1. INTRODUCTION:
Since the past decades, people have been dependent
on folk and recently herbal medicine for caring variety of diseases (1). They didn’t
care about the possible toxic effects of medicinal plants. Saffron is the long orange–red
dried stigma of Crocus saflower, is the most expensive spice in the world due to
its production costs (2). About 68 kg of flowers are required to make one kg of
saffron (3). Saffron (powder of deride stigma of Crocous sativus L) is habitually
used in many countries like Iraq and Iran as a spice by sprinkling it on cooked
rice. Pharmacological studies have revealed the extract of saffron has various therapeutic
effects like anticonvulsant (4), antidepressant (5), antinociceptive (6) and antioxidant
(7). Saffron as a medicinal plant has a wide range of therapeutic and adverse effects
(8). Last decade, there is the tendency for go into details of the plant’s adverse
and toxic effects (9).
Toxicological studies had shown that the LD50
of saffron stigma in mice is 1.6g/K.g.BW, but certain studies of sub-acute exposure
showed a decrease in hematocrit, erythrocyte and hemoglobin values, while it didn’t
cause pathological lesion in different organs (10). Nikethamide is a CNS stimulant
which induces convulsion (11). Since saffron is habitually used by many people worldwide,
we are concerned to study if there are possible toxic effects, because the safety
of saffron is very important in relation to their habitual consumption and medicinal
use.
2. MATERIALS AND METHODS:
2.1. Plant:
The powder of saffron stigma was purchased from Novin Zeferan
Co (Mashhad, Iran).
2.2. Preparation of methanolic extract of saffron by maceration method.
Briefly, 10 gm of stigma powder was macerated in 200 ml
of 80% v/v of methanol for 72 hours. The solution was subsequently filtered and
concentrated under negative pressure at 40°C.The extraction yield was 55%.
2.3. Animals:
Male Balb-C Mice with a body weight ranged 25-30 gm were
obtained from the animal house of the college of veterinary medicine, University
of Baghdad and were bred in an air- conditioned room (25°C ±1°C) with a 12: 12 hours
light-dark cycle. Standard pellet diet and water were provided ad –labitum.
2.4. Experimental Design: -
This work was performed with the approval of the College
of Veterinary Medicine/University of Baghdad in accordance with international ethical
standards of research for work with laboratory animals.
2.5. Saffron Dosage Solution: -
Each mouse was dosed 0.1 ml/10 gm body weight, by fitting
the concentration which contained the estimated and calculated doses in all experiments
included in this study.
2.6. Acute Toxicity assessment:
Range finding study (12) is carried out for the evaluation
of acute toxicity of methanolic extract of saffron. Twenty male Balb-C mice were
divided equally into four groups administered different methanolic extract doses
subsequently (2000, 3000, 4000, 5000) mg/Kg.BW by gastric gavage needle. Death considered
the criterium for determining the acute toxic after 7 days of dosing. Further three
animals were dosed with 5000 mg/Kg.BW of the plant extract subsequently and observed
for the same period.
2.7. Assessment of effective dose50% of methanolic extract
of saffron stigma as anticonvulsant:
At the first we had conducted a pilot study to choose the
dose of Nikethamide, (N, N-Diethylnicotinamide) a package of 25 ml contained 25gm
obtained from Sigma-Aldrich that only resulted in convulsion without death. Seven
subsequent doses (100, 120, 140, 160, 180, 200, 220) mg/Kg.Bw of Nikathamide dosed
orally by gastric gavage, each single dose was given to two mice. The effective
dose (ED50) was determined by Up and Down method (13).Five mice were
dosed with 200mg/kg.BW of Nikethamide orally to induce convulsion and then treated
simultaneously with methanolic extract of saffron stigma orally with initial dose
500mg/kg.BW for the first mouse, depending on the outcome, the dose of was increased
by 20% when the signs of convulsion didn't show any relief but the dose was decreased
also by 20% in case of recovery. The ED50 was calculated according to
the following equation.
