Content uploaded by Nora Abdullah ALFaris
Author content
All content in this area was uploaded by Nora Abdullah ALFaris on Jan 10, 2018
Content may be subject to copyright.
OPEN ACCESS Asian Journal of Clinical Nutrition
ISSN 1992-1470
DOI: 10.3923/ajcn.2017.64.70
Research Article
Effects of Chlorophyll on Body Functioning and Blood Glucose
Levels
Amnah Mohammed Alsuhaibani, Nora Mohammed ALkehayez, Amal Hassan Alshawi and
Nora Abdullah Al-Faris
Department of Nutrition and Food Science , College of Home Economics, Princess Nourah Bint Abdulrahman University, Riyadh,
Saudi Arabia
Abstract
Background and Objective: Blood glucose levels can be affected by the various types of chlorophyll. Experiments have proven that
chlorophyll has antioxidant activities that are present in various foods. This study aimed to identify the effects of chlorophyll type on blood
glucose and body functioning. Methodology: For the study, five different high-chlorophyll products were selected: Mint, broccoli, thyme,
grapes and bell peppers. The chlorophyll was extracted from each source. Two types of chlorophyll, chlorophyll a and b, were extracted.
The total chlorophyll content was determined. Toxicity tests were conducted using 40 Swiss albino male rats, 6-7 weeks old. The rats were
randomly split into 4 groups. The control group was fed
ad libitum
with a Purina® chow diet. The chlorophyll extracts were ground and
mixed with the standard pellets so that the feed contained 15% carbohydrate weight replacement with chlorophyll. Students t-test and
the chi-square test were used to assess the significance of the values obtained in both the treated and the control groups during the study.
Results: Body weights increased after feeding with chlorophyll from all sources except bell peppers. The weight before feeding was
334.10±26.5 g, after feeding, it was 318.7±26.96 g, which is interpreted as a low difference. The mean glucose level was monitored
0, 1, 2 and 3 h after the intake of chlorophyll. A diet rich in chlorophyll led to a slight decrease in the number of white blood cells,
haematocrit, haemoglobin and an increase in red blood cells compared with control. The results of the treatment did not show any
significant changes in the levels of total cholesterol, LDL , HDL , tr igl yce rid es, g luc ose , ur ea a nd cr eat ini ne am on g th e ex per im ent al
and control groups. Moreover, there was significant difference (p<0.05) in the weights of the animals organs among the groups.
Conclusion: It is concluded that chlorophyll extracts from mint, broccoli, thyme and bell pepper are likely to have important implications
regarding blood sugar. Bell pepper extracts and juice has benefits in body weight and further studies are warranted.
Key words: Body functioning, chlorophyll, extraction, juices, body weight
Received: January 05, 2017 Accepted: March 02, 2017 Published: March 15, 2017
Citation: Amnah Mohammed Alsuhaibani, Nora Mohammed Alkehayez, Amal Hassan Alshawi and Nora Abdullah Al-Faris, 2017. Effects of chlorophyll on
body functioning and blood glucose levels. Asian J. Clin. Nutr., 9: 64-70.
Corresponding Author: Nora Abdullah Al-Faris, Department of Nutrition and Food Science, College of Home Economics,
Princess Nourah Bint Abdulrahman University, Riyadh, Saudi Arabia Tel: +966118237437
Copyright: © 2017 Amnah Mohammed Alsuhaibani
et al
. This is an open access article distributed under the terms of the creative commons attribution
License, which permits unrestricted use, distribution and reproduction in any medium, provided the original author and source are credited.
Competing Interest: The authors have declared that no competing interest exists.
Data Availability: All relevant data are within the paper and its supporting information files.
Asian J. Clin. Nutr., 9 (2): 64-70, 2017
INTRODUCTION
Glucose is an instant source of energy but when
consumed in high concentration, it leads to major problems.
The blood glucose level is the amount of sugar or glucose
present in the blood stream. The body naturally regulates
glucose levels as a part the bodys metabolic processes. It is a
primary energy mechanism for cells and the blood lipid profile.
