ArticlePDF Available

Effect of hydroxycitric acid on serotonin release from isolated rat brain cortex

Authors:

Abstract and Figures

There is evidence that hydroxycitric acid (HCA), an extract of dried fruit rind of South Asian trees of the genus Garcinia cambogia, can reduce food intake in experimental animals. In the present study, we investigated the effect of HCA on basal and potassium-depolarization evoked increase in radiolabeled serotonin ([3H]-5-HT) release from rat brain cortex slices in vitro. HCA (10 microM-1 mM) altered the baseline of spontaneous tritium efflux but had no significant effect on potassium-evoked release of [3H]-5-HT. When applied on its own, HCA (10 microM-1 mM) elicited a concentration-dependent increase in efflux of [3H]-5-HT reaching a maximum at 300 microM. We conclude that HCA can increase the release of radiolabeled 5-HT from the isolated rat brain cortex.
Content may be subject to copyright.
89
Molecular and Cellular Biochemistry 238: 89–103, 2002.
© 2002 Kluwer Academic Publishers. Printed in the Netherlands.
Safety and mechanism of appetite suppression by a
novel hydroxycitric acid extract (HCA-SX)
Sunny E. Ohia, Catherine A. Opere, Angela M. LeDay, Manashi
Bagchi, Debasis Bagchi and Sidney J. Stohs
Department of Pharmacy Sciences, Creighton University School of Pharmacy and Allied Health Professions, Omaha,
NE, USA
Received 15 January 2002; accepted 16 April 2002
Abstract
A growing body of evidence demonstrates the efficacy of Garcinia cambogia-derived natural (–)-hydroxycitric acid (HCA) in
weight management by curbing appetite and inhibiting body fat biosynthesis. However, the exact mechanism of action of this
novel phytopharmaceutical has yet to be fully understood. In a previous study, we showed that in the rat brain cortex a novel
HCA extract (HCA-SX, Super CitriMaxTM ) increases the release/availability of radiolabeled 5-hydroxytryptamine or serotonin
([3H]-5-HT), a neurotransmitter implicated in the regulation of eating behavior and appetite control. The aim of the present study
was 2-fold: (a) to determine the effect of HCA-SX on 5-HT uptake in rat brain cortex in vitro; and (b) to evaluate the safety of
HCA-SX in vivo. Isolated rat brain cortex slices were incubated in oxygenated Krebs solution for 20 min and transferred to buffer
solutions containing [3H]-5-HT for different time intervals. In some experiments, tissues were exposed to HCA-SX (10 µM – 1
mM) and the serotonin receptor reuptake inhibitors (SRRI) fluoxetine (100 µM) plus clomipramine (10 µM). Uptake of [3H]-5-HT
was expressed as d.p.m./mg wet weight. A time-dependent uptake of [3H]-5-HT occurred in cortical slices reaching a maximum at
60 min. HCA-SX, and fluoxetine plus clomipramine inhibited the time-dependent uptake of [3H]-5-HT. At 90 min, HCA-SX (300
µM) caused a 20% decrease, whereas fluoxetine plus clomipramine inhibited [3H]-5-HT uptake by 30%. In safety studies, acute
oral toxicity, acute dermal toxicity, primary dermal irritation and primary eye irritation, were conducted in animals using various
doses of HCA-SX. Results indicate that the LD50 of HCA-SX is greater than 5,000 mg/kg when administered once orally via
gastric intubation to fasted male and female Albino rats. No gross toxicological findings were observed under the experimental
conditions. Taken together, these in vivo toxicological studies demonstrate that HCA-SX is a safe, natural supplement under the
conditions it was tested. Furthermore, HCA-SX can inhibit [3H]-5-HT uptake (and also increase 5-HT availability) in isolated rat
brain cortical slices in a manner similar to that of SRRIs, and thus may prove beneficial in controlling appetite, as well as treat-
ment of depression, insomnia, migraine headaches and other serotonin-deficient conditions. (Mol Cell Biochem 238: 89–
103, 2002)
Key words:Garcinia cambogia, (–)-Hydroxycitric acid, fluoxetine, clomipramine, rat brain cortex, serotonin release, appe-
tite, acute oral toxicity, acute dermal toxicity, primary dermal irritation, primary eye irritation
Introduction
(–)-Hydroxycitric acid (HCA), a popular dietary supplement
for weight loss, is a natural extract from the dried fruit rind of
Garcinia cambogia (family Guttiferae), a tree that is native
to southeast Asia. The dried fruit rind is also known as Mala-
bar tamarind, which is extensively used in southern India for
culinary purposes [1]. The fruit exhibits a distinctive sweet
and sour taste, and has been used for centuries with no harm-
ful effects in Asian countries for preparing culinary dishes. It
has been observed that when the dried fruit rind is sprinkled
on curries it makes the food more filling and satisfying [1, 2].
Address for offprints: S.E. Ohia, Creighton University School of Pharmacy and Allied Health Professions, 2500 California Plaza, Omaha, NE
68178, USA (E-mail: seohia@creighton.edu)
90
HCA has been shown to be a competitive inhibitor of ATP-
citrate lyase, the enzyme catalyzing the extramitochondrial
cleavage of citrate to oxaloacetate and acetyl-CoA. This en-
zyme is important in maintaining the acetyl-CoA pool for fatty
acid and cholesterol biosynthesis, particularly during the
hyperlipogenic nutritional state produced by high carbohy-
drate feeding. One study has shown that HCA administered
before meals causes weight loss in obese subjects [2] and
improves energy metabolism. HCA reduces food consump-
tion in humans by possibly diverting carbohydrates and fatty
acids that would have become fat in the liver into hepatic gly-
cogen. This metabolic change may send a signal to the brain
that results in reduced food intake, curbs the appetite, and
the desire for sweets is also eliminated [3, 4]. Research has
shown that HCA-SX inhibits fat production and decreases
body weight in animals and humans [5]. No toxicity, either
acute or chronic at any reasonable level of consumption has
been observed.
Recent studies have demonstrated that oral HCA supple-
mentation (as Super CitriMaxTM, a calcium/potassium salt of
60% HCA, which is tasteless, odorless and highly water solu-
ble, HCA-SX) is highly bioavailable in human plasma as
demonstrated by a gas chromatography-mass spectromet-
ric method [6]. HCA-SX remained in the blood for more than
4–9 h after oral ingestion. These investigators also demon-
strated that eating a meal shortly after taking HCA-SX reduced
food absorption by about 60% [6].
HCA-SX has been shown to increase the release/availability
of [3H]-5-hydroxytryptamine (serotonin; HT) from rat brain
cortical slices in vitro [7]. Since 5-HT has been implicated in
the regulation of eating behavior and body weight regulation
[8–10], it is reasonable to suggest that a mechanism of appe-
tite suppression induced by HCA-SX could be mediated by
this neurotransmitter. Indeed, stimulants of this monoamine
reduced food intake and weight gain and increased energy
expenditure in both experimental animals and humans [8].
The objective of the present study was 2-fold, (a) to fur-
ther investigate the effects of HCA-SX on 5-HT metabolism
by examining the action of this novel appetite suppressant
on the uptake of this monoamine in isolated rat brain cortex
in vitro, and (b) to determine the safety profile of HCA-SX.
For the latter, we conducted acute oral toxicity, acute dermal
toxicity, primary dermal irritation and primary eye irritation
studies in in vivo models.
Materials and methods
Chemicals
A natural, highly water soluble, calcium/potassium salt of 60%
HCA extract from Garcinia cambogia commercially known as
Super CitriMaxTM HCA-SXS (HCA-SX) was obtained from
InterHealth Nutraceuticals, Benicia, CA, USA. [3H]-5-HT was
purchased from NEN Life Sciences, Boston, MA, USA. Flu-
oxetine and clomipramine were obtained from Sigma-Aldrich,
Natick, MA, USA. All other chemicals and reagents were ob-
tained from Sigma Chemicals (St. Louis, MO, USA) and were
of analytical grade or the highest grade available.
Animals and treatment for 5-HT uptake study
Male Sprague–Dawley rats (weighing 150–200 g) were ob-
tained from Charles River Breeding Laboratories (Portage, MI,
USA). Animals were maintained and used in accordance with
the current National Institute of Health Guidelines and the
ARVO Resolution on the Use of Animals in Research.
The effect of HCA-SX on radiolabeled 5-HT (serotonin)
uptake was measured as described by Kirksey and Slotkin [11]
and Goodlet et al. [12] with some modifications. Briefly, Male
Sprague–Dawley rats were sacrificed by asphyxiation and the
whole brain was quickly dissected and placed in ice-cold sa-
line solution (0.9% w/v NaC1). The cortex was carefully re-
moved from ice-cold saline solution and cut into thin slices
of about 15–30 mg weight using surgical blades and scissors.
Although tissues obtained differed slightly in size, the results
are presented as d.p.m./mg wet weight in order to normalize
the data. Cortex slices were equilibrated in 2 ml of Krebs buffer
solution at 37°C under an atmosphere of 95% O2 and 5% CO2
for 20 min. The Krebs buffer solution was composed of the
following (millimolar); sodium chloride, 118; potassium chlo-
ride, 4.8; calcium chloride, 1.3; potassium dihydrogen phos-
phate, 1.2; sodium bicarbonate, 25; magnesium sulfate, 2.0;
and dextrose, 10 (pH 7.4). After equilibration, cortex slices were
incubated in Krebs buffer solution (gassed with 95% O2 and
5% CO2) at 37°C, containing 80 nM of [3H]-5-HT and pargyline
(10–5 M). During incubation, two slices were removed at times
0, 5, 10, 20, 25, 30, 60, and 90 min and rinsed in 10 ml ice-cold
Krebs buffer solution to stop uptake of [3H]-5-HT into neurons.
Each slice was blotted dry with Whatman paper #1, weighed
and digested in 1 ml NaOH (1 N) at 60°C for 20 min. Each sample
was combined with 12 ml of aqueous scintillation cocktail (Eco-
lume; ICN Radiochemicals, CA, USA), and analyzed for radio-
activity by liquid scintillation spectrometry. The amount of
[3H]-5-HT present in samples was expressed as d.p.m./mg wet
weight. Test drugs were present in the Krebs buffer solution
during the pre-incubation and incubation periods.
Animals and treatment for acute oral toxicity study
The objective was to determine the acute oral median lethal
dose and evaluate the potential systemic toxicity of HCA-SX
when administered as a single dose to Albino rats. The pro-
tocol was designed and the study was conducted in compli-
91
ance with the Environmental Protection Agency Guidelines for
Registering Industrial Chemicals in the US (Pesticide Assess-
ment Guidelines, Subdivision F, Hazard Evaluation: Human
and Domestic Animals Section 81-1), the Toxic Substances
Control Act (TSCA) Health Effects Test Guidelines, 40 CFR
798.1175.
