luvabunny":1fdjctsh said:
Michaels4Gardens - how much of the green onion / garlic top do you feed each one? And how often?
My rabbits are generally around 10 lbs as adults.
__________ Sun Sep 28, 2014 12:05 am __________
I just thought of this....the judge at the show is going to flip my rabbit over to view it's teeth and go "ugh! onion breath!"
Sorry...it's late and that just hit me as funny!
I feed about a quarter,to 50 cent size bunch of Chives , onion or garlic tops, about a foot long, so I suppose that would be about 1 1/4 inch diameter size bunch of tops. [for me that is a handful] probly about 1/4 to 1/2 oz
--..for most of the breed stock [dry does and bucks] I have fed it 1x/ week, but --I have fed it every day for 3 weeks when trying to treat a problem. and does with litters out of the box, and eating, I give 2 to 3 bunches, depending on how much they clean up in an hour. [my chores take about an hour,after I feed the critters, so -when I am done with chores, I go through and remove any chives from the cage they have not eaten] usually there is none to be found in the cage after an hour, but when feeding a doe with a litter, I sometimes remove some and toss it to the sheep.
-- ** I always feed the rabbits , or at least make sure the hay mangers are full before I toss the chives in, as I don't want overly hungry rabbits when I feed the chives,[or any green feed] this is to make sure they do not eat more then they really need, as overly hungry rabbits will sometimes gorge them selves on green feeds, and hurt them selves..
__________ Thu Oct 02, 2014 7:40 am __________
michaels4gardens":1fdjctsh said:
luvabunny":1fdjctsh said:
Michaels4Gardens - how much of the green onion / garlic top do you feed each one? And how often?
My rabbits are generally around 10 lbs as adults.
__________ Sun Sep 28, 2014 12:05 am __________
I just thought of this....the judge at the show is going to flip my rabbit over to view it's teeth and go "ugh! onion breath!"
Sorry...it's late and that just hit me as funny!
I feed about a quarter,to 50 cent size bunch of Chives , onion or garlic tops, about a foot long, so I suppose that would be about 1 1/4 inch diameter size bunch of tops. [for me that is a handful] probly 1/4 to 1/2 oz
--..for most of the breed stock [dry does and bucks] I have fed it 1x/ week, but --I have fed it every day for 3 weeks when trying to treat a problem. and does with litters out of the box, and eating, I give 2 to 3 bunches, depending on how much they clean up in an hour. [my chores take about an hour,after I feed the critters, so -when I am done with chores, I go through and remove any chives from the cage they have not eaten] usually there is none to be found in the cage after an hour, but when feeding a doe with a litter, I sometimes remove some and toss it to the sheep.
-- ** I always feed the rabbits , or at least make sure the hay mangers are full before I toss the chives in, as I don't want overly hungry rabbits when I feed the chives,[or any green feed] this is to make sure they do not eat more then they really need, as overly hungry rabbits will sometimes gorge them selves on green feeds, and hurt them selves..
European Journal of Medicinal Plants
2(4): 290-307, 2012
SCIENCEDOMAIN international
http://www.sciencedomain.org
Haematological and Hepatotoxic Potential of
Onion (Allium cepa) and Garlic (Allium sativum)
Extracts in Rats
Enitan Seyi Samson1*, Ajeigbe Kazeem Olasunkanmi1,
Josiah Sunday Joel2 and Ehiaghe Friday Alfred3
1Department of Physiology, School of Basic Medical Sciences, College of Health Sciences,
Igbinedion University Okada, P.M.B. 0006, Edo state, Nigeria.
2Department of Biochemistry, School of Basic Medical Sciences, College of Health
Sciences, Igbinedion University Okada, P.M.B. 0006, Edo state, Nigeria.
3Department of Haematology, School of Clinical Medicine, College of Health
Sciences, Igbinedion University Okada, P.M.B. 0006, Edo state, Nigeria.
Authors’ contributions
This work was carried out in collaboration between all authors. ESS designed the study,
managed the literature searches, wrote the protocol and the first draft of the manuscript,
AKO and JSJ managed the analyses of the study. EFA performed the statistical analysis. All
authors read and approved the final manuscript.
Received 28th April 2012
Accepted 3rd July 2012
Published 12th October 2012
ABSTRACT
Aims: To evaluate and compare the effects of single and combined oral administration of
fresh aqueous onion (Allium cepa) and garlic (Allium sativum) extracts at different
concentrations (200mg/kg/d, 400mg/kg/d and 600mg/kg/d) on some haematological and
hepatotoxicity indicator parameters in treated rats.
Study Design: Cross-sectional nonclinical study in animal model.
Place and Duration of Study: The study was carried out at the Department of
Physiology, Department of Biochemistry and Department of Haematology, College of
Health Sciences, Igbinedion University Okada, Edo state, Nigeria between the month of
July and August, 2011.
Methodology: Following 30 days post-oral administration of extracts in 36 treated male
albino rats as well as 4 controls, haematological parameters were determined using the
Research Article
European Journal of Medicinal Plants, 2(4): 290-307, 2012
291
Sysmex® Automated Haematology Analyzer, while serum levels of liver enzymes were
evaluated using the Reflotron® Plus Auto-Analyzer and liver weight was determined using
electronic sensitive weighing balance.
Results: Red blood cell count (RBC), Packed cell volume (PCV), Haemoglobin
concentration (HGB), Total White blood cell count (TWBC), Absolute Count of Neutrophil
(NEUT#), Absolute Count of Lymphocyte (LYM#), Absolute Count of the summation of
Monocyte, Eosinophil and Basophil (MXD#) and Platelet count (PLT) were significantly
increased (P<0.05) to varied extent, except Mean cell volume (MCV), Mean cell
haemoglobin (MCH) and Mean cell haemoglobin concentration (MCHC) while Alanine
aminotransferase (ALAT) and Aspartate aminotransferase (ASAT) serum levels were
significantly decreased (P<0.001) and liver weight (LW) was non-significantly (P>0.05)
reduced in a dose-dependent manner when compared to the control. Synergistic effect
was not observed in the haematological parameters when the two extracts were
combined.