ED50= XF + Kd
XF= the last dose
d= difference between doses. It is 100 mg in this experiment.
K= constant. It is - 0.305
2.8. Investigation of some toxic effects of methanolic
extract of saffron stigma in mice:
Fifteen mice were divided equally into three groups. The
first group (T1), the second group (T2) were dosed respectively,
with 4000 and 5000mg mg/Kg.BW of methanolic extract of saffron stigma(the two highly
nontoxic doses obtained from range finding study) orally by gastric gavage for five
weeks, the third group was considered control group (C) dosed with distilled water
for the same period. Body weights were recorded before treatment and weekly after
treatment till the end of the exposure. At the end of exposure, some hematological
parameters were measured which included red blood cells count (RBC) according to
(14), white blood cells count (WBC) according to the method mentioned by Coles (15),
Hemoglobin by converting to methemoglobin using drabkin’s reagent. Values were determined
by the method described by Varely and Bell (16), also some biochemical parameters
included Serum uric acid was assessed using a commercially available kit (Bio System-SPAIN).
The principle of this method depends on the coupled quinoneimine reaction results
that are measured by spectrophotometry (17). Blood urea nitrogen (BUN) was assessed
using a commercially available kit (RANDOX-UK). The modified urease—Berthelot method
was used, and is based on the formation of a green complex of 2-2 dicarboxylindophenol
(18). Serum creatinine was assessed using a commercially available kit (BioSystem-SPAIN).
The method depends on the reaction of serum creatinine with picrate in an alkaline
medium, forming a colored complex, which is then measured by spectrophotometry (19).
Liver enzymes included alkaline phosphatase by a commercial kit of LiNEAR
Chemicals-SPAIN which is depended on the formation of free 4-nitrophenol and
inorganic phosphate, the test has been formulated according to (20),
alanineamino transeferase (ALT) by commercial kit of BioSystems-SPAIN by means
of the Lactatedehydrogenase coupled reaction(21), aspartateamino traseferase
also by commercial kit of BioSystems-SPAIN but the test depends on coupled reaction
of Malatedehydrogenase (22) which were measured. Blood was collected from the animals
in both two groups by heart puncture after anesthesia with 0.1 ml/100 gm BW of ketamine
10% post treatment. The serum was obtained from blood samples by centrifugation
at 5000 r/per minute for five minutes.
2.9. Histopathology:
The animals in both groups were sacrificed at the end of
the dosing period. Liver and kidney were obtained and preserved in 10% formalin,
then sent to the Medicinal City Hospital laboratory for histopathology processing
as previously described (23).
2.10. Stastical analysis:
All data expressed as mean ±SE. Analysis was performed using SPSS version 24.00. A one-way
ANOVA, A P-value < 0.05 was considered statistically significant. LSD multiple
range tests were performed for comparisons between the means.
3. RESULTS AND DISCUSSION:
3.1. Acute toxicity:
The result of range finding study which conducted to evaluate
acute toxicity of methanolic extract of saffron stigma in male Balb-C mice revealed
there are no signs of toxicity and death in all mice of the four groups that were
administered methanolic extract of saffron orally throughout 7 days of the experiment
shown in Table (1). Further three additional mice that were dosed with 5000mg/Kg.BW
also didn’t show any signs of toxicity and death.
Table1. The outcomes of the range finding study of
methanolic extract of saffron stigma in mice orally.
|
Dose mg/Kg.BW
|
Number of mice
|
Number of dead mice After 7 days of
treatment with methanolic extract
|
Number of alive mice After 7 days of
treatment with methanolc extract
|
|
2000
|
5
|
0
|
5
|
|
3000
|
5
|
0
|
5
|
|
4000
|
5
|
0
|
5
|
|
5000
|
5
|
0
|
5
|
The range finding study is more favorable instead of LD50
estimation (24) for expectation toxicity of saffron, which is considered as aspic
and is being added to cooked rice and beverage in many countries all over the world.