Male and female Albino rats (weighing 208–216 g at study
initiation) were obtained from Charles River Breeding Labo-
ratories (Portage, MI, USA) and were allowed free access to
lab chow (Purina Certified Rodent Chow, No. 5002, St. Louis,
MO, USA) and municipal water ad libitum. Animals were ac-
climated to laboratory conditions for a minimum of 7 days prior
to initiation of dosing. The animal room was kept at a control-
led temperature (69–76°F), humidity (30–82%), and light (12 h
light/12 h dark).
The amount of HCA-SX administration was based on body
weights taken just prior to dosing using a dose volume of 10
ml/kg. HCA-SX was administered orally via gastric intubation
with a French rubber feeding tube (14-gauge), which was af-
fixed to a 3-ml syringe. The rats were fasted approximately 18–
20 h prior to dosing and returned to feeding 3–4 h after dosing.
One group of five male and one group of five female rats were
administered a single dose at a level of 5000 mg/kg. Control
animals received the vehicle.
The rats were observed for mortality at approximately 1, 3
and 4 h post-treatment on day 0 and twice daily (morning
and afternoon) thereafter for 14 days. For clinical observa-
tions, the rats were observed at approximately 1, 3 and 4 h
post-treatment on day 0 and once daily thereafter for 14
days. Body weights were obtained and recorded on study
days –1, 0 (initiation), 7 and 14 (study termination). Upon
study termination, all rats were euthanized by carbon diox-
ide asphyxiation. At the terminal necropsy, the major organ
systems of the cranial, thoracic and abdominal cavities were
examined for all animals.
Animals and treatment for acute dermal toxicity study
The objectives of this study was to determine the median le-
thal dose, evaluate the potential systemic toxicity and evalu-
ate the local irritative potential of HCA-SX when applied once
to the skin of Albino rabbits. The protocol was designed and
the study was conducted in compliance with the Environmen-
tal Protection Agency Guidelines for Registering Industrial
Chemicals in the US (Pesticide Assessment Guidelines, Sub-
division F, Hazard Evaluation: Human and Domestic Animals
Section 81-2), the Toxic Substances Control Act (TSCA) Health
Effects Test Guidelines 40 CFR 798.1100, and the Japanese
Agricultural Chemicals Laws and Regulations Testing Guide-
lines for Toxicology Studies published by the Society of
Agricultural Chemical Industry under the auspices of MAFF
(Ministry of Agriculture, Forestry and Fisheries).
Male and female New Zealand white Albino rabbits (weigh-
ing 2227–2466 g at study initiation) were obtained from Hazle-
ton Research Products (Denver, PA, USA) and allowed free
access to lab chow (Purina Certified Rodent Chow, No. 5322,
St. Louis, MO, USA) and municipal water ad libitum. Animals
were acclimated to laboratory conditions for a minimum of 6
days prior to initiation of dosing. The animal room was kept
at a controlled temperature (66–70°F), humidity (50–80%), and
light (12 h light/12 h dark).
Individual doses of the HCA-SX were calculated based on
body weight obtained just prior to dosing with a dose of 2000
mg/kg bodyweight. HCA-SX was applied directly to clipped,
unabraded skin. On the day prior to dosing, the hair was re-
moved from the backs of the rabbits using an Oster® small
animal clipper. Individual doses of HCA-SX moistened with
approximately 4.5 ml of deionized water, were applied to the
dorsal skin and covered approximately 15% of the total body
surface. Plastic restraint collars were applied and remained on
the rabbits for the duration of the exposure (24 h). Upon com-
pletion of the exposure period, the collars and bandages were
removed and the application sites were wiped with dispos-
able paper towels moistened with tepid water. There was one
group of five male Albino rabbits and one group of five fe-
male Albino rabbits that was dermally administered single
doses (24 h, semi-occluded exposure) of HCA-SX at a dose
level of 2000 mg/kg.
The rabbits were observed for mortality at approximately
1, 3 and 4 h post-treatment on day 0 and twice daily (morning
and evening) thereafter for 14 days. For clinical observations,
the rabbits were observed at approximately 1, 3 and 4 h post-
treatment on day 0 and once daily thereafter for 14 days. For
dermal observations, the application sites were examined for
erythema, edema (Table 1) and other dermal findings begin-
ning approximately 30–60 min after bandage removal and daily
thereafter for 13 days. The rabbits were shaved to facilitate
dermal observations on study days 3, 7, 10 and 14. The body
weights were obtained and recorded on days 0 (initiation), 7
and 14 (study termination). The rabbits were euthanized by
intravenous injection of sodium pentobarbital solution (150
mg/kg, i.v.) upon termination of the study. The major organ
systems of the cranial, thoracic and abdominal cavities were
examined for all animals.
Animals and treatment for primary dermal irritation study
The objective was to determine the irritative potential of HCA-
SX following a single exposure to the skin of Albino rabbits.
The protocol was designed and the study was conducted in
general compliance with the Environmental Protection Agency
Guidelines for Registering Industrial Chemicals in the US (Pes-
ticide Assessment Guidelines, Subdivision F, Hazard Evalua-
tion: Human and Domestic Animals Section 81-5); the Toxic
92
Substances Control Act (TSCA) Health Effects Test Guide-
lines, 40 CFR 798.4470; the Organization for Economic Coop-
eration and Development (OECD) Guideline for Testing of
Chemicals, Section 404; and the Japanese Agricultural Chemi-
cals Laws and Regulations Testing Guidelines for Toxicology
Studies published by the Society of Agricultural Chemical
Industry, under the auspices of MAFF (Ministry of Agricul-
ture, Forestry and Fisheries).
Two male and four female Albino New Zealand White rab-
bits (weighing 3477–3958 g at study initiation) were obtained
from Hazleton Research Products (Denver, PA, USA) and were
allowed free access to lab chow (Purina Certified Rabbit Chow,
No. 5322, St. Louis, MO, USA) and municipal water ad libi-
tum. Animals were acclimated to laboratory conditions for a
minimum of six days prior to initiation of dosing. The animal
room was kept at a constant temperature (70–72°F), humidity
(42–48%), and light (12 h light/12 h dark).
The route of HCA-SX administration was direct applica-
tion to shaved intact skin. This route of administration is
standard for assessment of local dermal irritative potential.
HCA-SX was assessed on two male and four female rabbits.
On the day prior to dosing, the hair was removed from the
backs of the rabbits using as Oster® small animal clipper. The
HCA-SX was moistened with approximately 0.5 ml deionized
water and applied to an area of skin approximately 2.5 × 2.5
cm under a secured 2-ply gauze patch, that was overwrapped
with a gauze binder and secured with Dermiform® tape. Plas-
tic restraint collars were applied and remained on the animals
for the duration of the exposure period. One intact site per
rabbit was used and the dosage level was 500 mg/site. Each
animal received a single, 4-h, semi-occluded exposure. At the
end of 4 h, the collars and bandages were removed and the
sites wiped with disposable towels moistened with deionized
water.
The rabbits were observed twice daily (morning and after-
noon) for mortality for the duration of the study. For dermal
observations, the application sites were observed for erythema,
edema and other dermal findings at approximately 30–60 min
and 24, 48 and 72 h after patch removal. Dermal irritation was
graded in accordance with the method of Draize [13]. In order
to facilitate dermal observations, the rabbits were shaved ap-
proximately 1 h prior to collecting the 72 h dermal scores.
The Primary Dermal Index was calculated from the scores re-
corded at 30–60 min, 24, 48 and 72 h after patch removal. The
mean scores for erythema and edema were calculated separately
to the nearest tenth and added together. Based on this value,
the Draize grading system (Table 2) was used to arrive at the
primary dermal irritation descriptive rating.
Body weights were obtained and recorded on study day 0
(initiation) and at each rabbit’s termination from the study.
Upon termination of the study, the rabbits were euthanized
by intravenous injection of sodium pentobarbital solution and
discarded.
Animals and treatment for primary eye irritation study
The objective was to determine the irritative potential of HCA-
SX following a single exposure to one eye of Albino rabbits.
The protocol was designed and the study was conducted in
compliance with the Environmental Protection Agency Guide-
lines for Registering Industrial Chemicals in the US (Pesticide
Assessment Guidelines, Subdivision F, Hazard Evaluation:
Human and Domestic Animals Section 81-4); the Toxic Sub-
stances Control Act (TSCA) Health Effects Test Guidelines,
40 CFR 798.4500; the Organization for Economic Cooperation
and Development (OECD) Guideline for Testing of Chemicals,
Section 405; and the Japanese Agricultural Chemicals Laws
and Regulations Testing Guidelines for Toxicology Studies
published by the Society of Agricultural Chemical Industry,
under the auspices of MAFF (Ministry of Agriculture, For-
estry and Fisheries).
Table 1. Acute dermal toxicity study of HCA-SX in Albino rabbits: Scoring criteria for dermal reactions
Evaluation of dermal reactions*
Value Erythema and eschar formation Computer designation Value Edema formation Computer designation
0 No erythema No erythema 0 No edema No edema
1 Very slight erythema (barely perceptible, Very slight erythema 1 Very slight edema (barely perceptible, Very slight edema
edges of area not well defined) edges of area not well defined)
2 Slight erythema (pale red in color and Slight erythema 2 Slight edema (edges of area well Slight edema
edges definable) defined by definite raising)
3 Moderate to severe erythema (definite Moderate erythema 3 Moderate edema (raised Moderate edema
red in color and area well defined) approximately 1 mm)
4 Severe erythema (beet or crimson red) Severe erythema 4 Severe edema (raised more than Severe edema
to slight eschar formation (injuries 1 mm and extending beyond area
in depth) of exposure)
*Source: Draize et al. [13].
93
Three male and three female New Zealand white Albino
rabbits (weighing 2172–2974 g at study initiation), obtained
from Hazleton Research Products (Denver, PA, USA), were al-
lowed free access to lab chow (Purina Certified Rodent Chow,
No. 5322, St. Louis, MO, USA) and municipal water ad libi-
tum. Animals were acclimated to laboratory conditions for a
minimum of six days prior to initiation of dosing. The animal
room was kept at a constant temperature (66–71°F), humidity
(46–86%), and light (12 h light/12 h dark).