Conclusion: The results of this study reveal the haematological potential of onion
and garlic extracts with no potential risk of hepatotoxicity (at the concentrations
tested) as earlier anticipated. It also further confirms the higher efficacy of garlic over
that of onion, but the molecular mechanism behind their combined effect would require
further investigation.
Keywords: Onion; garlic; haematological parameters; hepatotoxicity indicator parameters;
rats.
1. INTRODUCTION
The potency of onion (Allium cepa) and garlic (Allium sativum) as medicinal plants, due to
their high content of vitamins, trace elements, amino acids and several organosulphur
compounds has been acknowledged over 5000 years (Balick and Paul, 1996). These
“magic” drugs are well known for their: fibrinolytic effects (Chutani and Bordia, 1981),
hemodynamic and hemostatic effects (Brosche et al., 1990), platelet effects (Apitz-Castro et
al., 1992), immunologic effects (Chisty et al., 1996), lipid-lowering effects (Ide and Lau,
1997), anti-artherosclerotic effects (Koscielny et al., 1999), anti-oxidative effects (Borek,
2001), anti-cancer effects (Milner, 2001), vascular effects (Ashraf et al., 2004), anti-microbial
effects (Benkeblia, 2004), haematological effects (Fazlolahzadeh et al., 2011) and
hepatoprotective effects (Ugwu and Omale, 2011) among other health benefits.
It is quite alarming that the use of some medicinal plants seems to pose some health
hazards despite their overwhelming health benefits either due to their inherent toxic effect or
abuse resulting from over-dosage or prolong usage. Concerns over their safety were raised
after incidents of toxicity were reported following consumption of some Herbal preparations
but the underlying pathogenesis remained cryptic (Favreau et al., 2002; Stickel et al., 2005).
While the health benefits of most herbs have been strongly demonstrated, their safety is
poorly documented, and the awareness of consumers and health professionals towards
herbal preparations as a potential source of health damage is low. More worrisome are
recent reports about adverse effects, following the intake of some herbal preparations
containing ephedrine and green tea extracts resulting in acute and chronic liver injury.
Severe liver injury associated with long-term consumption of Herbalife-products possibly by
a yet unknown profibrotic component was described ranging from reversible cholestatic
European Journal of Medicinal Plants, 2(4): 290-307, 2012
292
hepatitis to acute liver failure requiring liver transplantation, and death from post-transplant
complications (Duque et al., 2007; Elinav et al., 2007).
Controversies exist regarding the health benefit and toxic effect of onion and garlic extracts.
Very little is known about their potential toxic effects on body organs, however garlic has
been shown to cause bad breath and body odor, Irritation of the stomach lining, nausea,
intestinal gas, diarrhea, heart burn (Desai et al., 1990), decrease of serum protein and
calcium (Shashikanth et al., 1986), anemia (Nakagawa et al., 1980) and inhibition of
spermatogenesis (Qian et al., 1986) particularly in people who lack the liver processing
system for detoxifying allicin (Mader, 1990), also significant damage to the epithelial mucosal
membrane, resulting in bleeding, shrinkage and ulcers, after 24 hours of exposure has been
reported in rats, while Onion on the other hand, has been reported to exacerbate
hyperacidity in patients with active gastric and duodenal ulcers and similarly worsen existing
heart burn (Allen et al., 1990). Consuming large quantities of onions can also lead to
stomach distress, gastrointestinal irritation and allergy manifesting as skin rash, redness and
itchy eyes (Valdivieso et al., 1994) particularly in people who lack the liver processing
system for detoxifying N propyl disulphide (toxin, sometimes found in onion and other
alliums).
Though, some studies have shown that regular intake of garlic and onion can help treat
haematotoxicity (resulting from chemicals, microbal or plant toxins in the blood), as well as
potentiate the haematopoietic activities of the bone marrow in the treatment of anemia and
leucopenia, nevertheless, adequate data on the potential risk of hepatotoxicity seems to be
lacking. Causes of hepatotoxicity include viral and parasitic infections, autoimmune diseases
and intoxication with various xenobiotics such as chlorinated solvents, alcohol, fungal toxins,
industrial pollutants, radioactive isotopes, drugs and herbal medicines (Evans, 2002). Liver
injury do occur, not necessarily as a result of over-dosage as earlier reported, but sometimes
it does occur, even when drugs/herbs are used within therapeutic limits. The predominant
pathologies in a given country depend on nature (genetics) and nurture (environment).
A primary purpose of nonclinical studies using animal model is to monitor for the onset of
adverse effects, discover target organ toxicity, determine the margin of safety, determine
whether the risk is acceptable for the therapeutic effect of the drug and for the patient
population and from this information stop the development of the compound or to utilize this
information for monitoring possible toxicities in human studies. Presently, there is a lack of
sufficiently organized data to make an informed conclusion on the predictivity of nonclinical
studies for identifying the risk of significant hepatotoxicity in clinical trials and in the postmarketing
population. Besides, lack of standardization, quality and safety shortcomings of
some herbal preparations and supplements are major concerns of advocates of a tighter
regulation of these products. If the use of herbal remedies must be fully embraced, they
must be subjected to toxicological scrutiny from time to time in order to ensure their safe use
both on short and long term bases.