The EPA (1994) recommended that 5 gm/Kg.BW of any substance that doesn’t cause death
to rollout any acute toxic effect and no need for LD50 estimation. So
methanolic extract of saffron stigma is considered practically not to have any acute
toxic effect. Also the safety of saffron is assured by (25) who found saffon is
generally nontoxic when ingested in culinary amount but the lethal dose is 20 gm.
We thought that the purpose of this study is to aid in the selection of the proper
starting dose for other studies. Also Hosseinzadeh and Younesi (4) Proofed the safety
of both watery and ethanolic extract of saffron, when they found the maximum nonfatal
doses of both extract were 3.6 and 8 gm/Kg.BW respectively.
Table2. The results of pilot study of Nikathamide dosed
orally to miceforroducing convulsion.
3.2. Effective dose of methanolic extract of saffron stigma
as anticonvulsion:
The result of pilot study that was conducted in mice with
suitable dose of Nikethamide that produced convulsion without death, is summarized
in the table 2 which revealed that 0.2 ml/Kg.BW of Nikethamide orally is the best
one.
The estimation of ED50 of methanolic extract
of saffron stigma which was conducted in mice through relieving the convulsive effect
of Nikethamide. The result of Up and Down method that used for this purpose is summarized
in the Table 3, which revealed the ED50 is 494 mg/Kg.BW orally in male
Balb-C mice according to the below equation.
Table3. The result of Up and Dow method for estimation
ED50.
|
Initial
dose
Mg/Kg.BW
|
Last
dose
Mg/Kg.BW
animals
|
Result
after plant extract dosing
|
ED50
mg/Kg.BW.
|
|
500
|
500
|
XOOXO
|
494
|
X= no relieve of convulsion. O= relieve of convulsion.
ED50 = Xf+ KdXf= last doseK= constant (-
0.305).
d= difference between doses = 100 mg.
The anticonvulsion and antinociceptive effects of saffron
also was ensured by (4&6) who found that both watery and ethanolic extract of
saffron exhibited anticonvulsive effect in electroshock seizure and pentylentetrazoline
induced convulsion in mice. We thought the anticonvulsive effect of the ethanolic
extract of saffron stigma is due to the safranal, the colouring agent of saffron,
which was found relieving both minimal clonic seizures and generalized tonic‐clonic
seizures following pentylenetetrazolen administration in rats (26). Also the anti
convulsent effect of saffron attributed to crocin, crocetin, picrocrocin and safranal.
These components especially crocin, have significant effects including antidepressant
and anticonvulsant, analgesic (27) The effect of safranal is mediated, through the
GABA (A)‐benzodiazepine receptor complex (28).
3.3. Clinical study of sub chronic exposure of methanolic
extract of saffron stigma:
3.3.1. Some hematological test (Total white blood cells
count WBC, Red blood cells count RBC and hemoglobin):
Mthanolic extract of saffron stigma with both doses 4000
and 5000mg/Kg.BW showed significant increase P<0.05
of total white blood cells count, significant decrease P<0.05 in red blood cells count and blood hemoglobin in comparison
with control group. There were no significant differences in WBC count and RBC count
between both treated groups with saffron, but there was significant decrease in
blood hemoglobin of mice that were treated with 5000mg/Kg.BW in comparison with
mice dosed with 4000mg/Kg.BW, table (4). Daryoush and colleagues (29) found the
intraperitoneally doses of 0.35, 0.70 and 1.05 g kg−1 of stigma alcoholic
extract for two weeks to male wistar rats caused significantly decrease in RBC count
and hemoglobin blood level and increase in WBC count in a dose dependent manner,
which are nearly coordinated with our present study findings.