Bloodglucose is transported from the intestines/liver to the
cells via the bloodstream. The absorption of glucose is
promoted in the presence of insulin, a hormone produced in
the pancreas. If blood glucose levels are not balanced, both
high and low blood glucose leads to the malfunctioning of
body organs1. Fruit and vegetables contain phytochemicals
that may be prevent diabetes by regulating glucosidase and
lipase activities, reducing the postprandial glycaemic level,
providing anti-inflammatory activity and improving pancreatic
function and synergistic action with hypoglycaemic drugs2.
Fruit and vegetables have been reported to contain
antioxidant compounds that are valuable in human health,
such as carotenoids, vitamin C, vitamin E and phenolic and
thiol (SH) compounds2-4. The consumption of fruit and
vegetables is part of the Arabic cultural heritage and they play
important roles in the customs, traditions and food culture of
the Saudi household. Mint (
Menthapulegium
), broccoli
(
Brassica oleracea
), thyme (
Thymus vulgaris
) and
Gongronema latfolium
are green leafy vegetables that
contain a fair amount of chlorophyll. The consumption of a
diet rich in fruits and vegetables that are rich in chlorophyll is
associated with lower incidences of oxidation-linked diseases,
such as diabetes and cardiovascular disease2-4.
Increased consumption of phytochemicals that have
antioxidant activity can suppress the development of
diabetes5. In contrast to other phytochemical compounds,
such as phenolic components, chlorophyll is present in large
quantities in plants (±1% dry weight); therefore, it has the
potential to be a functional food6. Chlorophyll is a component
that may help to prevent disease. Moreover, the chlorophylls
are the most abundant natural pigment and is considered a
plant-derived antioxidant7. They are called dihydroporphyrins
because they contain four pyrrole rings coordinated with
magnesium metal. Along with the porphyrin ring, they also
contain a long hydrophobic sidechain derived from an alcohol
called phytol (C20H39OH), which is responsible for the
hydrophobicity of the whole molecule. Generally, higher-level
plants contain two types of chlorophyll: a and b. These
chlorophyll types differ in terms of the R group, which may be
an aldehyde group in chlorophyll b or a methyl group in
chlorophyll a7.
The role of chlorophyll as a hypoglycemic agent occurs
through the inhibition of free radicals. Chlorophyll is an
antioxidant chain breaker that donates its electrons to free
radicals and forms complexes with peroxyl radicals to
generate a stable product8. Ferruzzi
et al
.9 reported that
dietary chlorophyll derivatives prevalent in both fresh and
processed foods and dietary supplements have antioxidant
and antimutagenic activities. There are many imported foods
based on chlorophyll and the highest chlorophyll content
occurs in the green fruits and vegetables mint, broccoli,
thyme, grapes and bell peppers. The aim of this study was to
evaluate the chlorophyll content of mint, broccoli, thyme,
grapes and bell peppers which are commonly consumed in
Saudi.
MATERIALS AND METHODS
Preparation of products
Extraction of chlorophyll: First, five products (mint, broccoli,
thyme, grapes and bell peppers) with a high phenol content
were collected from local vendors. They were referred to as
FP 1 (mint), FP 2 (broccoli), FP 3 (thyme), FP4 (grapes) and
FP 5 (bell peppers), respectively. Then, whole chlorophyll was
extracted and the chlorophyll a and b component contents
were determined according to AOAC10 methods. Table 1
shows the total chlorophyll a and b components per 100 g.
Preparation of samples: Fresh grapes juice was prepared by
homogenizing the fruit right before the start of the
experiments (1 mL of juice contained approximately 2 g of
fresh grapes). The mint, broccoli, thyme and bell pepper leaves
were washed, weighed (100 g LG1) and triturated with water in
a blender for 7 min. The juice was filtered and frozen in a flask
placed in a refrigerator. Each flask was thawed daily at
ambient temperature two hours prior to administration.