To address this issue it is necessary to determine whether there are data that would allow
accurate predictions to be made, whether there are early signals in animal studies, that
indicates a drug (in this case, plant extract) will later cause serious hepatotoxicity in humans,
and whether there are signals that indicate that the hepatotoxicity of the drug poses an
acceptable risk. In an attempt to explore the haematological potentials of onion and garlic
extracts, we also see the need to determine if these extracts cause hepatotoxicity at the
varied concentrations (200mg/kg/d, 400mg/kg/d and 600mg/kg/d) tested at the end of 30
days of oral administration. It is hope that these animal data can be extrapolated to predict
European Journal of Medicinal Plants, 2(4): 290-307, 2012
293
and prevent unanticipated hepatotoxicity in man. Besides, even though garlic and onion
extracts have been shown to possess haematological potential in some previous studies, the
controversy regarding which of them exhibit a higher efficacy needs to be investigated and
clarified. And owing to the fact that plausible evidence exist that garlic may interfere with the
action of some herbal remedies in modulating haematopoietic and hepatic activities
(Zlotogorski and Littner, 2004), it becomes paramount to investigate the combined effects of
garlic and onion extracts on some haematological and hepatotoxicity indicator parameters in
experimental models, with a view to establish if the anticipated effects are elicited either
singly or combined and if also dose-dependent. It is hoped that the outcome of this
investigation will provide adequate scientific justification on the continuous use of garlic and
onion extracts as haematopoietic modulating agents with little or no hepatotoxic potential.
2. MATERIALS AND METHODS
2.1 Study Area
This study was carried out at the College of Health Sciences, Igbinedion University Okada
located in Okada wonderland (a commercial, but costly recreational center equipped with
housing venture, comprehensive education institute and 300-bed space medical center) in
Okada, a town beside Benin City (capital city of Edo state), located in the Ova-North East
local government area of Edo state, coordinates: 6o30, N 6o00E in central southern Nigeria.
Predominant occupation among the people is farming and they are noted for the growing of
the following agricultural products: rubber, cocoa, cashew nuts, plantain etc. Despite the
availability of reliable medical service, the local populaces still rely on the use of herbs as
medicines for both curative and prophylaxis purpose.
2.2 Study Design
A total of 40 male albino rats weighing 150±40g (mean±SD) were purchased from the
Animal Production and Health Department, Federal University of Technology, Akure (Ondo
State, Nigeria) and housed in the Experimental Animal House, College of Health Sciences,
Igbinedion University Okada (Edo state, Nigeria) separately in well ventilated wire-bottom
steel cages under hygienic conditions, with proper aeration at 25±2ºC, and a relative
humidity of 45–50%. The rats were randomly assigned into 10 groups of 4 rats each and fed
on standard rat diet (10g/100g body weight) twice daily and tap water ad libitum. Prior to
commencement of administration, the rats were allowed to stabilize in the Animal House with
standard 12-hour light-dark cycle, for a period of 14 days, and were treated for 30 days with
different doses of the plant extracts. All studies on animal experimentation were conducted
in accordance with the Current Animal Care Regulations and Standards approved by the
Institute for Laboratory Animal Research (ILAR, 1996).
2.3 Preparation of Aqueous Onion and Garlic Extracts
A bulk of fresh onion bulbs and garlic cloves (single batch) sufficient for the study was
purchased from a retail store in Okada market (Edo state, Nigeria). Aqueous onion extract
was prepared from fresh onion by modified method of Martha et al., (1998). The light scaly
leaves were peeled with knife and thoroughly washed. 60g of the onion was then weighed,
sliced into tiny pieces and homogenized in 100 ml of cold sterile distilled water. This gave a
concentration of 600mg/ml, on the basis of the weight of the starting material (60g/100 ml).
The homogenates were filtered three times through sterile cheese cloth and centrifuged at
European Journal of Medicinal Plants, 2(4): 290-307, 2012
294
200 xg for 10 minutes and the supernatant was collected and then filtered into separate
sterile containers using a funnel containing sterile Whatman No. 1 filter paper for
clarification. The liquid filtrates were transferred into sterile Universal bottles. After
preparation of the crude extracts as described; additional concentrations (400mg/ml and
200mg/ml) were made from the stock (600mg/ml) with sterile distilled water. All these were
done aseptically. Protocol was repeated for garlic. It was ensured that fresh plant extracts
were prepared each day from the single batch of onion and garlic samples purchased before
administration as the active ingredients are well known to lose most of their potency with
time after extraction when exposed to higher temperature; otherwise, extracts were stored in
the refrigerator at 4ºC to maintain their potency.
2.4 Animal Treatment
A total of 40 rats were randomly assigned into 10 groups (n=4/group) and treated as shown
in Table 1.
Table 1. Experimental pharmacological protocol for onion and garlic treated rats
Groups (n=4) Treatments
G1 Control Group received no extracts at all
G2 200mg/kg/d aqueous extract of onion
G3 400mg/kg/d aqueous extract of onion
G4 600mg/kg/d aqueous extract of onion
G5 200mg/kg/d aqueous extract of garlic
G6 400mg/kg/d aqueous extract of garlic
G7 600mg/kg/d aqueous extract of garlic
G8 200mg/kg/d combined aqueous extract of onion and garlic
G9 400mg/kg/d combined aqueous extract of onion and garlic
G10 600mg/kg/d combined aqueous extract of onion and garlic
The volume of extract (2ml/100g body weight) to be administered to individual rat in each
group using intragastric tube was calculated, recorded and adjusted daily with changes in
body weight throughout the treatment period which lasted for 30 days. They were observed
daily for any observable change. Overnight prior to euthanasia, the animals were starved of
food. The animals were sacrificed by cervical dislocation as described by Ochei and
Kolhatkar (2006). Cardiac blood specimen was taken from each rat by terminal bleeding
from the heart. The first half of the blood collected was transferred into a clean edtacontainer
(thoroughly mixed) ready for evaluation of haematological parameters while the
second portion was transferred into an anticoagulant-free test tube and allowed to clot and
subsequently centrifuged at 750 g for 15 min to obtain serum component ready for
evaluation of serum level of liver enzymes.