The elevation of WBC count may be due to inflammatory effect
of saffron mthanolic extract revealed in histopathological changes in kidney of
both dosed groups, represented by dilation and congestion of interstitial blood
vessels with perivascular lymphatic cuffing, figure (1), and infiltration of inflammatory
cells within the lumina and interstitial tissue. Furthermore, there is severe mononuclear
interstitial infiltration with slight fibrosis and the formation of eosinophilichyaline,
figure (2) and figure (3).
The anemic effect of the extract is may be due to saffron
alkaloid colchicines (30) which has antimitotic i.e inhibitory role in cell division
of rapidly dividing cells (31). Since stem cells of bone marrow are rapidly dividing
cells and included precursor of red blood cells (erythroblast), we supposed that
methanolic extract of saffron stigma with both 4000 and 5000 mg/Kg. BW exerted its
anemic effect may be through this mechanism.
It is axiomatic that creatinine blood level inversely correlated
with kidney function, so when the kidney function declines, serum creatininerises.
Also the increase of blood urea nitrogen may be seen with decreased kidney function
and some other status like dehydration and high dietary protein intake. We could
conclude the last two reasons because there were no signs of diarrhea and dehydration
in mice of the treated and control groups, and also they had been fed with the same
food.
Figure 1: Histopathological section in kidney of mouse
dosed 4000 mg/Kg.BW methanolic extract of saffron stigma for one month. Severe
dilation and congestion of intersticial vessels (
)
with privascular lymphocytic cuffing (
)
(H & E 10X)
Figure 2: Histopathological section in kidney of mouse
dosed 5000 mg/kg. Bw methanolic extract of saffron stigma for one month. Severe
interstitial mononuclear cells infiltration ()
with atrophy of glomerular tuft ()
and slight interstial fibrosis (
)
(H&E 10 X)
The histopathological changes which included degenerative
changes of epithelial linings of the proximal and distal convoluted tubules with
atrophy of glomerular tufts, in addition to severe dilation and congestion of interstitial
blood vessels with perivascular lymphatic cuffing, figure(1, 2, 3, 4) which confirmed
that saffron high doses given for one month had led to renal toxicity.
Figure 3: Histopathological section in kidney of mouse
dosed 5000 mg/Kg. BW methanolic extgract of saffron stigma for one month.
Formation of eosinophilic hyaline case () (H & E 40X)
Figure 4 histopathological section in kidney of mouse
dosed 4000mg/Kg. BW methanolic extract of saffron stigma for one month. Atrophy
of glomerular tuft (
)
(H&E40X)
3.3.3. Liver injury enzyme aspartate aminotransferase (AST)
and alanineaminotrasferase (ALT)
There was a significant increase P<0.05 in the serum
activity of AST of mice of both treated groups with two doses 4000 and 5000 mg/Kg.BW
of methanolic extract of saffron stigma in comparison with the mice of the controlled
one, but there were no significant changes between them. The serum activity of ALT
of both dosed groups showed no significant changes in comparison with control group,
but we can observe there is no significant increase in it, table(6).The changes in the serum activity of these enzymes
may be considered due to harmful effect of methanolic extract of saffron stigma
in the liver tissue of both dosed groups, which included dilation of central veins with vacuolar degeneration
of hepatocytes accompanied with proliferation of Kupffers cells (figure 5), fibrous
thickening of Glasson's capsule with formation of multiple parenchematous granuloma
(figure 6) and granuloma (figure 7)
Table 6, Liver injury enzymes (AST)U/L and (ALT) U/L
|
Group
|
AST U/L
|
ALT U/L
|
|
T1 4000mg/Kg.BW
T2 5000 mg/Kg.BW
Control (C) DW
|
170.70±7.44
A
180.20±6.65
A
110.80±4.25
B
|
130.99±30.22
A
144.60±40.80
A
77.10±29.30
A
|
LSD of AST=18.30 LSD of ALT=70.60
The different capital letters denote significance differences
P<0.05 between groups.
