientific research that has been conducted so far, assessing the therapeutic properties of RK in several disease conditions as well as inspiring future research before RK can be considered safe and efficacious with limited side effects as an alternative to modern medicines in the treatment of major lifestyle-based diseases.
... In addition, raspberry ketone is utilized as an attractant in insect baits (Perez 1983;Metcalf et al. 1983;Deifel 1989) and as a component in perfumes (Dumont et al. 1996;Farwick et al. 2019), whereas further applications in cosmetic industry, e.g., as skin whitening or hair growth inducing agent, remain controversial (Harada et al. 2008;Kim et al. 2016). Other publications promote dose-dependent health benefits with regard to an anti-obese effect (Morimoto et al. 2005;Park 2015;Tsai et al. 2017;Zhao et al. 2019;Mir et al. 2019) or suggest medical applications due to its antioxidant and anti-inflammatory potential (Parmar and Tripathi 1991;Khan et al. 2018;Fouad et al. 2019;Mohamed et al. 2020;Hamdy et al. 2020). ...
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Raspberry ketone is a widely used flavor compound in food and cosmetic industry. Several processes for its biocatalytic production have already been described, but either with the use of genetically modified organisms (GMOs) or incomplete conversion of the variety of precursors that are available in nature. Such natural precursors are rhododendrol glycosides with different proportions of ( R )- and ( S )-rhododendrol depending on the origin. After hydrolysis of these rhododendrol glycosides, the formed rhododendrol enantiomers have to be oxidized to obtain the final product raspberry ketone. To be able to achieve a high conversion with different starting material, we assembled an alcohol dehydrogenase toolbox that can be accessed depending on the optical purity of the intermediate rhododendrol. This is demonstrated by converting racemic rhododendrol using a combination of ( R )- and ( S )-selective alcohol dehydrogenases together with a universal cofactor recycling system. Furthermore, we conducted a biocatalytic cascade reaction starting from naturally derived rhododendrol glycosides by the use of a glucosidase and an alcohol dehydrogenase to produce raspberry ketone in high yield. Key points • LB-ADH, LK-ADH and LS-ADH oxidize (R)-rhododendrol • RR-ADH and ADH1E oxidize (S)-rhododendrol • Raspberry ketone production via glucosidase and alcohol dehydrogenases from a toolbox Graphical abstract
... Raspberry ketone (4-(4-hydroxyphenyl) butan-2-one; RK) is one of the major natural aromatic compounds of red raspberry fruit (Rubus idaeus), responsible for the fruity aroma of raspberries (Giuggioli et al. 2015, Mir et al. 2019. It is used as a fragrance and flavoring substance for cosmetics, perfumes, foodstuffs, and soft drinks (Schwab et al. 2018). ...
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The current investigation was accomplished to evaluate the hepatoprotective effect of White tea and Raspberry Ketone against toxicity induced by acrylamide in rats. Sixty adult male rats were divided randomly into group (I) control; group (II) rats received RK with dose (6 mg/kg/day); Group III: rats received 5 ml of WT extract/kg/day; Group IV rats received AA (5 mg/kg/day); Group V: rats administrated with both AA (5 mg/kg/day) and RK (6 mg/kg/day) and Group VI: rats administrated AA (5 mg/kg/day) and 5 ml of WT extract/kg/day. The biochemical assays exhibited a significant increase in serum levels of Adiponectin, AST, ALT, ALP of the group treated with acrylamide if compared to the control group and an improvement in their levels of groups V and VI. The histopathological and immunohistochemical findings confirm the biochemical observations. In conclusion, the present investigation proved that the supplementation of WT and RK enhanced the liver histology, immunohistochemistry and biochemistry against the oxidative stress induced by acrylamide.
... Raspberry ketone (4-(4-hydroxyphenyl) butan-2-one; RK) is one of the major natural aromatic compounds of red raspberry fruit (Rubus idaeus), responsible for the fruity aroma of raspberries (Giuggioli et al. 2015, Mir et al. 2019. It is used as a fragrance and flavoring substance for cosmetics, perfumes, foodstuffs, and soft drinks (Schwab et al. 2018). ...
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Background Dietary supplements are used by more than half of adults, although to our knowledge, the reasons motivating use have not been previously examined in US adults using nationally representative data. The purpose of this analysis was to examine motivations for dietary supplement use, characterize the types of products used for the most commonly reported motivations, and to examine the role of physicians and health care practitioners in guiding choices about dietary supplements. Methods Data from adults (≥20 years; n = 11 956) were examined in the 2007-2010 National Health and Nutrition Examination Survey, a nationally representative, cross-sectional, population-based survey. Results The most commonly reported reasons for using supplements were to “improve” (45%) or “maintain” (33%) overall health. Women used calcium products for “bone health” (36%), whereas men were more likely to report supplement use for “heart health or to lower cholesterol” (18%). Older adults (≥60 years) were more likely than younger individuals to report motivations related to site-specific reasons like heart, bone and joint, and eye health. Only 23% of products were used based on recommendations of a health care provider. Multivitamin-mineral products were the most frequently reported type of supplement taken, followed by calcium and ω-3 or fish oil supplements. Supplement users are more likely to report very good or excellent health, have health insurance, use alcohol moderately, eschew cigarette smoking, and exercise more frequently than nonusers. Conclusions Supplement users reported motivations related to overall health more commonly than for supplementing nutrients from food intakes. Use of supplements was related to more favorable health and lifestyle choices. Less than a quarter of supplements used by adults were recommended by a physician or health care provider.