Animal testing methods: The study was approved by the
Department of Nutrition and Food Science, Princess Nourah
Bint Abdulrahman University, Riyadh, Saudiin September
2011. Using chronic modes, toxicity studies were conducted
with 40 male Swiss albino rats, aged 6-7 weeks and with
different weights. The animals were bred and raised at
the Animal House, Central Laboratory for Drug and Food
Analysis, Ministry of Health, Riyadh, Saudi Arabia. The
animals were handled in accordance with the guidelines for
selecting doses for long-term treatment and minimal
toxicity11. The rats were randomly split into 4 groups (10 rats
per group). The environment was maintained under standard
65
Asian J. Clin. Nutr., 9 (2): 64-70, 2017
conditions with a humidity of 50%, a temperature of 22±2EC
and light conditions of 12 h each for dark and light.
All the animals were given free access to water. The
control group was fed
ad libitum
with a Purina® chow diet
purchased from the Arabian Agricultural Services Company
(Arasco), Riyadh, Saudi Arabia. The feeds given to the
experimental groups were prepared according to the
American Institute of Nutrition (AIN) guidelines12. Dose
selection was based on the regulatory guidelines requiring
minimal toxicity during long-term treatment. As body weight
and water intake increased over time, the daily dose was
adjusted to approximate the initial human therapeutic dose
specified by the manufacturer. The extracts were ground and
mixed with standard pellets so that 15% of the feeds
carbohydrate weight was replaced with the experimental
products. The feed was stored at 5EC until it was used.
Table 2 summarizes the control and experimental diets. The
juices were administered orally at a dose of 0.29 g kgG1 once
a day. The dose administered to the animals was based on
100 g LG1, which corresponds to a daily intake of 200 mL of
juice by an adult man weighing 70.0 kg (this intake was based
o n p op u la t io n c o ns u lt at i on ) . T h e d i et s we r e l a be l le d FP 1 mi n t,
FP 2 broccoli, FP 3 thyme, FP 4 grapes and FP 5 bell peppers.
Parameters assessed
Toxicity: It was measured as suggested by the WHO Scientific
Group13 and Wilson
et al
.11.
Weight change: Body weights were recorded before and after
the experimental period to examine the impact of the
different drugs on body weight. Furthermore, at the end of the
treatment, the weights of the vital organs (heart, lungs,
kidneys, spleen, liver and testes) were weighed and reported
per 100 g b. wt., to facilitate comparisons between the normal
and control animals.
Haematological and biochemical analysis: These were
assessed before and after the experimental period. The
haematological profiles included White Blood Cell count
(WBC), Red Blood Cell count (RBC), haemoglobin (Hb),
haematocrit (HCT) and the Mean Corpuscular Volume (MCV).
Blood was also collected and the serum was separated and
stored at -20EC for later biochemical analysis of liver and
kidney function and lipid profile. Specifically, alanine
aminotransferase (ALT), aspartate aminotransferase (AST)14,
glucose15, urea16, creatinine17, cholesterol; triglycerides18,
High Density Lipoprotein (HDL) cholesterol19 and Low
Density Lipoprotein (LDL) cholesterol20 were measured. The
parameters were analysed with enzymatic colorimetry using
test combination reagents (Boehringer Mannheim GmbH,
Diagnostica, Germany). Measurements were made using a
spectrophotometer (Introspect II, LKB).