The route of HCA-SX administration was direct conjuncti-
val instillation. This route of administration is standard for
assessment of local ocular irritative potential. HCA-SX was
placed directly into the cupped lower conjunctival sac of the
right (test) eye at a dose of 54 mg/right eye. The eyelid was
held closed for approximately 1 sec after instillation. The left
eye was manipulated in an identical manner to simulate the
dosing of the right eye. There was one group of six rabbits
that received a single, unwashed exposure.
The rabbits were observed twice daily (morning and after-
noon) for mortality for the duration of the study. For ocular
observations, both eyes of the rabbits were examined for ocu-
lar abnormalities prior to study initiation. The pre-initiation
examination included the use of sodium fluorescein and ultra-
violet light for detection of corneal abnormalities. Rabbits
assigned to the study had no pre-existing abnormalities. Fol-
lowing treatment, both eyes of the rabbits were examined
macroscopically for ocular irritation using a handheld pen light
in accordance with the method of Draize (Table 3) at approxi-
mately 1, 24, 48 and 72 h after dosing and on days 4, 7, 14,
and 21 if irritation persisted. In addition, both eyes were fur-
ther examined at 72 h and on days 7, 14 and 21 with sodium
fluorescein and ultraviolet light.
Body weights were obtained and recorded on study day
0 (initiation) and at each animal’s termination from study.
Upon termination, the rabbits were euthanized by intrave-
nous injection of sodium pentobarbital solution and dis-
carded.
Statistics
Data from different experiments (control and test) were pooled
and subjected to statistical analysis. Except where indicated
otherwise, values given are arithmetic means ± S.E.M. Sig-
nificance of differences between control and test values were
evaluated using analysis of variance (ANOVA) followed by
Dunnett’s test (GraphPad Software, San Diego, CA, USA).
Differences with p values < 0.05 were accepted as statistically
significant.
Results
The objectives of this study were to determine the effects of
HCA-SX on 5-HT uptake in rat brain cortex in vitro, and to
determine the safety profile of HCA-SX in in vivo models.
Table 2. Primary dermal irritation study of HCA-SX Albino rabbits: Scoring criteria for dermal reactions
Evaluation of dermal reactions*
Value Erythema and eschar formation Value Edema formation
0 No erythema 0 No edema
1 Very slight erythema (barely perceptible, edges of area not 1 Very slight edema (barely perceptible, edges of area not
well defined) well defined)
2 Slight erythema (pale red in color and edges definable) 2 Slight edema (edges of area well defined by definite raising)
3 Moderate to severe erythema (defined in color and area 3 Moderate edema (raised approximately 1 mm)
well defined)
4 Severe erythema (beet or crimson red) to slight eschar 4 Severe edema (raised more than 1 mm and extending beyond
formation (injuries in depth) area of exposure)
4 Total possible erythema score 4 Total possible edema score
8 Total possible primary irritation score
Descriptive ratings
Mean primary dermal irritation index
Range of values Descriptive ratings
0 Non-irritating
0.1–2.0 Slightly irritating
2.1–5.0 Moderately irritating
5.1–8.0 Severely irritating
*Source: Draize 1965 [13].
94
Effect of HCA-SX on [3H]-5-HT uptake
This study was conducted to determine the effects of HCA-SX
on 5-HT uptake in rat brain cortex in vitro. As shown in Fig. 1,
[3H]-5-HT was taken up into neuronal stores in rat cortical slices
in a time-dependent manner reaching a maximum at approxi-
mately 90 min. After 40 min, HCA-SX (300 and 1 mM), and
the 5-HT uptake inhibitors fluoxetine (100 µM) plus clomi-
pramine (10 µM) decreased the time-dependent uptake of [3H]-
5-HT. At 90 min, HCA-SX (300 µM) caused a 20% decrease in
[3H]-5-HT uptake, whereas fluoxetine plus clomipramine sig-
nificantly inhibited (p < 0.001) [3H]-5-HT uptake by 30% (Fig.
2). At 90 min, significant inhibition of 5-HT uptake was pro-
duced by 300 µM HCA-SX (p < 0.01); however, no signifi-
cant inhibition was observed following incubation with 1 mM
HCA-SX (Fig. 2). Thus, kinetics studies were performed on
HCA and fluoxetine plus clomipramine in order to determine
optimal uptake at equilibrium. Uptake at equilibrium is a re-
flection of total uptake under our experimental conditions. Re-
gardless of initial differences observed for the agents before
Table 3. Primary eye irritation study of HCA-SX in Albino rabbits: Scale for scoring ocular irritationa
I. Cornea
(A) Opacity-degree of density (area most dense taken for reading)
No ulceration or opacity 0
Dulling of normal luster, details of iris clearly visible 1 *
Easily discernible translucent areas, details of iris slightly obscured 2 *
Nacreous areas, no details of iris visible, size of pupil barely discernible 3 *
Opaque cornea, iris not discernible through the opacity 4 *
(B) Area of cornea involved
No ulceration or opacity 0
One quarter or less but not zero 1*
Greater than one quarter, but less than half 2*
Greater than half, but less than three quarters 3*
Greater than three quarters, up to whole area 4*
Score equals A × B × 5 Total maximum = 80
II. Iris
(A) Values
Normal 0
Markedly deepened rugae, congestion, swelling, circumcorneal injection (any or all of these 1 *
or combination of any thereof), iris still reacting to light (sluggish reaction is positive)
No reaction to light, hemorrhage, gross destruction (any or all of these) 2 *
Score equals A × 5 Total maximum = 10
III. Conjunctivae
(A) Redness (refers to palpebral and bulbar conjuctivae excluding cornea and iris)
Blood vessels normal 0
Some blood vessels definitely hyperemic (injected above) normal 1
Diffuse, deeper crimson color, individual vessels not easily discernible 2 *
Diffuse beefy red 3*
(B) Chemosis: lids and/or nictitating membranes
No swelling 0
Any swelling above normal (includes nictitating membrane) 1
Obvious swelling with partial eversion of lids 2*
Swelling with lids about half closed 3*
Swelling with lids more than half closed 4*
(C) Discharge
No discharge 0
Any amount different from normal (does not include small amounts observed in inner canthus 1
of normal animals)
Discharge with moistening of the lids and hairs just adjacent to the lids 2
Discharge with moistening of the lids and hair, and considerable area around the eye 3
Score equals (A + B + C) × 2 Total maximum = 2 0
Total maximum Score Possible = 110
aDraize scale for scoring ocular lesions, as published in the guidelines in Subsection F. Hazard Evaluation: Human and Domestic Animals distributed
in 1982 and the OECD Guidelines for Testing of Chemicals distributed by 1987.
*Starred figures indicate positive effect.
95
90 min, only data obtained at or after 90 min is reflective of
uptake at equilibrium.
HCA-SX safety studies
In this paper, we have conducted a number of in vivo studies
to demonstrate the safety of HCA-SX. Acute oral toxicity,
acute dermal toxicity, primary dermal irritation and primary
eye irritation studies were conducted in animals to determine
the safety profile of HCA-SX.
Fig. 1. Time-dependent uptake of [3H]-5-HT in isolated rat brain cortex: control and in the presence of (–)-hydroxycitric acid (HCA-SX, 300
µM and 1 mM) and fluoxetine (100 µM) plus clomipramine (10 µM). Each data point is the mean ± S.E.M. of 3–6 replicates. *p < 0.01; **p <
0.001 significantly different from untreated control.
Fig. 2. [3H]-5-HT uptake in isolated rat brain cortex slices at 90 min control and in the presence of (–)-hydroxycitric acid (HCA-SX, 300 µM and
1 mM) and fluoxetine (100 µM) plus clomipramine (10 µM). Vertical bars represent mean ± S.E.M. Number of observations was 4 in each case.
*p < 0.01; **p < 0.001 significantly different from untreated control.
Table 4. Acute oral toxicity study of HCA-SX in Albino rats follow-
ing a single oral dose of 5000 mg/kg: Summary of clinical findings:
Total occurrence/number of animals
Group Male Female
Days 0–14 Days 0–14
Acute
Appeared normal 73/5 79/5
Scabbing dorsal head 12/1 –
Rales – 5/ 2
Soft stool 1/1
See Materials and methods for details.
96
Acute oral toxicity study
Mortality
No deaths were observed during the study from any of the
doses of HCA-SX which were used.
Clinical observations (Tables 4 and 5)
All clinical findings were noted on the day of dosing, with
the exception of scabbing of the dorsal head that was present
for one male on day 3 and throughout the remainder of the
study. Soft stool and rales were observed for one and two
rats, respectively. There were no other clinical findings.
Body weights (Table 6)
There were no remarkable changes or differences observed
in body weights under any of the experimental conditions.
Necropsy (Table 7)
There were no significant changes for all examined tissues at
terminal necropsy for any of the doses of HCA-SX which were
used.
There were no toxicological gross findings for any exam-
ined tissues at the scheduled necropsy. The data indicates
that the LD50 of HCA-SX is greater than 5000 mg/kg when
administered once orally via gastric intubation to fasted male
and female Albino rats.
Table 5. Acute oral toxicity study of HCA-SX in Albino rats following a single oral dose of 5,000 mg/kg: Individual clinical observations
Observation Sex Hour post-dose Day post-dose
134 1234567891011121314
Appeared normal Male P P P PPPPPPPPPPPPPP
Male P P P PPPPPPPPPPPPPP
Male P P P PPPPPPPPPPPPPP
Male P P P P P –––––––––––
Male P P P PPPPPPPPPPPPPP
Female – – – PPPPPPPPPPPPPP
Female – – P PPPPPPPPPPPPPP
Female P P P PPPPPPPPPPPPPP
Female P P – PPPPPPPPPPPPPP
Female P P P PPPPPPPPPPPPPP
Rales Female P P P –––––––––––––
Female P P – –––––––––––––
Soft stool Female – – P –––––––––––––
Scabbing dorsal head Male – – – – – PPPPPPPPPPPP
Grade code: P = present; S = slight; M = moderate; V = severe; – = not seen at that interval. See Materials and methods for details.
Table 6. Acute oral toxicity study of HCA-SX in Albino rats following a single oral dose of 5000 mg/kg: Individual and mean ± S.D. of body weights
(g) on days post-treatment
Sex –1 day 0 days 7 days 14 days Sex –1 day 0 days 7 days 14 days
Male 239 210 29 2 34 6 Female 232 211 240 260
Male 244 216 30 3 34 9 Female 232 208 237 240
Male 239 210 28 8 32 3 Female 245 215 245 258
Male 244 215 29 6 33 9 Female 234 213 244 261
Male 238 213 29 4 33 7 Female 230 211 249 256
Mean ± S.D. 241 ± 2.9 213 ± 2.8 295 ± 5.5 339 ± 10.1 235 ± 6.0 212 ± 2.6 243 ± 4.6 255 ± 8.6
See Materials and methods for details.