Figure 5: Histopathological section in liver of mouse
dosed 4000mg/Kg. BW methanolic extract of saffron stigma for one month.
Dilation of central vein () with vacuolar degeneration of hepatocytes () (H
& E 40X)
Figure 6: Histopathological section in liver of mouse
dosed 5000mg/Kg. BW methanolic extract of saffron stigma for one month.
Vacuolar degeneration of hepatocytes () with moderate fibrous thickening of
Glasson’s capsule () (H & E 40X)
3.4. The effect of methanolic extract of saffron stigma
in body Weight
The mice treated with 5000mg/Kg.BW of methanolic extract
of saffron stigma showed, significant decrease P<0.05
in body weight after 3, 4 and 5 weeks of treatment in comparison with its pretreatment
(day zero) body weight and control mice body weight, while the mice that were treated
with 4000 mg/Kg.BW of methanolic extract of saffron stigma showed significant decrease
P<0.05 in body weight only after five weeks of treatment in comparison
with its pretreatment(day zero) body weight and control group mice body weight,
table (7).
Saffron (Crocus Sativus L.) is considered
a source of polyphenols and carotenoids which have antioxidant ability (32). Natural
extracts from plants such as polyphenols and carotenoids reported to enhance body
weight loss (33,34).Carotenoids and polyphenols are the antioxidants which have
the ability to reduce blood glucose, triglyceride, blood LDL cholesterol, increase
energy expenditure and fat oxidation consequently reducing body weight (35,36),
and they are capable of inhibition of pancreatic lipase, Lipoprotein lipase and
glycerophosphate dehydrogenase (37,38).Capsule 176.5 mg/day of ethanolic saffron
extract given for two months reported a decrease in snacking and body weight in
overweight women(32 ). Crocin, a saffron carotene is a reversible lipase inhibitor
in comparison with a commercially available anti –obesity Orlista which is irreversible
(39). Ethanolic extract of saffron stigma was found to significantly reducing body
weight in rats through decreasing appetite as a side effect(40, 41). We thought
the possible reduction in body weight of both dose 4000 and 5000mg/Kg.BW of methanolic
extract of saffron stigma for Blab-C mice in our present study is dose dependent,
and may have contributed to decreasing fat metabolism through inhibition of pancreatic
lipase which resulted in decreasing calorie intake and also satiety effect which
suppressed appetite that noticed in mice of both treated groups.
4. REFERENCES:
1. Talha J, Priyanka M, Akanksha A.
Hypertension and herbal plants. Int Res J Pharm.; 2011; 2:26–30.
2. Alonso, G.L., Zalacain, A. and Carmona, M.
2012.Saffron. In Handbook of Herbs and Spices, 2nd Ed., (K.V. Peter, ed.)Woodhead
Publishing, Cambridge.
3. Serrano-Diaz, J., Sanchez, A.M., Martinez-Tome,
M., Winterhalter, P. and Alonso, G.L. 2013. A contribution to nutritional studies on Crocus sativus flowers and their value as food. J. Food Comp. Anal. 31, 101–108.
4. Hosseinzadeh H, Khosravan V.(2002).
Anticonvulsant effects of aqueous and ethanolic extracts of Crocus sativus L.
stigmas in mice.; Arch Irn Med; 5:44–7.
5. Hosseinzadeh H, Khosravan V. Anticonvulsant
effects of aqueous and ethanolic
extracts of Crocus sativus L. stigmas in mice. Arc Iran Med. 2002; 5(1):44.
6. Hosseinzadeh, H., Younesi HM.(2002)
Antinociceptive and anti-inflammatory effects of Crocus Sativus L stigma and
petal extract in mice. BMC Pharmacol.,
2; 2:7.
7. Hosseinzadeh H, Shamsaie F, Mehri
S.(2009).Antioxidant activity of aqueous and ethanolic extracts of Crocus
sativus L. stigma and its bioactive constituents, crocin and safranal.