Oral Glucose Tolerance Test (OGTT): Oral glucose tolerance
was measured after fasting rats were given a glucose solution
Table 1: Total chlorophyll a and b components per 100 g
Sources Test-ref Chlorophyll a (ppm) Chlorophyll b (ppm) Total
FP 1 AOAC 972.04 549.00 201.78 750.78
FB 2 AOAC 972.04 75.47 32.00 107.47
FP 3 AOAC 972.04 393.10 585.88 978.98
FP 4 AOAC 9772.04 3.79 1.45 5.24
FP 5 AOAC 972.04 31.66 9.69 41.35
FP 1: Mint, FP 2: Broccoli, FP 3: Thyme, FP 4: Grapes, FP 5: Bell pepper
Table 2: Constituents of control and experimental diets in g/100 g
Quantity of nutrients per 100 g
----------------------------------------------------------------------------------------------------------------------------------------------------------------------
Chemical components Control FP 1 FP 2 FP 3 FP 4 FP 5
Corn-starch 46.57 39.58 39.58 39.58 39.58 39.58
Casein 14.00 14.00 14.00 14.00 14.00 14.00
Dextrinised corn-starch 15.50 15.50 15.50 15.50 15.50 15.50
Sucrose 10.00 10.00 10.00 10.00 10.00 10.00
Soybean oil 4.00 4.00 4.00 4.00 4.00 4.00
Fiber 5.00 5.00 5.00 5.00 5.00 5.00
Mineral mix 3.50 3.50 3.50 3.50 3.50 3.50
Vitamin mix 1.00 1.00 1.00 1.00 1.00 1.00
L-cystine 0.18 0.18 0.18 0.18 0.18 0.18
Choline bitartrate 0.25 0.25 0.25 0.25 0.25 0.25
Extract (15% carbohydrate) - 6.99 6.99 6.99 6.99 6.99
Total 100.00 100.00 100.00 100.00 100.00 100.00
FP 1: Mint, FP 2: Broccoli, FP 3: Thyme, FP 4: Grapes, FP 5: Bell pepper
66
Asian J. Clin. Nutr., 9 (2): 64-70, 2017
(1.75 g kgG1 of weight of the rats). Blood glucose was
measured after 1, 2 and 3 h and the test was repeated once a
week. The average of the readings was taken.
Statistical analysis: Statistical analysis was performed using
SPSS v.16 (Statistical Package for Social Sciences, version 16,
SPSS, Inc., Chicago, USA)21. Students t-test and the chi-square
test were used to assess the significance of the values
obtained in both the treated and the control groups during
the study. p values of <0.05 were considered statistically
significant22.
RESULTS AND DISCUSSION
Effect of chlorophyll extraction and juices on body weight:
Table 3 shows that the rats weightsincreased gradually after
consuming chlorophyll in the form of mint, broccoli, thyme,
grapes but not as a result of bell pepper extract consumption.
The pre-treatment value was 334.10±26.50 g, whereas the
post-treatment value was 318.70±26.96 g. Both the difference
and the standard deviation were low. The consumption of
chlorophyll extracts led to growth and the increase in weight
reached as much as 1.27 g dayG1 8.
Additionally, changes in the body weights of male rats
were also observed with the administration of chlorophyll
in the form of juices made from mint, broccoli, thyme,
grapes and bell peppers. Again, the intake of bell pepper
juice was associated with a decrease inthe body weight
of the male rats. Dias23 demonstrated that bell peppers
contain substances that lead to increases in bodys heat
production and oxy gen c ons ump tion for ap pro ximat ely
20 min after eating. Under such circumstances, the body burns
extra calories, which helps with weight loss.
Effect of chlorophyll extracts and juices on the Oral Glucose
Tolerance Test (OGTT): The mean glucose level of the male
rats was monitored at 0, 1, 2 and 3 h after the intake of
chlorophyll in the form of mint, broccoli, thyme, grape and bell
pepper extracts and juices. The resultsindicated that treatment
with chlorophyll extract led to decreased blood glucose after
3 h (Fig. 1). The highest decrease occurred in the rats that were
fed thyme extract. Nissa
et al
.8 demonstrated that higher
doses of chlorophyll extract led to blood glucose levels. The
mechanism of chlorophyll as ahypoglycemic agent occurs
through its inhibition of free radicals. Chlorophyll is an
antioxidant chain breaker that donates its electrons to free
r ad ic a ls a nd fo r ms c om p le x es w it h pe r ox y ra d ic a ls t o g e ne r at e
Table 3: Effect of chlorophyll extracts and juices on weight
Body weight (g)
-------------------------------------------------------------
Treatment groups Pre-treatment** Post-treatment**
Control 339.24±19.39 391.89±24.41
Mint extract 339.24±19.39 394.80±10.52
Broccoli flower extract 336.90±21.50 395.80±27.44
Bell pepper extract 334.10±26.50 318.70±26.96*
Thyme extract 338.80±27.92 396.80±32.64
Grape extract 333.40±11.40 389.40±9.00
Control 2 339.24±19.39 391.89±24.41
Mint juice 338.06±19.90 395.06±10.79
Broccoli flower juice 336.18±22.06 396.09±28.15
Bell pepper juice 339.93±27.19 326.99±27.66*
Thyme juice 334.75±28.65 388.82±33.49
Grape juice 339.21±11.70 394.40±9.23
*Compare between the experimental group and control, *p<0.05 (student
t-test), **Mean±Standard Deviation
Fig. 1: Effect of chlorophyll extracts and juices on oral glucose tolerance test results
67
0
20
40
60
80
100
120
140
160
180
0123
Blood glucose (mg dLG1)
Time (h)
Control Mint extract Broccoli flower extract
Bell pepper extract Thyme extract Grape extract
Mint juice Broccoli flower juice Bell pepper juice
Thyme juice Grape juice
Asian J. Clin. Nutr., 9 (2): 64-70, 2017
a stable product7. In addition to chlorophyll, chloroplasts
also contain exogenous superoxide dismutase24 and ascorbic
acid25, which also have antioxidant capacity. Paradoxically,
grape juice led to an increase in blood glucose, while grape
extract had the opposite effect, possibly because of the high
sugar content of grapes compared with the other fruits and
vegetables used.