Table 7. Acute oral toxicity study of HCA-SX in Albino rats follow-
ing a single oral dose of 5000 mg/kg: Gross necropsy observations
incidence summary
Scheduled necropsy Male Female
Number of animals in dose group 5 5
Number of animals terminally euthanized 5 5
No significant changes observed 5 5
– all examined tissues
See Materials and methods for details.
97
Acute dermal toxicity study
Mortality
There were no deaths during the study under the experimen-
tal conditions which were employed.
Clinical observations (Tables 8 and 9)
All animals appeared normal throughout the study with the
exception of one male rabbit, which had mucoid feces on days
0–2. No other clinical findings were observed for animals re-
ceiving a dermal application of 2000 mg HCA-SX/kg.
Dermal observations (Table 10)
HCA-SX induced very slight to slight erythema on all rab-
bits. No edema was observed. Desquamation was noted on
eight sites out of 10 animals by day 7. All treatment sites were
stained yellow. All dermal irritation completely subsided by
day 12 or earlier, and no other dermal findings were observed.
Body weights (Table 11)
No remarkable changes or differences in body weight oc-
curred.
Table 8. Acute dermal toxicity study of HCA-SX in Albino rabbits
following a single dose of 2000 mg/kg: Summary of clinical findings:
Total occurrence/number of animals
Male Female
Acute Days 0–14 Days 0–14
Appeared normal 82/5 85/5
Mucoid feces 3/ 1
See Materials and methods for details.
Table 9. Acute dermal toxicity study of HCA-SX in Albino rabbits following a single dose of 2000 mg/kg: Individual clinical observations
Observation Sex Hour post-dose Day post-dose
134 1234567891011121314
Appeared normal Male P P P PPPPPPPPPPPPPP
Male P P P PPPPPPPPPPPPPP
Male P P P PPPPPPPPPPPPPP
Male – P P – – PPPPPPPPPPPP
Male P P P PPPPPPPPPPPPPP
Female P P P PPPPPPPPPPPPPP
Female P P P PPPPPPPPPPPPPP
Female P P P PPPPPPPPPPPPPP
Female P P P PPPPPPPPPPPPPP
Female P P P PPPPPPPPPPPPPP
Mucoid feces Male P – – –––––––––––––
Grade code: P = present; S = slight; M = moderate; V = severe; = not seen at that interval. See Materials and methods section for details.
Table 10. Acute dermal toxicity study of HCA-SX in Albino rabbits following a single dose of 2000 mg/kg: Individual dermal observations
Erythema+/edema+/other findings
Study day Male Male Male Male Male Female Female Female Female Female
1 2/0/h 1/0/h 1/0/h 1/0/h 2/0/h 2/0/h 1/0/h 2/0/h 2/0/h 2/0/h
2 2/0/h 1/0/h 1/0/h 1/0/h 2/0/h 2/0/h 1/0/h 1/0/h 2/0/h 1/0/h
3 2/0 1/0 1/0 1/0 2/0 2/0 1/0 1/0 1/0 1/0
4 2/0 1/0 1/0 1/0 2/0 2/0 1/0 1/0 1/0 1/0
5 2/0 1/0 1/0 1/0 2/0 2/0 1/0 1/0 1/0 1/0
6 2/0/d 1/0 1/0/d 1 /0 2/0/d 1/0/d 1/0 1/0 1/0/d 1/0
7 2/0/d 1/0/d 1/0/d 1/0/d 1/0/d 1/0/d 1/0/d SNR SNR SNR
8 1/0/d 1/0/d 1/0 1/0 1/0/d SNR 1/0 SNR SNR SNR
9 1/0/d SNR 1/0 SNR 1/0/d SNR SNR SNR SNR SNR
1 0 1/0 SNR 1 / 0 SNR SNR SNR SNR SNR SNR SNR
11 1/0 SNR SNR SNR 0/0/d SNR SNR SNR SNR SNR
1 2 SNR SNR SNR SNR SNR SNR SNR SNR SNR SNR
1 3 SNR SNR SNR SNR SNR SNR SNR SNR SNR SNR
1 4 SNR SNR SNR SNR SNR SNR SNR SNR SNR SNR
+ = Refer to Draize scale for dermal scoring criteria; d = desquamation; SNR = scored, not remarkable; h = application site stained yellow. See
Materials and methods section for details.
98
Necropsy (Tables 12 and 13)
Reddened application sites were noted for two rabbits at the
terminal necropsy. Single occurrences of pale kidneys, mot-
tled lungs and hair loss were noted for one rabbit each. Two
rabbits had accessory spleens, a common congenital abnor-
mality. There were no other gross necropsy findings for all
examined tissues at the terminal necropsy.
The data indicated that the LD50 of HCA-SX is greater than
2000 mg/kg when it is applied once for 24 h to the shaved,
intact skin of male and female Albino rabbits. There was no
evidence of acute systemic toxicity among rabbits when 2000
mg/kg HCA-SX is dermally administered.
Primary dermal irritation study
Mortality
No deaths occurred during this study.
Dermal observations (Table 14)
HCA-SX induced very slight erythema on one animal and
stained all treatment sites yellow. All erythema completely
subsided by the end of day 1. No edema or other dermal find-
ings were observed.
Body weights (Table 15)
No significant body weight changes occurred during the study
period.
The Primary Irritation Index was calculated to be 0.0. HCA-
SX received a descriptive rating classification (Table 2) of non-
irritating.
Primary eye irritation study
Mortality
There were no deaths.
Ocular irritation (Tables 16 and 17)
Number with positive effect/Number treated
Group Cornea Iris Conjunctiva Total Maximum
average score
(M.A.S.)
0.1 ml/ 0/6 6 /6 6/6 6/ 6 15.0 at 1 h
right eye,
unwashed
None of the rabbits vocalized upon instillation of the test
material (HCA-SX). The left (control) eyes were free of evi-
dence of ocular irritation and other findings for the duration
of the study. Individual and average ocular irritation scores
for the treated eyes are presented in Table 16. Individual ani-
mal results (other findings), including sodium fluorescein ex-
amination results, are presented in Table 17. The scale for
scoring ocular irritation and method of score calculation are pre-
sented in Table 3 with a total maximum possible score of 110.
A small area of inflammatory exudate with enlarged blood
vessels was present at the apex of the lower conjuctival sac
for three rabbits on day 7. This inflammatory exudate com-
pletely subsided by study termination (day 21) for two animals.
One of the three rabbits also had inflammatory exudate on the
nictitating membrane that was present on day 7 through study
termination (day 21). The Maximum Average Score (M.A.S.)
for HCA-SX was 15.0 at 1 h. The test material induced posi-
tive iridal and conjunctival reactions for all rabbits. There
were no corneal reactions noted. Iridal irritation subsided by
48 h or earlier. With the exception of inflammatory exudate,
conjunctival irritation completely subsided by study termi-
nation (day 21) or earlier for all animals.
Table 11. Acute dermal toxicity study of HCA-SX in Albino rabbits: Individual and mean + S.D. body weights (g)
Sex Day 0 Day 7 Day 14 Sex Day 0 Day 7 Day 14
Male 2323 2835 2851 Female 2227 2322 2529
Male 2266 2622 2751 Female 2295 2557 2777
Male 2330 2656 2769 Female 2372 2742 2854
Male 2466 2953 3004 Female 2236 2655 2697
Male 2259 2623 2721 Female 2351 2674 2826
Mean ± S.D. 2329 ± 83.2 2738 ± 149.2 2819 ± 114.0 2296 ± 65.5 2590 ± 163.8 2737 ± 130.5
See Materials and methods for details.
Table 12. Acute dermal toxicity study of HCA-SX in Albino rabbits
following a single dose of 2000 mg/kg: Summary of gross necropsy
observations incidence
Male Female
Number of animals in dose group 5 5
Number of animals terminally euthanized 5 5
Application site – reddened 1 1
Kidneys – pale 0 1
Spleen – accessory 1 1
External surface – hair loss 1 0
All examined tissues – no significant changes 2 2
observed
See Materials and methods for details.
99
Table 13. Acute dermal toxicity study of HCA-SX in Albino rabbits: Individual gross description of organs
Sex Organ findings Grade Organ – no significant findings Sex Organ findings Grade Organ – no significant findings
Male None NA adrenal glands, application site, brain, Female Kidneys gross: P adrenal glands, application site,
intestine, epididymides, esophagus, pale bilateral brain, intestine, esophagus, eyes,
eyes, gall bladder, heart, kidneys, gall bladder, heart, liver, lymph
liver, lymph node, mesenteric, lungs, node, mesenteric, lungs,
mammary gland, pancreas, pituitary, mammary gland, ovaries,
prostate, salivary glands, seminal pancreas, pituitary, salivary
vesicles, skin, spleen, stomach, glands, skin, spleen, stomach,
testes, thymus gland, thyroid glands, thymus gland, thyroid glands,
trachea, urinary bladder trachea, urinary bladder, uterus
Male Lungs gross: P adrenal glands, application site, Female None NA adrenal glands, application site,
mottled all brain, intestine, epididymides, brain, intestine, esophagus, eyes,
lobes esophagus, eyes, gall bladder, heart, gall bladder, heart, kidneys, liver,
kidneys, liver, lymph node, lymph node, mesenteric, lungs,
mesenteric, mammary gland, mammary gland, ovaries,
pancreas, pituitary, prostate, pancreas, pituitary, salivary
salivary glands, seminal vesicles, glands, skin, spleen, stomach,
skin, spleen, stomach, testes, thymus gland, thyroid glands,
thymus gland, thyroid glands, trachea, urinary bladder, uterus
trachea, urinary bladder
Male Application P adrenal glands, brain, intestine, Female None NA adrenal glands, application site,
site gross: epididymides, esophagus, eyes, brain, intestine, esophagus, eyes,
reddened gall bladder, heart, kidneys, liver, gall bladder, heart, kidneys, liver,
lymph node, mesenteric, lungs, lymph node, mesenteric, lungs,
mammary gland, pancreas, mammary gland, ovaries,
pituitary, prostate, salivary pancreas, pituitary, salivary
Spleen gross: P glands, seminal vesicles, skin, glands, skin, spleen, stomach,
accessory two, stomach, testes, thymus gland, thymus gland, thyroid glands,
1 and 2 mm thyroid glands, trachea, urinary trachea, urinary bladder, uterus
diameter bladder
Male None NA adrenal glands, application site, Female Application site P adrenal glands, brain, intestine,
brain, intestine, epididymides, gross: reddened esophagus, eyes, gall bladder,
esophagus, eyes, gall bladder, heart, kidneys, liver, lymph node,
heart, kidneys, liver, lymph node, mesenteric., lungs, mammary
mesentericsenteric, lungs, gland, ovaries, pancreas,
mammary gland, pancreas, pituitary, salivary glands, skin,
pituitary, prostate, salivary glands, spleen, stomach, thymus gland,
seminal vesicles, skin, spleen, thyroid glands, trachea, urinary
stomach, testes, thymus gland, bladder, uterus
thyroid glands, trachea, urinary
bladder
Male External P adrenal glands, application site, Female Spleen gross: P adrenal glands, application site,
surface gross: brain, intestine, epididymides, accessory two, brain, intestine, esophagus, eyes,
hair loss esophagus, eyes, gall bladder, heart, 2 mm in gall bladder, heart, kidneys, liver,
ventral kidneys, liver, lymph node, diameter lymph node, mesenteric, lungs,
abdominal mesenteric, lungs, mammary gland, mammary gland, ovaries,
pancreas, pituitary, prostate, pancreas, pituitary, salivary
salivary glands, seminal vesicles, glands, skin, stomach, thymus
skin, spleen, stomach, testes, gland, thyroid glands, trachea,
thymus gland, thyroid glands, urinary bladder, uterus
trachea, urinary bladder
P = present; NA = none appeared. See Materials and methods for details.