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Obesity constitutes a major worldwide problem in which hyperlipidemia and insulin resistance represents adverse metabolic consequences of it. The present study was conducted to elucidate the role of raspberry ketones (RKs) in controlling body weight gain, hyperlipidemia and insulin resistance in male obese rats through affecting the expression of various adipocytokines. As Aquaporin-7 is co-related with the expression of various adipocytokines and has recently emerged as a modulator of adipocyte metabolism, the present study evaluated the effect of RKs on adipose tissue expression of aquaporin-7(AQP7) in high-fat (HF) diet-fed rats. Groups of male rats were assigned to normal, HF diet-fed control rats and RKs-treated (250 and 500mg/kg) groups. RKs administration effectively abrogated hyperlipidemia and oxidative burden and enhanced insulin sensitivity. In addition, treatment with RKs ameliorated adipose tissue and liver indices and the reduced adipocyte diameters. Moreover, administration of the low dose of RKs ameliorated the expression of apelin and its receptor, and visfatin with upregulating adiponectin expression compared to HF diet control rats. However, both doses effectively downregulated leptin expression. It was obvious that both RKs doses revealed effectiveness in upregulating the AQP7 expression. The present data suggest the promising therapeutic role of RKs in HF diet-induced obesity that is likely attributable, at least in part, to upregulation of AQP7 expression.
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Aim: The cardioprotective role of raspberry ketone (RK) against isoproterenol (ISO)-induced myocardial infarction (MI) in rats was assessed. Materials and methods: Rats were randomly divided into Group I - Vehicle control; Group II - Toxic control ISO (85mg/kg, s.c.); Group III, IV and V - RK (50, 100 and 200mg/kg, respectively) with ISO; Group VI- RK (200mg/kg) alone; Group VII - Propranolol (10mg/kg) with ISO; and Group VIII - Propranolol (10mg/kg) alone. After twenty-four hours of the last dose, animals were sacrificed and creatine kinase-MB, lactate dehydrogenase, total cholesterol, triglycerides, high-density-lipoprotein, low-density-lipoprotein, very-low-density-lipoprotein, malondialdehyde, reduced glutathione, superoxide dismutase, catalase, Na+, K+-ATPase, nitric oxide, histopathological and immunohistochemical analysis (tumor necrosis factor-α and inducible nitric oxide synthase) were performed. Key findings: Treatment with ISO significantly deviate the biochemical parameters from the normal levels, which was considerably restored by RK at 100 and 200mg/kg doses. 50mg/kg dose, however, did not demonstrate any significant cardioprotective action. The histopathological and immunohistochemical analysis further substantiated these findings. Significance: Our study showed a dose-dependent reduction in oxidative stress, inflammation and dyslipidemia by RK in ISO-intoxicated rats, which signifies that RK from the European red raspberry plant might be a valuable entity for the management of MI.
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Background: Obesity is caused by excessive accumulation of body fat and is closely related to complex metabolic diseases. Raspberry ketone (RK), a major aromatic compound in red raspberry, was recently reported to possess anti-obesity effects. However, its mechanisms are unclear. Aim: Adipogenesis plays a critical role in obesity and, therefore, this study aimed to investigate the effect and mechanisms of action of RK on adipogenesis in 3T3-L1 preadipocytes. Materials and methods: 3T3-L1 preadipocytes were differentiated in medium containing insulin, dexamethasone, and 1-methyl-3-isobutylxanthine. Adipocyte lipid contents were determined using oil-red O staining while adipogenic transcription factor and lipogenic protein expressions were determined using western blotting. Results: RK (300-400µM) strongly inhibited lipid accumulation during 3T3-L1 preadipocyte differentiation into adipocytes. RK reduced the CCAAT/enhancer-binding protein-α (C/EBP-α), peroxisome proliferation-activated receptor-γ (PPAR-γ), fatty acid synthase (FAS), and fatty acid-binding protein 4 (FABP4) expressions and increased heme oxygenase-1 (HO-1), Wnt10b, and β-catenin expressions in 3T3-L1 adipocytes. Additionally, RK inhibited lipid accumulation, and adipogenic transcription factor and lipogenic protein expressions were all decreased by inhibiting HO-1 or β-catenin using tin protoporphyrin (SnPP) or β-catenin short-interfering RNA (siRNA), respectively. Furthermore, Wnt10b and β-catenin expressions were negatively regulation by SnPP. Conclusion: RK may exert anti-adipogenic effects through modulation of the HO-1/Wnt/beta-catenin signaling pathway.