2.5 Evaluation of Haematological and Hepatotoxicity Indicator Parameters
Haematological parameters were determined using the Sysmex® Automated Haematology
Analyzer kx-21n, Sysmex Corporation, (Kobe-Japan) as described by Samuel et al. (2010).
while, the serum level of alanine aminotransferase (ALAT) and aspartate aminotransferase
(ASAT) were evaluated photometrically with Reflotron® Plus auto-analyzer (Boehringer,
Mannheim, FRG, Roche, Germany) according to the method as described by Deneke and
Rittersdorf (1984, 1985). The auto-analyzers were operated as instructed in the user’s
European Journal of Medicinal Plants, 2(4): 290-307, 2012
295
operational manual. Following dissection of the peritoneal cavity, the liver was isolated as
described by Fricker et al. (1994) and weighed with the aid of electronic sensitive analytical
balance.
2.6 Statistical Analyses
All numerical results were obtained from the ten (10) groups (control and treated). Data were
presented as mean±SEM and analysed using one way analysis of variance (ANOVA) and
Tukey-Kramer Multiple Comparisons Test using SPSS-18.0 (Statistical packages for social
Scientists – version 18.0) statistical program. P values<0.05 were considered significant.
3. RESULTS AND DISCUSSION
RBC, PCV, HGB, TWBC, NEUT#, LYM#, MXD#, and PLT were significantly increased, while
MCV, MCH and MCHC were significantly decreased to different extent (P<0.05) in a dosedependent
manner in both onion and garlic treated groups when compared to the control.
Extract of garlic at 600mg/kg/d proved to be the most potent while the combined extract of
onion and garlic at 200mg/kg/d was the least effective. A partial antagonistic effect on
haematological indices was observed when the two extracts were combined (Table 2). The
serum level of Liver enzymes (ALAT and ASAT) were significantly decreased (P<0.001) in a
dose-dependent manner in both onion and garlic treated groups when compared to the
control. The extracts appear to be non-hepatotoxic when used singly and combined.
Meanwhile, a non-significant decrease (P>0.05) in liver weight (LW) was observed in the
onion, garlic and combined and treated groups when compared to the control (Table 3).
The outcome of this present study shows that extracts of fresh garlic and onion (especially at
higher concentration) have significant (P<0.05) impacts on some haematological parameters
as evident by the active proliferations of blood components to varied extent as measured in
the treated groups, compared to the control. While, the significant increase (P<0.001) in
RBC, TWBC and PLT counts observed in this present study did not agree with that of
Michael et al. (2009) who reported a significant decrease (P<0.05) in total WBC and Ugwu
and Omale (2011), who reported a non-significant decrease (P>0.05) in the WBC and RBC
counts; it does agrees with the works of: Iranloye (2002), who reported increases in total
leucocyte count, neutrophils, lymphocytes, monocytes, RBC and Hb concentration in rats
following 30-days of garlic consumption; Sahu (2004) observed an increase in erythrocytic
count and Salah et al. (2008) reported a significant increase in leucocytic count after
administering garlic.
It also strongly confirms the findings of Tatfeng and Enitan (2012) who reported significant
increases in total leucocyte count, absolute count of lymphocytes, neutrophils and
summation of monocytes, eosinophils and basophils in their work on effects of onion and
garlic extracts on some immunologic cells.
European Journal of Medicinal Plants, 2(4): 290-307, 2012
296
Table 2. Haematological parameters of rats after onion, garlic and combined extracts were administered
Automated Method (Samuel et al., 2010)
Keys: CONC. = Concentration, RBC= Red blood cell count, PCV= Packed cell volume, HGB= Haemoglobin concentration, MCV= Mean cell
volume, MCH= Mean cell Haemoglobin, MCHC= Mean cell haemoglobin concentration, TWBC= Total White blood cell count, NEUT#= Absolute
neutrophil count, LYM#= Absolute lymphocyte count, MXD#= Absolute count of the summation of monocyte, eosinophil and basophil, PLT= Platelet
count, O+G = combined extracts of onion and garlic. All values are expressed in Mean ±SEM of 4 animals. Group values differ significantly from
control at 5 percent level (a = P<0.05), 1 percent level (b = P<0.01), 0.1 percent level (c = P<0.001) and d= P>0.05 (Not Significant).
Group
Parameters
1 2 3 4 5 6 7 8 9 10
CONTROL 200mg/
Kg
ONION
400mg/
Kg
ONION
600mg/
Kg
ONION
200mg/
Kg
GARLIC
400mg/
Kg
GARLIC
600mg/
Kg
GARLIC
200mg/
Kg
O+G
400mg/
Kg
European Journal of Medicinal Plants, 2(4): 290-307, 2012
297
Table 3. Hepatotoxicity indicator parameters of rats after onion, garlic and combined extracts were administered
Group
Indices
1 2 3 4 5 6 7 8 9 10
CONTROL 200mg/
Kg
ONION
400mg/
Kg
ONION
600mg/
Kg
ONION
200mg/
Kg
GARLIC
400mg/
Kg
GARLIC
600mg/
Kg
GARLIC
200mg/
Kg
O+G
400mg/
Kg
O+G
600mg/
Kg
Automated method (Deneke and Rittersdorf, 1984 and 1985)
Keys: CONC. = Concentration, ALAT = Alanine aminotransferase, ASAT = Aspartate aminotransferase, LW= Liver weight, O+G = combined
extracts of onion and garlic. All values are expressed in Mean ±SEM of 4 animals. Group values differ significantly from control at 1 percent level
(b = P<0.01), 0.1 percent level (c = P<0.001) and d= P>0.05 (Not Significant).