Effects of chlorophyll extracts and juices on haematological
parameters: Table 4 shows the effects of the chlorophyll
products on the haematological profile of experimental
animals. Itshows that the group fed with chlorophyll
experienced a slight decrease in the number of white
blood cells and an increase in red blood cells compared
with controls. The decreases in white blood cells may
have occurred b ec au se c hl oro ph yll pl ays ananti-bacterial
role26. The results indicated that the treatment did
not lead to any significant changes.
Data analysis indicated that there was no significant
difference between the treatment groups and the control
group in terms of lipid profile and blood glucose levels.
Figure 2 shows that there were no significant differences in
the levels of total cholesterol, LDL, HDL, triglyceride, glucose,
urea and creatinine among the experimental and control
groups. These results agree with the study of Aminian
et al
.27,
who showed that unripe gra pe juice consumption had no
measurable effect on serum HDL-C levels in healthy
individuals. They also stated that grape juice had no effect
on TG, total cholesterol and LDL-C27. Some studies found
that daily consumption of grapes or grape juice reduced
body weight, blood pressure and lipids compared with the
control28.
Table 4: Impact of chlorophyll extracts and juices on the haematological test results
Haematology tests
---------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Treatment groups WBC** (×101 LG1) RBC** (×1012 LG1) Hb** (g dLG1) MCV** (fl) HCT** (%)
Control 8.98±0.11 8.08±0.13* 16.78±0.23 51.32±0.92 46.58±0.66
Mint extract 8.67±0.10 8.74±0.16 15.04±0.27 53.07±0.95 42.66±0.76
Broccoli flower extract 8.92±0.37* 8.70±0.16 16.32±0.26 49.56±0.89 41.42±0.74
Bell pepper extract 8.51±0.37* 8.57±0.15 16.53±0.26 48.53±0.87 42.45±0.76
Thyme extract 8.23±0.20* 8.18±0.15 15.84±0.27 49.36±0.88 42.45±0.76
Grape extract 8.11±0.13* 8.13±0.15 16.32±0.26 51.52±0.92 40.39±0.72
Mint juice 8.78±0.10 8.91±0.16 16.34±0.27 54.12±0.97 43.51±0.78
Broccoli flower juice 8.33±0.38 8.87±0.16 16.61±0.26 50.55±0.91 42.25±0.76
Bell pepper juice 8.91±0.37 8.74±0.16 15.82±0.27 49.50±0.89 43.30±0.78
Thyme juice 8.46±0.21 8.34±0.15 15.13±0.27 50.34±0.90 43.30±0.78
Grape juice 8.25±0.13 8.29±0.15 16.61±0.26 52.55±0.94 41.20±0.74
WBC: White blood cell, RBC: Red blood cell, Hb: Haemoglobin, MCV: Mean corpuscular, HCT: Haematocrit, *Compare between the experimental group and control,
*p<0.05 (student t-test), **Mean±Standard Deviation
Fig. 2: Effect of chlorophyll extracts and juices on lipids profile and glucose level
68
0
20
40
60
80
100
120
140
160
Control Mint
extract Broccoli
flower
extract
Bell pepper
extract Thyme
extract Grape
extract Mint juice Broccoli
flower
juice
Bell pepper
juice Thyme
juice Grape juice
Blood level (mg dLG1)
Treatements
Glucose Urea Creatinine Cholesterol HDL cholesterol LDL cholesterol Triglycerides
Asian J. Clin. Nutr., 9 (2): 64-70, 2017
Table 5: Effect of chlorophyll extracts and juices on organ weights
Organ weight changes (g/100 g b.wt.)