100
Table 16. Primary eye irritation study of HCA-SX in Albino rabbits: Individual ocular irritation scores
Examination intervals
Sex Tissue 1 h 24 h 48 h 72 h 4 days 7 days 14 days 21 days
Male Cornea (O-A) 0 0 0 0 0 0 0 0 0 0 0 0
Iris 110000
Conjunctiva (R-C-D) 2 2 2 1 0 0 1 0 0 1 0 0 1 0 0 0 0 0
Male Cornea (O-A) 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Iris 11000000
Conjunctiva (R-C-D) 1 2 2 1 0 0 2 0 0 2 0 0 2 0 0 2 0 0 1 0 0 0 0 0
Male Cornea (O-A) 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Iris 11000000
Conjunctiva (R-C-D) 1 2 1 1 0 0 2 0 0 2 0 0 2 0 0 1 0 0 0 0 0 0 0 0
Female Cornea (O-A) 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Iris 1000000
Conjunctiva (R-C-D) 2 1 2 1 0 0 1 0 0 1 0 0 1 0 0 1 0 0 0 0 0
Female Cornea (O-A) 0 0 0 0 0 0 0 0 0 0 0 0
Iris 110000
Conjunctiva (R-C-D) 1 2 2 1 1 0 2 1 0 1 0 0 1 0 0 0 0 0
Female Cornea (O-A) 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Iris 10000000
Conjunctiva (R-C-D) 1 2 2 1 1 0 2 0 0 2 0 0 2 0 0 2 0 0 0 0 0 0 0 0
Group: 54 mg/right eye, unwashed. *Fluorescein solution applied; O = opacity; A = area; R = redness; C = chemosis; D = discharge.
See Materials and methods for details.
Table 15. Primary dermal irritation study of HCA-SX in Albino rab-
bits: Individual body weights (g)
Sex Initiation Day 0 Terminal Day 3
Male 3952 3937
Male 3550 3465
Female 3738 3690
Female 3958 3754
Female 3477 3531
Female 3691 3629
See Materials and methods for details.
Table 14. Primary dermal irritation study of HCA-SX in Albino rabbits: Individual dermal scores
Sex Site Erythema Edema
1 h 24 h 48 h 72 h 1 h 24 h 48 h 72 h
Male A 0000 0000
Male A 0000 0000
Female A 0000 0000
Female A 1000 0000
Female A 0000 0000
Female A 0000 0000
Total 1000 0000
The dosage level was 0.5 g HCA-SX/site. *Dose site stained yellow. See Materials and methods for details.
PII calculated using test periods: 1, 24, 48, 72 h.
Primary irritation index: (PII) = (1 + 0 + 0 + 0)/24 + (0 + 0 + 0 + 0)/24
PII = (1/24) + (0/24) = 0.0 + 0.0
PII = 0.0 = non-irritating.
Body weights (Table 17)
There were no significant changes or differences observed
in body weights during the study period.
Discussion
(–)-Hydroxycitric acid (HCA) is the natural primary organic acid
found in the fruits and rinds of Garcinia cambogia. HCA works
by inhibiting lipogenesis, the process by which the body con-
verts carbohydrates into fat by temporarily inhibiting ATP
101
Table 17. Primary eye irritation study of HCA-SX in Albino rabbits: Individual animal results – other findings
Sex Initial body Terminal body Examination Other Sex Initial body Terminal body Examination Other
wt. (g) wt. (g) interval findings wt. (g) wt. (g) interval findings
Day 0 Day 0 Day 0 Day 0
Male 2214 2449 (day 7) 1 h b,s Female 2974 3423 (day 14) 1 h b,s
24 h 24 h
48 h 48 h
72 h r(0%) 72 h r(0%)
4 days 4 days
7 days r(0%) 7 days r(0%)
14 days r(0%)
Male 2239 2934 (day 21) 1 h b,s Female 2346 2700 (day 7) 1 h b,s
24 h 24 h
48 h 48 h
72 h r(0%) 72 h r(0%)
4 days 4 days
7 days e,r(0%) 7 days r(0%)
14 days f,r(0%)
21 days r(0%)
Male 2172 2884 (day 21) 1 h b,s Female 2334 2755 (day 21) 1 h b,s
24 h 24 h
48 h 48 h
72 h r(0%) 72 h r(0%)
4 days 4 days
7 days g,r(0%) 7 days e,f,r(0%)
14 days g,r(0%) 14 days e,f,r(0%)
21 days r(0%) 21 days f,r(0%)
Group: 54 mg/right eye, unwashed; b = clear discharge; r = sodium fluorescein stain retention (0% of area); s = small amount of residual test material
present in the eye; e = inflammatory exudate covering an area of approximately 4 × 2 mm containing enlarged blood vessels present at apex of
lower conjuctival sac; f = inflammatory exudate covering an area of approximately 1 × 1 mm containing enlarged blood vessels present at apex
of lower conjunctival sac; g = inflammatory exudate covering an area of approximately 5 × 3 mm containing enlarged blood vessels present at
apex of lower conjunctival sac.
citrate lyase, the enzyme that converts excess glucose into
fat. Furthermore, by inhibiting ATP citrate lyase, HCA reduces
the availability of acetyl-CoA, the building block for fat syn-
thesis. Earlier studies in humans have shown that HCA con-
sumption (500 mg, t.i.d.) before meals for 8 weeks resulted in
215% greater weight loss than those taking a placebo with-
out any adverse side effects [5]. Mattes and Bormann [2]
conducted a placebo-controlled, double-blind study involv-
ing 89 moderately overweight female human subjects who
ingested HCA (1200 mg/day) or placebo for 12 weeks and
received a daily diet of 5020 kJ. A significant loss of body
weight was observed as compared to the placebo group (3.7
± 3.1 kg vs. 2.4 ± 2.9 kg) for subjects ingesting HCA.
We have recently shown that HCA (as Super CitriMaxTM,
HCA-SX) can increase the release of 5-HT from rat brain cor-
tex slices, in vitro, provided the first scientific evidence offer-
ing an explanation for the ability of HCA to suppress appetite
[7]. In the present study, we now report that HCA-SX inhib-
ited the time-dependent uptake of [3H]-5-HT, an action that
was mimicked by the well-known serotonin receptor reuptake
inhibitors (SRRIs), fluoxetine and clomipramine. Fluoxetine and
clomipramine are potent selective inhibitors of [3H]-5-HT up-
take in neuronal tissue both in vivo and in vitro [14, 15]. For
instance, Stauderman and Jones [16], demonstrated that flu-
oxetine inhibited accumulation of [3H]-5-HT into rat spinal
serotonergic nerve terminals in a sodium-dependent manner.
Taken together, these results support the view that HCA-SX
increases the release/availability of [3H]-5-HT from neuronal
serotonergic nerve terminals presumably via an effect on neu-
ronal uptake of this monoamine.
Since increased brain levels of 5-HT are involved in regu-
lation of sleep, mood changes and appetite suppression, the
results strongly suggest that an effect on this monoamine could
underlie the mechanism of appetite suppression and food in-
take induced by HCA-SX. Furthermore, HCA-SX may provide
a therapeutic advantage by alleviating emotional issues of
overweight people, including binge-eating and depression.
It has been demonstrated by GC-MS that HCA-SX is bio-
available in human plasma [6]. However, a major question
remains whether HCA-SX can cross the blood–brain barrier.
Recently, we have completed a human clinical study on HCA-
SX, which demonstrated that HCA-SX supplementation over
8 weeks increases serum serotonin levels significantly in hu-
man volunteers (unpublished results). The other advantage
102
of HCA-SX is that many of the ‘natural’ diet products on the
market, including Ma Huang (a natural source of ephedrine),
or Kola Nut (a natural source of caffeine) and Guarana extract
(a natural source of caffeine) are central nervous system stimu-
lants known to cause serious adverse side effects, including
increased heart rate, high blood pressure, upset stomach, se-
vere headaches/migraines, vomiting, and others. HCA-SX has
been in the market for more than 8 years with no such adverse
side effects reported so far. In addition, high concentrations
(20–30%) of HCA in the dried fruit rind of Garcinia cambogia
have been consumed for centuries in the diets of South Asian
people as a food condiment to make meals more ‘filling’ [1].
A number of safety studies have been conducted on HCA-
SX. In the acute oral toxicity study, no deaths, remarkable body
weight changes or gross necropsy findings were observed.
Clinical findings were limited to soft stool and rales for one
and two rats, respectively. A male rat with scabbing on the
dorsal head was noted on days 3–14. All other animals ap-
peared normal on day 1 and throughout the remainder of the
study. There were no other clinical findings. These results
demonstrate that the LD50 of HCA-SX is greater than 5000 mg/
kg when administered once orally via gastric intubation to
fasted male and female albino rats.
In the dermal toxicity study, there were no deaths, HCA-SX-
related clinical findings or remarkable body weight changes.
The HCA-SX induced very slight to slight erythema with no
edema. Desquamation was noted on eight sites. All dermal
irritation completely subsided by day 12 or earlier. There were
no other dermal findings. Reddened application sites were
noted for two rabbits at the terminal necropsy. There were no
other gross necropsy findings related to HCA-SX for all ex-
amined tissues at the terminal necropsy. The LD50 of HCA-
SX was found to be greater than 2,000 mg/kg when applied
once for 24 h to the shaved, intact skin of male and female
albino rabbits. There was no evidence of acute systemic tox-
icity among rabbits that were dermally administered HCA-SX
at 2,000 mg/kg.