European Journal of Medicinal Plants, 2(4): 290-307, 2012
298
Still, the significant increase observed in the PCV of rats in this present study, concur with
that of Salah et al., (2008) who reported a non-significant increase in PCV values of treated
fish (Oreochromis Niloticus), but, however, contradicts the results of Banerjee and Maulik
(2002), Adebolu et al. (2011) and Ugwu and Omale (2011), who all observed significant
decreased PCV in the treated rats when compared with that of the control and thereby
conclude that garlic is toxic to red blood cells and at very high concentration may induce
anaemia in animals on prolong feeding.
Meanwhile, red cell indices (MCV, MCH and MCHC) on the other hand, are particularly
important for the diagnosis of anemia in humans and most animals. Significant decreases
(P<0.01) in these red cell indices as observed in the treated rats in this current study is in
agreement with the findings made by Corzo-Martinez et al., 2007), who similarly observed a
significant decrease of MCH and MCV in fish fed on the highest dose of Allium sativum. It is
assumed that the decrease or increase of blood indices may be attributed to a defence
reaction against garlic or onion, which occurs by stimulation of erythropoiesis.
According to Lampe (1999), the mechanisms of action under-pining these diverse
haematological effects of plant extracts remain disjointed and relatively poorly understood.
Nevertheless, postulated mechanisms of action supposedly revolve around their stimulatory
effects on some cytokines, their role in iron bio-availability, and presence of some vitamins,
essential amino acids and phytochemicals.
First, garlic and onion compounds seem to have a stimulatory effect along particular
pathways on some haematopoetic growth factors (cytokines) which interact with specific
receptors on the surface of haematopoietic cells, regulating the proliferation and
differentiation of progenitor cells and the maturation and functioning of mature cells. Normal
erythropoiesis for instant; is dependent on there being adequate amounts of erythropoietin
among other factors. Chemical components of garlic and onion seem to act as active oxygen
scarvenger in vivo, It is thus possible that they compete with haemoglobin in the RBC for
oxygen resulting in tissue hypoxia which in turn stimulates the kidney directly to cause
formation and secretion of erythropoietin or that the end product of metabolism of garlic and
onion in the body may step up Hb synthesis and RBC production by their indirect effect on
erythropoietin. The hormone erythropoietin is crucial to erythropoesis by increasing the
number of erythrocytes precursors. This is made possible by stimulating stem cells to
transform into pro-erythroblast (the first morphologically recognisable cell of the erythrocyte
series). Production of erythrocytes is restricted to the bone marrow in adult life, however
Akgul et al. (2010), reported that dietary garlic can enhance erythropoiesis in the spleen (i.e
garlic-induced extramedullary haemopoiesis) but not in the bone marrow. Stimulation of
splenic erythropoiesis gene expression in mice by garlic components is most likely mediated
by hypoxia which steps up the production of erythropoietin. Similarly, effects of garlic and
onion extracts on leucopoiesis may be attributed to their ability to stimulate production of
some colony stimulating factors (CSF) like the Granulocyte-CSF, Monocyte-CSF and
Granulocyte-Macrophage-CSF, or it may be the effect of their end product of metabolism on
the regulation of the cytokine production system, maintenance of the normal systemic
defence and bio-regulation. This may explain why these cytokines are used in boosting
immune function and in the treatment of some infectious diseases. These soluble
membrane-bound glycoproteins of the hematopoietic microenvironment are produced in the
cells lining of the blood vessels and they serve to stimulate the bone marrow to synthesize
white blood cells. They activate intracellular signaling pathways that can cause the myeloid
and lymphoid cells to proliferate and differentiate into a specific kind of white blood cells
(Albert et al., 2002). Garlic has been found to stimulate proliferation of several white blood
European Journal of Medicinal Plants, 2(4): 290-307, 2012
299
cells lines and induced the infiltration of tumours by white blood cells such as natural killer
cells and macrophages (Kandil et al., 1987). Mean while, the effects of garlic and onion
extracts on thrombopoiesis may be attributed to their ability to stimulate production of
thrombopoietin. This cytokine (also known as megakaryocytes growth and development
factor) is a glycoprotein hormone produced mainly by the liver (parenchymal cells and
sinusoidal cells), the kidney (proximal convoluted tubule cells) and bone marrow (stromal
cells). It is similarly believed that the chemical components of garlic and onion or the end
products of their metabolism seem to act on thrombopoietin which is responsible for the
production, proliferation and maturation of megakaryocytes (megakaryocytopoiesis) and
differentiation of megakaryocytes into large numbers of platelets (thrombopoiesis). Reduced
platelet number (thrombocytopenia) in the blood circulation is often therapy resistant in
individuals with AIDS; yet, Platelets numbers have been reported to increase, sometimes
greater than 100 000 during 4 months of garlic supplementation.
Second, it is also believe that both garlic and onion compounds help to improve iron
metabolism. The bone marrow produces over 2 million RBC per seconds. This tremendous
effort is only possible when there is enough iron available, because it is needed to produce
the haem component of haemoglobin and it is therefore crucial to synthesis of haemoglobin.