------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Treatment groups Heart** Lungs** Liver** Spleen** Testes** Kidneys**
Control 1.7±0.03 1.59±0.04 10.99±0.27 1.02±0.03 1.88±0.05 1.29±0.03
Mint extract 1.33±0.03 1.79±0.04 10.25±0.28 1.13±0.03 1.88±0.05 1.32±0.03
Broccoli flower extract 1.28±0.03 1.73±0.04 10.96±0.32 1.06±0.03 1.83±0.05 1.31±0.03
Bell pepper extract 1.58±0.03 1.67±0.04 10.58±0.26 1.04±0.03 1.81±0.05 1.10±0.03
Thyme extract 1. 5±0.03 1.63±0.04 10.98±0.27 1.04±0.03 1.92±0.05 1.05±0.03
Grape extract 1.59±0.028 1.465±0.036 10.577±0.263 0.983±0.024 1.846±0.05 1.015±0.025
Mint juice 1.129±0.033 1.789±0.044 11.247±0.280 1.130±0.028 1.883±0.05 1.318±0.033
Broccoli flower juice 1.38±0.03 1.73±0.04 11.96±0.32 1.06±0.03 1.83±0.05 1.31±0.03
Bell pepper juice 1.18±0.03 1.67±0.04 10.58±0.26 1.04±0.03 1.81±0.05 1.10±0.03
Thyme juice 1.015±0.025 1.632±0.041 10.975±0.27 1.036±0.03 1.915±0.05 1.046±0.026
Grape juice 1.12±0.03 1.47±0.04 10.58±0.26 0.98±0.02 1.95±0.05 1.02±0.025
*Compare between the experimental group and control, *p<0.05 (student t-test), **Mean±Standard Deviation
There was also no significant difference in the weights of
the animals organs among the groups (Table 5).
CONCLUSION
Consumption of chlorophyll of mint, broccoli, thyme,
grapes led to increase of weight while chlorophyll of bell
pepper demonstrated decreased the weight. Based on the
findings in this study, a key observation was that chlorophyll
was the key factor related to increase in weight and decreased
blood glucose. It can be concluded that include chlorophyll
rich food and its extraction in diet positively influence on
health.
SIGNIFICANCE STATEMENT
This study discovers the potential of chlorophyll in the
form of mint, broccoli, thyme, grapes as a source of
phytochemicals that can be beneficial for humannutrition.
This study will help the researcher to uncover the critical areas
of food applications that can use chlorophyll chemical
constituents. Thus, new information about chlorophyll in the
form of macro- and micronutrients can be obtained and used
in a practical way.
REFERENCES
1. Ettinger, S., 2004. Macronutrients: Carbohydrates, Proteins
and Lipids. In: Krause's Food, Nutrition and Diet Therapy,
Mahan, L.K. and S. Escott-Stump (Eds.). 11th Edn., Saunders,
USA., ISBN: 9780721697840, pp: 37-73.
2. Zhang, Y.J., R.Y. Gan, S. Li, Y. Zhou, A.N. Li, D.P. Xu and H.B. Li,
2015. Antioxidant phytochemicals for the prevention and
treatment of chronic diseases. Molecules, 20: 21138-21156.
3. Arshiya, S., 2013. The antioxidant effect of certain fruits:
A review. J. Pharm. Sci. Res., 5: 265-268.
4. Parashar, S., H. Sharma and M. Garg, 2014. Antimicrobial and
antioxidant activities of fruits and vegetable peels: A review.
J. Pharmacogn. Phytochem., 3: 160-164.
5. Pandey, K.B. and S.I. Rizvi, 2009. Plant polyphenols as dietary
antioxidants in human health and disease. Oxidative Med.