In the dermal irritation study, no deaths or significant body
weight changes were observed during the study period. HCA-
SX induced very slight erythema on one animal. No edema or
other dermal findings were noted, and all irritation was revers-
ible and completely subsided by the end of day 1. The Pri-
mary Irritation Index was calculated to be 0.0. Based on these
observations, HCA-SX received a descriptive rating classifi-
cation (Table 2) of non-irritating.
In the eye irritation study, no deaths or remarkable changes
in body weights occurred during the study period. The re-
sults indicate that HCA-SX causes ocular irritation with pro-
duction of inflammatory exudate in some animals. Positive
iridal and conjuctival reactions were present in all animals,
which subsided within 48 h. A total maximum score of 110 is
possible. A score of 15 was obtained in the study, indicating
mild irritation.
Taken together, this study had two major objectives (a) to
determine the effect of HCA-SX on 5-HT uptake in the rat
brain cortex in vitro, and (b) to assess the safety profile of
HCA-SX. The in vitro, serotonin release and reuptake stud-
ies demonstrate that HCA-SX can inhibit [3H]-5-HT uptake
(and increase 5-HT availability) in a manner similar to that of
serotonin receptor reuptake inhibitors (SRRIs), and thus may
prove beneficial in controlling appetite, as well as, in the treat-
ment of depression, insomnia, migraine headaches, and other
serotonin deficient conditions. However, fluoxetine plus clomi-
pramine exhibited significantly higher potency as compared
to HCA-SX. Acute oral toxicity, acute dermal toxicity, primary
dermal irritation and primary eye irritation studies indicate that
HCA-SX is a safe, natural supplement under the conditions it
was tested.
Acknowledgements
The authors acknowledge WIL Research Laboratories, Ash-
land, OH, USA, for performing the acute toxicity studies. The
authors thank Ms. Kristine Strong for technical assistance.
References
1 . Sergio W: A natural food, the Malabar Tamarind, may be effec-
tive in the treatment of obesity. Med Hypotheses 29: 39–40, 1988
2 . Mattes DR, Bormann L: Effects of (–)-hydroxycitric acid on
appetitive variables. Physiol Behav 71: 87–94, 2000
3 . Sullivan AC, Triscari J, Hamilton JG, Miller ON, Wheatley VR:
Effect of (–)-hydroxycitrate upon the accumulation of lipid in
the rat. I. Lipogenesis. Lipids 9: 121–128, 1974
4 . Sullivan AC, Triscari J, Hamilton JG, Miller ON: Effect of (–)-
hydroxycitrate upon the accumulation of lipid in the rat. II. Ap-
petite. Lipids 9: 129–134, 1974
5 . Ramos RR, Saenz JLS, Agular RJA: Extract of Garcinia Cambo-
gia in controlling obesity. Invest Med Int 22: 97–100, 1995
6 . Loe YC, Bergeron N, Rodriguez N, Schwarz J-M: Gas chromatog-
raphy/mass spectrometry method to quantify blood hydroxycitrate
concentration. Analyt Biochem 292: 148–154, 2001
7 . Ohia SE, Awe O, LeDay AM, Opere CA, Bagchi D: Effect of
hydroxycitric acid on serotonin release from isolated rat brain
cortex. Res Commun Mol Pathol Pharmacol 109: 210–216,
2001
8 . Leibowtz SF, Alexander JT: Hypothalamic serotonin in control
of eating behavior, meal size, and body weight. Biol Psychiatry
44: 851–864, 1998
9 . Mauri MC, Rudelli R, Somasehini E, Roncoroni L, Papa R, Man-
tero M, Penati G: Neurobiological and psychopharmacological
basis in the therapy of bulimia and anorexia. Prog Neuropsycho-
pharmacol Biol Psychiatry 20: 207–240, 1996
10 . Wilding J, Widdowson P, Williams G: Neurobiology. Br Med Bull
53: 286–306, 1997
11. Kirksey DF, Slotkin TA: Concomitant development of [3H]-
dopamine and [3H]-5-hydroxytryptamine uptake systems in rat
brain regions. Br J Pharmacol 67: 387–391, 1979
103
12 . Goodlet I, Mireylees SE, Sugrue MF: Effects of mianserin, a new
antidepressant, on the in vitro and in vivo uptake of monoamines.
Br J Pharmacol 61: 307–313, 1977
13 . Draize JH: The Appraisal of the Safety of Chemicals in Foods,
Drugs and Cosmetics. Dermal Toxicity. Association of Food and
Drug Officials of the US, Topeka, KA, 1965, pp 46–59
14 . Fuller RW, Wong DT, Robertson DW: Fluoxetine, a selective in-
hibitor of serotonin uptake. Med Res Rev 11: 17–34, 1991
15 . Lingjaerde O, Kildemo O: Dopamine uptake in platelets: Two dif-
ferent low affinity, saturable mechanisms. Agents Actions 11: 410–
416, 1981
16 . Stauderman KA, Jones DJ: Characterization of sodium-depend-
ent, high-affinity serotonin uptake in rat spinal cord synapto-
somes. Brain Res 330: 11–20, 1985
104
... Among all the reported compounds, hydroxycitric acid, garcinol, kaempferol, epicatechin, amentoflavone, apigenin, isovitexin, orientin, caffeic acid, and luteolin are expected to play the key role in conferring neuroprotection which is exhibited by G. morella extract in our study (Choudhury et al. 2018;Pandey et al. 2015;Bheemaiah and Kushalappa 2019;Murthy et al. 2020). Fascinatingly, these compounds could cross the blood-brain barrier and are known to ameliorate mitochondrial dysfunctions, inflammation, and oxidative stress via modulating different cell signalling pathways (Cao et al. 2017;Chetia Phukan et al. 2022;Siddique and Jyoti 2017;Bitu Pinto et al. 2015;Pan et al. 2020;Wang et al. 2017;Siddique 2021;Liu et al. 2019;Ohia et al. 2001). Prevention of dopaminergic neuronal loss was shown in both cell culture and animal models with these compounds by upregulation of Nrf2 and PI3K/ Akt signaling pathways, thereby reducing the oxidant and inflammatory responses and promoting cell survival and growth (Chung et al. 2019;Chetia Phukan et al. 2022;Hou et al. 2022;Cao et al. 2017;Li et al. 2020;Cheng et al. 2016;Liu et al. 2019;Pan et al. 2020;Xiao et al. 2018;Tsai et al. 2011;Ma et al. 2020;Antala et al. 2012). ...
Article
Full-text available
Dopaminergic neuroprotection is the main interest in designing novel therapeutics against Parkinson’s disease (PD). In the process of dopaminergic degeneration, mitochondrial dysfunctions and inflammation are significant. While the existing drugs provide symptomatic relief against PD, a therapy conferring total neuroprotection by targeting multiple degenerative pathways is still lacking. Garcinia morella is a common constituent of Ayurvedic medication and has been used for the treatment of inflammatory disorders. The present study investigates whether administration of G. morella fruit extract (GME) in MPTP mouse model of PD protects against dopaminergic neurodegeneration, including the underlying pathophysiologies, and reverses the motor behavioural abnormalities. Administration of GME prevented the loss of dopaminergic cell bodies in the substantia nigra and its terminals in the corpus striatum of PD mice. Subsequently, reversal of parkinsonian behavioural abnormalities, viz. akinesia, catalepsy, and rearing, was observed along with the recovery of striatal dopamine and its metabolites in the experimental model. Furthermore, reduced activity of the mitochondrial complex II in the nigrostriatal pathway of brain of the mice was restored after the administration of GME. Also, MPTP-induced enhanced activation of Glial fibrillary acidic protein (GFAP) and neuronal nitric oxide synthase (nNOS) in the nigrostriatal pathway, which are the markers of inflammatory stress, were found to be ameliorated on GME treatment. Thus, our study presented a novel mode of dopaminergic neuroprotection by G. morella in PD by targeting the mitochondrial dysfunctions and neuroinflammation, which are considered to be intricately associated with the loss of dopaminergic neurons.
... GC extract enhanced the time spent in the corners versus time spent in the center zone in OF test as well as increased immobility time of treated groups when compared to normal control ones (Fig. 1). Surprisingly, these findings are in contrast with other studies that revealed anti-anxiety potential of HCA containing herbal products [28][29][30]. Patients suffering from schizophrenia experience cognitive dysfunction, social withdrawal, and diminished emotional expressiveness, which manifests as lack of motivation, depression, and anxiety [31,32]. In threechamber social interaction test, GC extract treatment at both doses lead to both reduced sociability and social novelty (Fig. 2). ...
Article
Full-text available
Obesity is a global concern, closely allied with somatic and psychosomatic disorders. Herbal drugs are available in modern medicine to treat obesity. Garcinia camobogia being used by so many people trying to lose weight produces various systemic side effects. The study was conducted to assess its effect on anxiety, sociability, and dopamine turnover in male mice. Twenty-one male Swiss albino mice of either were divided into three groups with seven mice in each group. Control group was given distilled water (0.5 ml p.o.) and the other two groups received Garcinia cambogia extract at two different doses, a low and a higher dose (100 mg/kg and 500 mg/kg. p.o.) Each animal received a single oral dose daily, which was administered using an oral gavage for fourteen consecutive days. Effect of test drugs on anxiety was evaluated using open field test. Sociability and social novelty were evaluated using three chambers test. Results (mean ± SD) were analyzed using one-way ANOVA test followed by Tukey’s test. Garcinia cambogia extract significantly increased the time spent in the corners in the open field test, significantly reduced sociability and social novelty in the three-chamber test, significantly reduced dopamine turnover in the brain with a significant decrease in dopamine metabolite homovanillic acid (HVA) and increased D 2 receptor expression in ventral tegmental area. Garcinia cambogia extract have significant anxiogenic effect along with reduced sociability and social novelty in male mice. Moreover, these effects could be related to the altered dopamine turnover and D 2 receptor expression in mice brain. Article Highlights Chronic used of alcoholic extract of Garcinia campbogia lead to a significant increase in anxiety that was manifested by the reduced time in the center zone and increased immobility in the open field test. Garcinia camobogia chronic administration has a profound impact on sociability and social novelty with a significant decrease in both behavioral patterns compared to the control group. These effects could be attributed to the noticed change in the dopamine turnover in the brain with a significant decrease in dopamine metabolite (HVA) and an upward expression of D2 receptors in return.