Besides, it also assists in cellular immune response by stimulating the production of T-cells
in our immune system. There is considerable variation in the availability of iron for absorption
in different foodstuffs. Haem iron from animal origin is said to be easily absorbed than nonhaem
iron of plant origin. However, recently, some Researchers in India found that
compounds in both onion and garlic seem to have a promoting influence on the
bioavailability of iron from food grains. These alliums, they believe can help the body absorb
more dietary iron from cereals and pulses than it otherwise would. This is possible because
the diallyl sufides in garlic and onion may induce ferroportin mRNA, this can result in
increased production of ferroportin located on the basolateral surface of gut enterocytes and
the plasma membrane of reticuloendothelial cells. Ferroportin is an iron channel, a protein
that runs across the cell membrane, forming a passageway that allows stored iron to leave
the cells and become available for the synthesis of haemoglobin. This is important because
the bio-availablity of iron from plant foods like cereal is low. Research has shown that if
onion and garlic are consumed alongside cereal, the body absorbs up to 70 percent more
iron from the cereal. This discovery has the potential for global health, as non-availability of
iron for Hb production leads to iron deficiency anaemia, a serious problem worldwide. It is
therefore assumed that the extracts of garlic and onion administered in this current study
might have facilitated the absorption of more dietary iron (30-35mg/kg) present in the
pelletized feed upon which the rats were fed, thereby stepping up Hb synthesis and resulting
in an elevated haemoglobin concentration in the treated group compared to the control. It is
also speculated that garlic and onion compounds may inhibit hepcidin mRNA responsible for
the transcription of hepcidin (an iron shuttling protein, critical to iron homeostasis, which
prevents absorption of iron by binding to ferroportin), resulting in little or no production of
hepcidin. The lower the concentration of hepcidin, the more iron channels that are opened
for more iron to be absorbed by the hepatic portal system and can be made available for Hb
synthesis. Anemia of inflammation has been linked to high activity of hepcidin (Papanikolaou
et al., 2005).
Third, Fenwick and Hanley (1985) reported that several vitamins like vitamin B1, B2, C and E
are present in both garlic and onion while vitamin B6 and B9 are exclusively found in garlic. Of
particular interests are those that help with red blood cell formation and maintenance. For
instance, deficiency of Vitamin B2 (Riboflavin) has been reported to results in a decrease in
red cell glutathione reductase activity, since this enzyme requires flavin adenine dinucleotide
European Journal of Medicinal Plants, 2(4): 290-307, 2012
300
for activation (Powers et al., 1983). Glutathione reductase is an enzyme that reduces
glutathione disulfide (GSSG) to the sulphydryl form (GSH), which is an important cellular
antioxidant scavenging reactive oxygen species in cells like red blood cells. It plays an
important role in protecting Hb, red cell enzymes and biological cell membrane against
oxidative damage by increasing the level of reduced glutathione (GSSGR) in the process of
aerobic glycolysis. The enzyme deficiency may result in mild to moderately severe
haemolytic anemia upon exposure to certain drugs or chemicals. Still, Researchers at the
University of Maryland Medical Center, believe that cloves of garlic contain vitamin B6
(Pyridoxine) and deliver around 6% of the pyridoxine require per day by the average adult. A
hypochromic microcytic anemia has been associated with Vitamin B6 deficiency induced in
children. While others believe that garlic in addition, contain trace amount of folic acid
(vitamin B9) which is essential for DNA synthesis and are therefore needed by all dividing
cells in the body, particularly haematopoietic cells in the bone marrow. Deficiency of folate
leads to the death of many precursor cells in the bone marrow resulting in megaloblastic
anaemia. Vitamin C (ascorbic acid) is believed to help in the absorption of non-haem iron
from plant origin for the synthesis of Hb. Deficiency of dietary Vitamin C is associated with
deficiency of dietary iron, because Vitamin C serves to facilitate intestinal iron absorption.
Combination of iron and Vitamin C is needed to correct a hypochromic microcytic anemia.
Besides, Vitamin C is required for the maintenance of folic acid reductase in its reduced or
active form. Impaired folic acid reductase activity results in an inability to form tetrahydrofolic
acid, the metabolically active form of folic acid. It also serves to prevent the irreversible
oxidation of methyltetrahydrofolic acid to formylfolic acid. Failure to synthesis tetrahydrofolic
acid or protect it from oxidation ultimately results in the appearance of a megalobastic
anemia. While Vitamin E (tocopherol) strictly serves as an antioxidant; its role had been
demonstrated in its deficiency to compensate for genetic defects that limit the eryhtrocytes’s
defense against oxidant injury. Vitamin E has been found to lengthen the red cell life span
and increase level of haemoglobin. Deficiency of Vitamin E may result in haemolytic
anaemia, associated with morphologic alterations of the eryrocytes and more irreversibly
sickled cells in children with sickle cell anemia (Fishman et al., 2000).
Fourth, garlic and onion have been screened to possess some important phytochemicals
such as flavonoids, steroidal glycosides, alkaloids, saponins, tannins, phenolics, pectin,
amino acids etc, with their biological and physiological roles well documented by Fenwick
and Hanley (1985). They are believed to play a crucial a role in gene expression, enzyme
activity, stimulation of the immune system and organs related to blood cell formation
particularly the bone marrow as reported by Jeorg and Lee (1998). Flavonoids, glycosides
and alkaloids in particular act as antioxidants. They have been demonstrated to possess
good free radical scavenging properties, protecting the red blood cells against oxidative
damage. Addition of extract of aged garlic to red blood cell suspension prior to the addition
of an oxidant has been found to minimize oxidation and cell ruptures. About 0.07% of Steroid
saponins found in garlic have been shown to have antinutritive effects, particularly mid
chronic hemolysis without anaemia resulting in accelerated RBC turnover. However, as the
dosage of saponins (garlic) exceeds the rodent’ erythropoeitic capacity, anemia may ensue.
However, there appears to be no evidence of harmful effects of Alllium saponins in humans
(Sparg et al., 2004). According to Matsuura, (2001) and Oboh, (2004), accelerated RBC
turnover has been proposed as the crux of the mechanisms by which garlic sustains and
amplies its multiple biological effects. As garlic enhanced rates of turnovers, each degraded
RBC may release up to approximately 6X108 heme molecules, of which a significant
proportion are then converted to CO, to alleviate any heme toxicity. Normally, CO is
produced through metabolic processes and CO by itself can stimulate increase in RBC
counts, Hb content and PCV level. It is believe that metabolism of garlic likely result in
European Journal of Medicinal Plants, 2(4): 290-307, 2012
301
production of extra CO resulting in accelerated RBC turnover. Besides, the free amino acids
(mainly arginine) present in garlic and onion is also believed to be contributory to the aminoacid
pool needed for the synthesis of Hb. The globin part of the Hb molecule consists of
chains of amino-acids. The larger the amino-acid pool is, the more chances there are that
more Hb will be synthesized and more RBC produced (Block, 1985).