Pharmacogn Mag., 5(20):419.
8. Toktam, Z., Bibi, M, R. and; Hossein, H.
(2014). Saffron Reduced Toxic Effects of its Constituent, Safranal, in Acute
and Subacute Toxicities in Rats. Jundishapur J Nat Pha; 9(1): 3-8
9. Posadzki P, Watson LK, Ernst E.(2013).
Adverse effects of herbal medicines. an overview of systematic reviews. Clin Med.; 13(1):7–12.
10. Karimi G, Tabibi N, Hosseinzadeh H, Shirzad
F(2004).Sub-acute toxicity of Saffron (Crocus Sativus L) stigma and petal in
rat. J Med Plants.; 12 :32–9.
11. Gao, W., Zhong, Y., Guo,
J., Yao. X., Wang, L., Jin, Y. and Qi, N.(2008).Studies on the Main Pharmacological Effects of Tongren' anshen Pill in vivo. Journal Of Capital
Medical University, 29(1): 74-77.
12. Frank. C,
LU. And Sam, K.(2009).Lu, S Basic
Toxicology Fundamental, Target organs and Risk assessment. Informa Health care
USA, Inc, Pp:92.
13. Dixon, W. J. (1980). Efficient Analysis of
Experimental Observation. Am. Rev. Pharmacol. Toxicol., 20:441-462.
14. Dacie, J. V. and Lewis, S. M. (1984).
Practical hematology. Churchill Livingston Ed, Selecto Printing Co. Ltd., New
York, pp.445.
15. Coles, E. H. (1974). Hemostasis and
coagulation of blood. In: Veterinary Clinical Phathology. 2nd
edition. Chapter 5. W.B. Saunders Company. Printed in Philadelphia,
London, Toronto. PP.155, 161.
16. Varely, H.A and Bell, M. (1980).Practical
Clinical Biochemistry, 5thEd.London.WilliamHeine.Medical Book
Ltd.PP:107-122.
17. Fossati, P., Prencipe, L. and Berti, G.
(1980) Use of 3, 5-Dichloro-2-Hydroxybenzenesulfonic Acid/14-Aminophenazone
Chromogenic System in Direct Enzymic Assay of Uric Acid
in Serum and Urine. Clinical Chemistry, 26, 227-231.
18. Fawcett, J.K. and Scott, J.E. (1960) A Rapid
and Precise Method for the Determination of Urea. Journal of Clinical
Pathology, 13, 156-159.
19. Bartels, H. and Bohmer, M. (1971) Eine Mikromethodzur Kreatinine Test Immung. Clinical Chimica Acta,
32, 81-85.
20. German Society for Clinical chemistry.(1972).Recommendations
of the enzyme commission.Z.Klin.Chem.Klin.Biochem.10:281.
21. Approved Recommendation.(1985). IFCC
Methods for the Measurement of Catalytic concentration of Enzymes Part 3: IFCC
Method for Alanineameno trasferase (EC 2.6.1.2).J. Clin Chem Clin Biochem 1986; 24:481-495.
22. Gilla, F, J., Olivella, T., Cruz Pastor,
M., Arenas, J., Morino, R., Dorban, R. and Gomez, JA.(1985).A simple procedure
for routine determination of aspartate aminotrasfderase
and alanineamino traseferase with Pyridoxalphosphate. Clin Chem Acta;
153:241-247.
23. Luna, L.G. (1968) Manual of Histology
Staining Methods of the Armed Forces Institute of Pathology. 3rd
Edition, McGraw Hill Book Co., New York.
24. EPA.(1994). Health
Effect Test Guidelines, Hazard evaluation, Code of Fedral Regulations Title 40,
Part 792, 798.