... Purported effects of GCE such as appetite suppression in humans via increased levels of serotonin (Ohia et al. 2001), inhibition of lipid synthesis in vivo (Peng et al. 2018), reduction of carbohydrate metabolism in human cells (Yamada et al. 2007), and fat burning via thermogenesis in humans (Campbell et al. 2016) are attributed to its principal component, hydroxycitric acid (HCA) (Figure 1). GCE is approximately 30-70% HCA by weight Sakariah 1998, 2000;Semwal et al. 2015). ...
Article
Garcinia cambogia extract (GCE) is a popular botanical supplement used in weight loss products. Hydroxycitric acid (HCA) is the principal component in GCE. Due to lack of adequate toxicity data to assess the safe use of GCE, the National Toxicology Program is testing GCE in Hsd:Sprague Dawley SD rats following perinatal exposure and in adult B6C3F1/N mice. We report a validated method utilizing sample clean up with ultrafiltration followed by liquid chromatography—tandem mass spectrometry (LC-MS/MS) analysis to quantify HCA in rat plasma across the concentration range from 20 to 800 ng/mL. The method was linear (r² ≥ 0.99) with limits of quantitation (LOQ) and detection (LOD) equal to 20.0 and 3.9 ng/mL plasma, respectively. The accuracy (determined as the relative error, RE) and precision (determined as the relative standard deviation, RSD) using quality control standards analyzed over multiple days were ≤ ± 7.5% and ≤9.5%, respectively. The method was applied to quantify HCA in study matrices (relative error ≤ ± 23.0%; relative standard deviation ≤ 6.0) except gestational day (GD)18 fetus. The method was partially validated in GD18 fetal homogenate across the concentration range from 60 to 3000 ng/g (r² ≥ 0.99, relative error ≤ ± 11.9%, and relative standard deviation ≤ 5.5%; LOQ 60.0 ng/g; LOD 7.77 ng/g). Standards as high as 20,000 ng/mL (plasma) and 502,000 ng/g (fetus) were successfully quantified after dilution into the validated range (relative error ≤ ± 2.6%; relative standard deviation ≤ 5.2%). These results demonstrate that the method is suitable to quantify HCA in rodent matrices and may be adapted to other biological matrices.
... This phytochemical act by increasing the release of 5hydroxytryptamine and serotonin which are implicated in the regulation of eating behavior and appetite control. [3] Citrus aurantium flavonoids (CAF) were studied on the inhibition of adipogenesis and adipocyte differentiation in 3T3-L1 cells. CAF suppressed adipogenesis in 3T3-L1 ...
Article
Full-text available
Life style related diseases like obesity, diabetes and cancer threaten the health worldwide. Attention has been focused recently on the study of relation between the food/plant derived components and their effect on target gene to identify the novel pathway involved in the disorder and their future use as therapeutics. Obesity is a complex multifactorial disorder associated with many health problems including hyperlipidemia, hypertension, type2 diabetes, coronary heart disease and cancer. Obesity is caused by the imbalance between the calories intake and the expenditure that lead to the pathological growth and deposition of adipocytes. Adipocytes play a central role in regulating the adipose mass. In obesity, the increased mass of adipose tissue leads to the adipose tissue hyperplasia, triggering the transformation of preadipocytes to adipocytes. The programmed differentiation of preadipocytes to adipocytes is accompanied by increase in the expression of various transcription factors and adipocyte specific-gene. This review highlights on the studies to explain the mechanism of action of various dietary components interacting with proteins involved in adipogenesis. KEYWORDS: Obesity occurs as a result of genetic and acquired.
... 3 Another study even showed HCA reduced serotonin uptake in mice cortex. 4 Second, HCA has been found to inhibit adinosine triphosphate-citrate lyase, an enzyme involved in the first step of fatty acid synthesis. 5 However, the results from human trials have been mixed. ...
Article
Full-text available
Herbal and dietary supplements are frequently used as weight loss supplements. However, they account for 20% of drug-induced liver injury. Garcinia cambogia's (GC) active compound, hydroxycitric acid, can be found among those supplements. We report a 26-year-old woman who had been taking GC for 7 months when she presented with subacute liver failure and ultimately required a liver transplantation. This report highlights the risk of liver injury after long-term use of GC and demonstrates the importance of considering a close and prolonged monitoring of patients in a tertiary liver transplant center.
... 105 G. cambogia intensifies the release of serotonin from the cortex of the brain of rats. 106 A study reports that exogenous histamine formation in skipjack was prevented by the addition of G. cambogia. 107 Fresh and immobilized biomass of G. ...
Article
The modern popular ideology is that plant-based products do not have adverse effects. Hence, people are fond of using herbal products of their choice to treat their own ailments or anyone else. As majority of the population are not aware of herbal toxicity concept, the use of formulated single or combined medicinal and/or nutritional plant extracts or isolated compounds to treat chronic diseases are increasingly popular due to the widespread concerns regarding the adverse effects of pharmaceutical drugs. Awareness from the scientific community to bring society to its senses regarding the safety issue of a herbal product is rare. Garcinia gummigutta (more popularly known by the synonym Garcinia cambogia in commercial preparations), belonging to the family of Clusiaceae (alt. Guttiferae) is a popularly consumed weight-loss nutraceutical. This review aims to highlight the possible adverse effects of G. cambogia. For the said purpose, 147 articles were collected from PubMed, Web of Science and Google scholar. Literature review revealed a plethora of beneficial actions. Investigational outcomes and clinical evidences hint the possible adverse effects likely to be linked with the use of G. cambogia. However, the use of G. cambogia as an anti-obesity agent is advisable as long as the therapeutic value outweighs the adverse effect.
... The markedly decreased body weight in treated rats can be associated with the reduction in food intake, so this could suggest that the combination of citric acid, garcinol and morelloflavone in G. dulcis rind triggers the appetite satiety mechanisms that inhibit food intake, although there are no studies on these responses. Possible mechanisms include similar responses as hydroxycitric acid which reduced appetite and body weight in human subjects [62], increased release of serotonin from rat brain cortex slices [63] and inhibited serotonin uptake [64]. ...
Article
Full-text available
Metabolic syndrome is a cluster of disorders that increase the risk of cardiovascular disease and diabetes. This study has investigated the responses to rind of yellow mangosteen (Garcinia dulcis), usually discarded as waste, in a rat model of human metabolic syndrome. The rind contains higher concentrations of phytochemicals (such as garcinol, morelloflavone and citric acid) than the pulp. Male Wistar rats aged 8-9 weeks were fed either corn starch diet or high-carbohydrate, high-fat diet for 16 weeks, which were supplemented with 5% freeze-dried G. dulcis fruit rind powder during the last 8 weeks. We characterised metabolic, cardiovascular, liver and gut microbiota parameters. High-carbohydrate, high-fat diet-fed rats developed abdominal obesity, hypertension, increased left ventricular diastolic stiffness, decreased glucose tolerance, fatty liver and reduced Bacteroidia with increased Clostridia in the colonic microbiota. G. dulcis fruit rind powder attenuated these changes, improved cardiovascular and liver structure and function, and attenuated changes in colonic microbiota. G. dulcis fruit rind powder may be effective in metabolic syndrome by appetite suppression, inhibition of inflammatory processes and increased fat metabolism, possibly related to changes in the colonic microbiota. Hence, we propose the use of G. dulcis fruit rind as a functional food to ameliorate symptoms of metabolic syndrome.
... Moreover, in some non-clinical studies in rodents, G. cambogia fruit extracts have also been associated with weight loss and appetite suppression activity, probably as a result of the https://doi.org/10.1016/j.fct.2019.03.051 Received 15 December 2018; Received in revised form 25 March 2019; Accepted 27 March 2019 Abbreviations: AED, antiepileptic drug; AUC, area under the concentration-time curve; AUC 0-24 , AUC from time zero to 24 h; AUC 0-t , AUC from time zero to the last measurable concentration; AUC 0-∞ , AUC from time zero to infinity; AUMC 0-∞ , area under the first moment of the concentration-time curve from zero to infinity; CL/ F, apparent clearance; C max , peak concentration; HPLC-DAD, high-performance liquid chromatography-diode array detection; IS, internal standard; k el , apparent elimination rate constant; LTG, lamotrigine; MEPS, microextraction by packed sorbent; MRT, mean residence time; p.o., per os; t 1/2el , elimination half-life; t max , time to reach peak concentration; V d /F, apparent volume of distribution increase in brain serotonin levels, reduction in plasma insulin levels and inhibition of the enteral absorption of glucose ( Hayamizu et al., 2003;Ohia et al., 2001;Wielinga et al., 2005). Although most studies have shown efficacy on weight loss, in some human studies no significant weight or fat reduction was observed even when high doses of G. cambogia or HCA supplements were used for long periods of time (Kim et al., 2011;Onakpoya et al., 2011;Semwal et al., 2015;Yonei et al., 2008). ...
Preprint
Full-text available
Background: Obesity is a global concern, closely allied with somatic and psychosomatic disorders. Herbal drugs are available in modern medicine to treat obesity. Garcinia camobogia being used by so many people trying to lose weight produces various systemic side effects. The study was conducted to assess its effect on anxiety, sociability, and dopamine turnover in male mice. Methodology: Male Swiss albino mice of either were divided into three groups with seven mice in each group. Different groups were given distilled water (0.5ml p.o.) and Garcinia cambogia extract at two different doses (100mg/kg and 500 mg/kg p.o.). Effect of test drugs on anxiety was evaluated using open field test. Sociability and social novelty were evaluated using three chambers test. Results (mean ± standard deviation) were analyzed using one-way ANOVA test followed by Tukey’s test. Result: Garcinia cambogia extract significantly increased the time spent in the corners in the open field test, significantly reduced sociability and social novelty in the three chamber test, significantly reduced dopamine turnover and increased D2 receptor expression in ventral tegmental area. Conclusion: Garcinia cambogia extract have significant anxiogenic effect along with reduced sociability and social novelty in male mice. Moreover, these effects could be related to the altered dopamine turnover and D2 receptor expression in mice brain.
Chapter
Garcinia cambogia, with its active component hydroxycitric acid (HCA), is widely used for weight loss due to its anorexigenic effect, increased fat oxidation, and regulation of endogenous lipid biosynthesis. The potential effectiveness of G. cambogia in reducing body weight and fat has been concluded in several studies in both experimental animals and humans. However, the results of some randomized placebo-controlled human clinical trials have either not reported the same outcome or there has been a marginal reduction in body weight on a short-term basis, not beyond 12 weeks. Other beneficial effects of Garcinia in experimental studies include antiinflammatory, antiulcerogenic, antioxidant, hepatoprotective, cytotoxic, erythropoietic, and miscellaneous other effects. Studies with G. cambogia conducted in experimental animals have not reported mortality or significant toxicity. Further, at doses usually recommended for humans no side effects have been reported. However, G. cambogia should not be concurrently used with other drugs to circumvent drug interactions. Currently, a large number of G. cambogia/HCA-containing dietary supplements are being marketed, although the reports of toxicity associated with the regular use of these supplements have raised concerns. In most cases complaints have been related to formulations containing multiple ingredients including G. cambogia. Another reason for adverse effects of G. cambogia/HCA can be related to idiosyncratic reactions by individual patients or other drugs used concurrently with these supplements resulting into in a drug/herb-induced toxic interaction.