On the other hand, the complex chemistry of alliums necessitates assessments of their
potential toxic effects on body organs, particularly the liver because of the crucial role it plays
in drug-detoxification. The liver is the central site for the biotransformation of xenobiotics and
therefore is involved in the detoxifying mechanism of the body. It is responsible for
detoxifying the chemical substances in the blood and in this process it is exposed to high
concentrations of toxicants and toxic metabolites making it susceptible to injury. Recently,
liver injury after consumption of herbal remedies was reported but the underlying
pathogenesis remained cryptic (Evans, 2002).
In Clinical chemistry, indicator parameters routinely implemented during nonclinical
assessments for hepatotoxicity include: ALAT, ASAT (Hepatocellular leakage enzymes),
bilirubin, ALP, GGT (Cholestasis indicators), albumin, urea, nitrogen (Function indicators),
electrolytes, total CO2, glucose, triglyceride, and cholesterol (Metabolism indicators) among
others. Increases in these indicator parameters, even in the absence of histologic changes,
are considered adverse, unless the pathogenesis indicates to the contrary. It is difficult to
identify an absolute cut-off between ‘adverse’ and ‘not adverse’ for an individual parameter.
However, certain findings in individual parameters are considered adverse, unless they can
be associated with a mechanism that is non-adverse. It is known that serum level of liver
enzymes reflect liver cell function and integrity and increase in their levels indicate hepatic
cellular damage. The biochemical analysis of the liver can provide some valuable
information on the status or condition of the liver. >3-5X increase in the serum level of ALAT
and ASAT in particular, has been found to be associated with hepatocellular degeneration
and hepatocellular necrosis (including apoptosis) respectively (Ballet, 1997; Zbinden, 1991).
In this present study, the serum levels of ALAT and ASAT were significantly reduced
(P<0.001) in the treated group following 30 days administration of onion and garlic extracts
when compared to the control. Here, a rise in the serum level of the liver enzyme (i.e
transminitis) as was initially anticipated was absent, it can therefore be inferred that the
extracts did not cause hepatotoxicity at the concentration tested. The outcome of this
present study, appear to agree with the results of other similar studies: Hamlaoui-Gasmi et
al. (2011) reported a decrease in the plasma level of transaminases following oral treatment
of garlic (5g/Kg bw) against intra-peritoneal route treatment which resulted in hepatotoxicity
as indicated by the elevation in the enzymes level; Sankaran et al. (2010) showed that
administration of raw garlic significantly decreased the activity of markers enzymes in
alcoholic patients with severe liver damage; El-Demerdash et al. (2005) also found out that
the activities of ASAT, ALAT, LDH, AlP and AcP were significantly (P<0.05) decreased in the
liver tissue of alloxan-diabetic rats fed garlic when compared to the control values; Augusti et
al. (2001) reported that lipid parameters and enzyme activities (ASAT, ALAT and ALP) in rat
serum decreased significantly when they were fed a diet containing 5% garlic; while Ohaeri
(2001) showed that the levels of various enzyme in the serum and tissue of streptozocin
diabetic rats were significantly reduced by garlic oil.
Plasma concentration of liver enzymes is normally low, but is elevated in the event of
hepatotoxcity, which may occur in alcohol, drug or herb-induced liver damage leading to
necrosis of hepatocytes, increased permeability and leakage of cellular enzymes from the
European Journal of Medicinal Plants, 2(4): 290-307, 2012
302
liver cytosol into the blood stream. The main value of the Aminotransferases is in detecting
hepatocellular damage and monitoring the patient's progress as the levels rapidly return to
normal following resolution of the factors causing hepatocellular damage.
The elucidating mechanism of action behind the non-toxic effects of onion and garlic extracts
is also not fully understood and the wide discrepancy in the literatures may be due to many
factors such as the species of garlic and onion used its concentration, route of administration
or the nutritional status of rats. However, most researchers believe that the presence of
essential nutrients like the vitamins, trace elements and phytochemical contents are
responsible for their inherent-antioxidant activities as reported by Rahman (2003) and Yang
et al. (2004). These nutrients seem to chelate metal ion, inhibit lipid peroxidation, increase
the amount or increase the activity of antioxidant and detoxifying enzymes thereby
accelerate the regenerative capacity of the hepatocytes (under the influence of hepatocyte
growth factor), cause stabilization of the cell membrane of the hepatocytes and ultimately
protect the liver cells against deleterious agents and free radical-mediated toxic damages.
According to Gibson (1998), high concentration of fructans (35-40% dry weight), which
constitute a major portion of the water-soluble carbohydrates in onion also contribute to the
reduction of toxic metabolites and detrimental enzymes, protection of liver function and
reduction of serum cholesterol. Another side to it; is that hepatotoxins (like N propyl
disulphide) might be entirely absent or seem to be present in a very low concentration (i.e,
not significant enough to induce hepatotoxicity) in the extracts or that the liver is well fortified
to detoxify them. The ability of the extracts to induce a significant decrease in liver enzymes
activities hereby suggests that: (1) they can be considered safe and non-hepatotoxic. (2)
They pose no risk of hepatotoxicity at the concentration tested, however the use of higher
dosage is recommended for further investigation in the future.