25. Schmidt, M., Betti, G. and Hensel, A. (2007).Saffron in phytotherapy: Pharmacology and clinical
uses. Wien Med Wochenschr.; 157:315–9. [PubMed]
26. VIJAYA, B, K.(2011).Medicinal Uses and Pharmacological properties of
Crocus Sativus Linn(Saffron). International Journal of Pharmacy and Pharmaceutical
Sciences., 3(3):0975-1491.
27. Khorasany, A., Rand Hosseinzadeh,
H.(2016).Therapeutic effects of saffron (Crocus sativus L.) in digestive
disorders: a review. Iran J Basic Med Sci.; 19(5):455-69.
28. Hosseinzadeh, H. and Ghenaati, J.(2006).
Evaluation of the antitussive effect of stigma and petals of saffron (Crocus
sativus) and its components, safranal and crocin in guinea pigs. Fitoterapia
., 77: 446–448.
29. Daryoush, M., Ghafour, M., Mehran, M.,
Yousef, D. and Mir Hadi, K.(2007). Subacute Toxicity of Crocus Sativus L.
(Saffron) Stigma Ethanolic Extract in Rats. American Journal of Pharmacology
and Toxicology 2 (4):189-193.
30. Colchicine for acute gout update
information about dosing and drug interactions.(2010).National Prescribing
Service, 14 may.
31. Folpini, A. and Furfori. (1995).Colchicine
toxicity. Clinical Toxicology., 33(1):71-77.
32. Maryam, M., Azrina, A., Huzwah, K.,
Barakatun, N., Yousif, M. and Sabriah,
M.(2013).Saffron: A Natural Potent Antioxidant as promising Antioxidants drug. Antioxidants., 2:293-308.
33. Moro, C., Basile, G.(2000). Obesity and medicinal
plants. Fitoterapia, 71, S73–S82.
34. Rayalam, S., Della-Fera, M.A.,
Baile, C.A.(2008). Phytochemicals and regulation of the adipocyte lifecycle. J.
Nutr. Biochem, 19, 717–726.
35. García-Lafuente, A., Guillamón,
E.; Villares, A., Rostagno, M.A.; Martínez, J.A.(2009). Flavonoids as anti-inflammatory agents: Implications in cancer and cardiovascular
disease. Inflamm. Res, 58, 537–552.
36. Terra, X.; Montagut, G.; Bustos, M.;
Llopiz, N.; Ardèvol, A.; Bladé, C.; Fernández-Larrea, J.; Pujadas, G.; Salvadó,
J.; Arola, L.(2009). Grape-seed procyanidins prevent low-grade inflammation by modulating cytokine expression in rats fed a high-fat diet. J. Nutr.
Biochem, 20, 210–218.
37. Slanc, P.; Doljak, B.; Kreft, S.; Lunder, M.;
Janeš, D.; Štrukelj, B. Screening of selected food and medicinal
plant extracts for pancreatic lipase inhibition. Phytother. Res. 2009,
23, 874–877.
38. Birari, R.B,; Bhutani, K.K. Pancreatic
lipase inhibitors from natural sources: Unexplored potential.
Drug Discov. Today 2007, 12, 879–889.
39. Sheng, L.; Qian, Z.; Zheng, S.; Xi, L.
Mechanism of hypolipidemic effect of crocin in rats: Crocin inhibits pancreatic
lipase. Eur. J. Pharmacol. 2006, 543, 116–122.
40. Akhondzadeh, B.A.; Ghoreishi, S.A.;
Noorbala, A.A.; Akhondzadeh, S.H.; Rezazadeh, S.H. Petaland stigma of Crocus
sativusL. in the treatment of depression: A pilot double-blind randomized
trial. J. Med. Plants 2008, 7, 29–36.
41. Xi, L.; Qian, Z.; Xu, G.; Zheng, S.; Sun,
S.; Wen, N.; Sheng, L.; Shi, Y.; Zhang, Y. Beneficial impact of crocetin, a
carotenoid from saffron, on insulin sensitivity in fructose-fed rats.J.
Nutr. Biochem. 2007, 18, 64–72.