Article
Full-text available
1.1. Eating disorders can be found in several psychiatric pathologies: schizophrenia, delusional disorder (somatic type), bipolar disorders, major depressive disorder, borderline personality disorder, generalized anxiety disorder, body dysmorphic disorder, somatization disorder and conversion disorder.2.2. Although their clinical features have been defined, relatively little is known about the role of neurobiological patterns in the pathogenesis of these disorders. Several CNS neurotransmitters and neuromodulators are involved in the regulation of eating behavior in animals and have been implicated in symptoms such as depression and anxiety often observed in patients with eating disorders. The authors will review some studies on NA, DA, 5-HT, beta-endorphines, CRH, VP, OT, CCK, NPY and PYY envolved in eating disorders. Furthermore, we will highlight some of the studies on drug therapy of eating disorders taking into account the effects of these agents on neurotransmitters and neuromodulators.3.3. Antidepressant drugs have long been used for anorexia nervosa and bulimia, these disorders been claimed to be affective equivalent. Antidepressant agents seem to be effective in reducing the frequency of binge-eating episodes, purging behavior and depressive symptomatology. It is notable that antidepressant agents have been proved to be effective in patients with chronic bulimic symptoms, even in cases persisting for many years and in patients who had repeatedly failed courses of alternative therapeutic approaches. In all of the positive studies, antidepressant agents appeared effective even in bulimic subjects who did not display concomitant depression.4.4. Few controlled studies on use of medications for anorexia nervosa have been published. Central serotonergic receptor-blocking compounds such as cyproheptadine cause marked increase in appetite and body weight. Zinc supplementation or cisapride could be a therapeutic option in addition to psychological and other approaches in anorexia nervosa.5.5. There is no therapy as yet which is fully effective in alimentary disorders. Psychotropic drugs give some relief from symptoms, but they cannot cure the disorders. An integrated approach, either pharmacological or psychological, is still recommendable.
Article
: Slices of rat caudate nucleus were incubated in a solution of 123 mM-NaCl, 5 mM-KCl, 1.2 mM-MgCl2, 1.2 mM-NaH2PO4, 25 mM-NaHCO3, 0.2 mM-choline chloride, 0.058 mM-paraoxon, 1 mM-EGTA, and oxidizable substrates. (−)-Hydroxycitrate, a specific inhibitor of ATP-citrate lyase (EC 4.1.3.8), used at a concentration of 2.5 mM, inhibited the synthesis of acetylcholine (ACh) from [1,5-14C]citrate by 82–86%, but that from [U-14C]glucose by only 33%, from [2-14C]pyruvate by 24% and from [1-14C-acetyl]carnitine by 8%; the production of 14CO2 from these substrates was not substantially changed. The synthesis of ACh from glucose and pyruvate was in hibited also by citrate; 2.5 mM- and 5 mM-citrate diminished it by 43% and 66%, respectively; the production of from [U-14C]glucose and from [1-14C]pyruvate was not affected. The mechanism of the inhibitory effect of citrate on the synthesis of ACh is not clear; the possibility is discussed that citrate alters the intracellular milieu in cholinergic neurons by chelating the intracellular Ca2+ and decreases the supply of mitochondrial acetyl-CoA to the cytosol. The results with (−)-hydroxycitrate indicate that the cleavage of citrate by ATP-citrate lyase is not responsible for the supply of more than about one-third of the acetyl-CoA which is used for the synthesis of ACh when glucose or pyruvate are the main oxidizable substrates. This proportion may be even smaller, since (−)-hydroxycitrate possibly affects the synthesis of ACh from glucose and pyruvate by a mechanism (unknown) similar to that of citrate, rather than by the inhibition of ATP-citrate lyase.
Article
The effects of (-)-hydroxycitrate (OHC) and citrate on the concentration of acetylcoenzyme A (acetyl-CoA) and acetylcholine (ACh) in the tissue and on the release of ACh into the medium were investigated in experiments on slices of rat caudate nuclei incubated in media with 6.2 or 31.2 mM K+, 0 or 2.5 mM Ca2+, and 0, 1, or 10 mM EGTA. OHC diminished the concentration of acetyl-CoA in the slices under all conditions used: in experiments with 2.5 mM OHC, the concentration of acetyl-CoA was lowered by 25-38%. Citrate, in contrast, had no effect on the level of acetyl-CoA in the tissue. Although both OHC and citrate lowered the concentration of ACh in the slices during incubations with 6.2 mM K+ and 1 mM EGTA, they had different effects on the content of ACh during incubations in the presence of Ca2+. The concentration of ACh in the slices was increased by citrate during incubations with 2.5 mM Ca2+ and 31.2 or 6.2 mM K+, but it was lowered or unchanged by OHC under the same conditions. The release of ACh into the medium was lowered or unchanged by OHC and lowered, unchanged, or increased by citrate. It is concluded that most effects of OHC on the metabolism of ACh can be explained by the inhibition of ATP-citrate lyase; with glucose as the main metabolic substrate, ATP-citrate lyase appears to provide about one-third of the acetyl-CoA used for the synthesis of ACh. Experiments with citrate indicate that an increased supply of citrate may increase the synthesis of ACh. The inhibitory effect of citrate on the synthesis of ACh, observed during incubations without Ca+2, is interpreted to be a consequence of the chelation of intracellular Ca2+; this interpretation is supported by the observation of a similar effect caused by 10 mM EGTA.
Article
Hydroxycitrate (HCA), a popular dietary supplement for weight loss, is a competitive inhibitor of ATP-citrate lyase, an extramitochondrial enzyme involved in the initial steps of de novo lipogenesis (DNL). Although animal studies have shown that HCA effectively inhibits DNL and induces weight loss, these findings have not been consistent in humans. This raises the possibility that the bioavailability of HCA may differ among species. We developed a new GC/MS method to measure HCA levels in blood, using [U-13C]citrate (CA*) as internal standard to account for losses associated with the isolation, derivatization, and measurement of HCA. HCA and CA* were derivatized with BSTFA + 10% TMCS and analyzed using PCI/GC/MS (CA*, m/z 471; and HCA, m/z 553). The plasma HCA concentration was measured over a 3.5-h period in four subjects having ingested 2 g of HCA. Their plasma HCA concentration ranged from 0.8 to 8.4 μg/ml 30 min and 2 h after ingestion, respectively. These results demonstrate that when taken acutely, HCA is absorbed, yet present in small quantities in human plasma. This simple method requiring minimal sample preparation is able to measure trace amounts of HCA with accuracy and precision.
Article
Albino rats of Wistar strain were fed lipogenic diet with and without the addition of (−)hydroxycitrate for a period of 15 days. Daily food intake and weekly body weight gain of the animals were recorded during the feeding period. The animals were then sacrificed, blood, liver and epididymal fat were collected and analysed for selected parameters. It was observed that inclusion of (−)hydroxycitrate in the diet resulted in significant reduction in food intake, body weight, epididymal fat and serum triglyceride in the animals and also decrease in the feed efficiency ratio. The decrease in food intake, body weight gain and feed efficiency ratio brought about by (−)hydroxycitrate was dependent on the content of this compound in the diet.
Article
1 Synaptosomal uptake mechanisms of 5-hydroxytryptamine and dopamine were examined in cerebral cortex, corpus striatum and midbrain plus brainstem of developing rats. 2 In all regions, there was generally a parallel biphasic development of both uptake systems; the most rapid increases occurred in the first two weeks postpartum, followed by a slower rate of increase. 3 Kinetic studies with dopamine indicated that the maturation involved increases in maximal uptake without a change in the substrate Km, suggesting that there is a change in the number or terminals but not in the uptake system per se.
Article
1 The effects of mianserin and of selected tricyclic antidepressants were compared in a number of monoamine uptake models. 2 The ability of mianserin to block the noradrenergic neurone membrane amine pump of rabbit brain stem slices was comparable to that of imipramine and amitriptyline and less than that of desipramine and nortriptyline. Both mianserin and desipramine were competitive inhibitors of noradrenaline uptake in vitro. The effect of mianserin on noradrenaline uptake in vivo was studied both peripherally and centrally. The ability of 6-hydroxydopamine to lower rat heart noradrenaline levels was found to be very sensitive to inhibition by tricyclic antidepressants. Mianserin was active in this model. However, its ability to block the 6-hydroxydopamine-induced fall in rat heart noradrenaline concentration was appreciably less than that of the tricyclics studied. 3 Mianserin, like tricyclic antidepressants, was essentially devoid of effect on dopamine uptake both in vitro and in vivo. 4 The ability of mianserin to inhibit [3H]-5-hydroxytryptamine uptake by rat hypothalamic synaptosomes was appreciably less than that of the tricyclic antidepressants studied. Mianserin was essentially devoid of effect on rat brain 5-hydroxytryptamine uptake in vivo. 5 It is concluded that in certain situations large doses of mianserin may block noradrenaline uptake in vivo. However, in no way does mianserin rival tricyclic antidepressants in blocking monoamine uptake in vivo. The clinical efficacy of mianserin cannot be attributed to inhibition of monoamine uptake.
Article
Anaplerotic reactions leading to the de novo synthesis of fatty acids, were recently proposed to participate in the coupling of metabolic to secretory events in the process of glucose-stimulated insulin release. In an attempt to validate such a proposal, the effect of (-)-hydroxycitrate upon fatty acid synthesis and insulin release was investigated in glucose-stimulated rat pancreatic islets. The inhibitor of ATP citrate-lyase, when tested in the 1.0-2.0 mM range, failed to affect glucose-stimulated insulin release, but also failed to inhibit the incorporation of 14C-labelled acetyl residues derived from L-[U-14C]leucine into islet lipids. A partial inhibition of fatty acid labelling by 3H2O was only observed in islets incubated for 120 min in the presence of 5.0 mM (-)-hydroxycitrate and absence of CaCl2. These findings suggest that (-)-hydroxycitrate is not, under the present experimental conditions, a useful tool to abolish fatty acid synthesis in intact pancreatic islets.