There are issues beyond the scope of this study that require further investigation. We hereby
recommend that the toxic doses of these extracts be properly determined, that experimental
subjects be exposed to extracts for a longer duration (>30days), the number of animals used
in toxicity studies be increased, test for sex and specie differences in order to ease the
detection of hepatotoxicity occurring at a low incidence. Also, an assessment of the
histological profiles of the liver is very crucial, as liver architecture will help to elucidate the
result of the liver enzymes. Besides a better understanding of the mechanism(s) of liver
toxicity as well as the underlying reasons why nonclinical studies fail to prevent compounds
which produce serious human hepatotoxicity from proceeding in the clinic could result in
developing a more predictive nonclinical testing strategy. Where available, technologies
such as In Vitro Cellular Assays, Covalent Binding or Toxicogenomics/Proteomics should be
considered for delineating the mechanisms of hepatotoxicities. Such methods could aid in
identifying the molecular pathways of hepatotoxicity, covalent binding of compounds or
metabolites to proteins and possibly neoantigens (produced from drug treatment) eliciting
immune mediated toxicity. Their utility needs to be further assessed.
It should also be bore in mind that nonclinical studies like this do not provide adequate
assessment of all hepatotoxic liabilities in man. It is particularly important to understand
unique factors responsible for human differences from laboratory animal models and to
modify models or testing strategies to account for such differences. Such factors include:
genetic variability, lifestyle factors, formation of unique metabolites with hepatotoxic
potential, immune mediated events, drug-herb or drug-food or drug-environment
interactions, or endogenous or exogenous factors that modify or compromise organ or tissue
function. Further studies will also require the isolation and characterization of potential
hepatotoxins in garlic and onion extracts and preparations. Since, adequate data on toxic
European Journal of Medicinal Plants, 2(4): 290-307, 2012
303
doses seems not to be available; patients’ serum level of liver enzymes must be evaluated
and monitored continuously in order to avoid the potential risk of hepatotoxicity associated
with long term use of garlic and onion components. Besides, herb-drug interaction has been
well reported, for instance, both garlic and acetaminophen are processed by the same set of
enzymes in the liver when they are taken together, large amounts of garlic may interfere with
the break of acetaminophen and as a result too much acetaminophen may stay in the blood,
for this reason, patients are therefore advised to check with their doctors first before taking
onion and garlic preparation. Critical assessment of their liver function status must be carried
out and those with history of liver problem should be properly informed on the impact of
herbal-drug interaction on their health.
Regarding the degree of efficacy, this present study showed garlic to be more potent than
onion. Extracts of fresh garlic, particularly at the highest dose tested (600mg/kg/d) were
shown to be more effective than that of onion; this confirmed a higher efficacy of garlic as
reported by Micheal et al. (2009) who observed a significantly higher effect in garlic than
ginger; Tatfeng and Enitan (2012) who also observed a significantly higher effect in garlic
than onion. The significantly higher effect (P<0.01) on RBC and Total WBC counts observed
in garlic treated rats at various concentrations tested contradicts the works of Ugwu and
Omale (2011), who show that there were no significant differences (P>0.05) between the
garlic and onion extracts consumption on the RBC and Total WBC counts. While the nonsignificant
difference (P>0.05) in PCV between the garlic and onion treated rats strongly
support the findings of the same on equal treatments. Lesser effects as observed in onion
treated rats may be due to the presence of the extracted phytochemicals in lower degrees or
concentrations, since the same phytochemicals have been found to be also present in garlic
extracts. It is interesting to know that the effects of these spices on haematological
parameters were dose-dependent as higher values were obtained for haematological
parameters measured (except, MCV, MCH and MCHC) with increasing concentration of the
extracts administered compared to the control; suggesting that there may be a correlation
between these responses and dosage. This work agrees with the findings of Banerjee et al.
(2002) and Tatfeng and Enitan (2012).
The combination of both onion and garlic extracts does not appear to be really beneficial to
haematopoietic activities; as some of the haematological parameters were significantly
reduced compared to the single outcome of each extract. The reason for this is not clear,
however, it is possible that the action of some organosulphur compounds found exclusively
in garlic counteract those of onion resulting in a partial antagonistic relationship between
them. For example, quercetin is the main flavonol in onions while myricetin is the main
flavonol in garlic. These two compounds differ slightly in structure and it is possible that they
both compete for the same available receptors on the haematopoietic factors. The possible
mutual opposition in action between them may have resulted in a lesser or reduced
haematological effect as observed in this study. The molecular mechanism behind such
herbal–herbal interaction needs further investigation.
The non-significant decrease (P>0.05) in liver weight (LW) observed in the treated groups
when compared to the control, seems not to agree with the result of Abdullah et al. (2001),
who observed a significant decrease (P<0.05) in the weights of liver, heart and spleen of rats
administered aqueous extract of garlic (100mg/kg/day) for 90 days; it is therefore
recommended that, the dosage and the cumulative length of administration, which are
known to cause both toxicity and atrophy of the hepatocytes and the need for withdrawal or
dosage adjustment based on plasma level, must be considered in order to optimize the
European Journal of Medicinal Plants, 2(4): 290-307, 2012
304
medicinal benefits of these extracts. This is a challenge for the present and future
generations of hepatologists and therefore warrants further investigation.
4. CONCLUSION
The outcome of this current study further confirms the inherent-haematological and nonhepatotoxic
potentials of these extracts and therefore they can serve as natural haematinics
and antioxidants to step up haematopoiesis without risk of potential hepatotoxicity at the
concentrations tested. This study further strengthened the earlier works on the efficacy of
these extracts in this regard.
What I don't know is how much I could really feed without adverse effects , as this study used a very low dose [as far as I can figure = .02 OZ/ Kg / day [ a 10 lb rabbit would get 0.1 oz/ day]] but it is significant to see that there was no evidence of anemia or any other negative outcome, in the rats tested, and- in this study garlic and onion were administered by mouth and not put inside of the intestinal cavity with a needle , so it is much more like something we would do when feeding our rabbits.