Cell. Longevity, 2: 270-278.
6. Prangdimurti, E., D. Muchtadi, M. Astawan and F.R. Zakaria,
2006. Aktivitas antioksidan ekstrak daun suji (
Pleomele
angustifolia
N.E. Brown) [antioxidant activity of suji (
Pleomele
angustifolia
N.E. Brown) leaf extract]. J. Teknologi dan
Industri Pangan, 17: 79-86.
7. Inanc, A.L., 2011. Chlorophyll: Structural properties, health
benefits and its occurrence in virgin olive oils. Akademik G2da,
9: 26-32.
8. Nissa, C., M.I. Kartasurya and B. Rahmawati, 2016. Effects of
chlorophyll in papaya leaves on superoxide dismutation and
blood glucose level of diabetic rats. Makara J. Health Re s.,
19: 75-80.
9. Ferruzzi, M.G., V. Bohm, P.D. Courtney and S.J. Schwartz, 2002.
Antioxidant and antimutagenic activity of dietary chlorophyll
derivatives determined by radical scavenging and bacterial
reverse mutagenesis assays. J. Food Sci., 67: 2589-2595.
10. AOAC., 2012. Official Method of Analysis. 19th Edn.,
Association of Official Analytical Chemists, Washington DC.,
USA.
11. Wilson, D.T., J.F. Hardisty, J.R. Hayes and N.H. Wilson, 2014.
Short-Term, Subchronic and Chronic Toxicology Studies.
In: Princip les and M ethods o f Tox icology , Hay es, A.W a nd
C.L. Kruger (Eds.). 6th Edn., CRC Press, New York,
pp: 1205-1250.
12. Lien, E.L., F.G. Boyle, J.M. Wrenn, R.W. Perry, C.A. Thompson
and J.F. Borzelleca, 2001. Comparison of AIN-76A and
AIN-93G diets: A 13-week study in rats. Food Chem. Toxicol.,
39: 385-392.
13. Allmark, M.G., 2005. World Health Organization Technical
Report Series, No. 348: Procedures for Investigating
Intentional and Unintentional Food Additives. World Health
Organization, Geneva.
69
Asian J. Clin. Nutr., 9 (2): 64-70, 2017
14. Huang, X.J., Y.K. Choi, H.S. Im, O. Yarimaga, E. Yoon and
H.S. Kim, 2006. Aspartate aminotransferase (AST/GOT) and
alanine aminotransferase (ALT/GPT) detection techniques.
Sensors, 6: 756-782.
15. Rocco, R.M., 2006. Enzymaticglucose. In: Landmark Papers in
Clinical Chemistry, Rocco, R.M. (Ed.). 1st Edn., Elsevier,
Amsterdam, pp: 271-275.
16. Francis, P.S., S.W. Lewis and K.F. Lim, 2002. Analytical
methodology for the determination of urea: Current practice
and future trends. TrAC Trends Anal. Chem., 21: 389-400.
17. Delaney, M.P., C.P. Price and E.J. Lamb, 2007. Kidney Function
and Disease. In: Tietz Fundamentals of Clinical Chemistry
Passcode Only, Burtis, C.A., E.R. Ashwood and D.E. Bruns (Eds.).
6th Edn., W.B. Saunders Co., Philadelphia, pp: 631-655.
18. Kalia, V. and C.S. Pundir, 2004. Determination of serum
triglycerides using lipase, glycerol kinase, glycerol-3-
phosphate oxidase and peroxidase co-immobilized onto
alkylamine glass beads. Indian J. Biochem. Biophys,
41: 326-328.
19. Nauck, M., D. Wiebe and G.R. Warnick, 2000. Measurement of
High-Density-Lipoprotein Cholesterol. In: Handbook of
Lipoprotein Testing, Rifai, N., G.R. Warnick and
M.H. Dominiczak (Eds.). 2nd Edn., AACC Press, Washington,
DC., pp: 227-230.
20. Nauck, M., G.R. Warnick and N. Rifai, 2002. Methods for
measurement of LDL-cholesterol: A critical assessment of
direct measurement by homogeneous assays versus
calculation. Clin. Chem., 48: 236-254.
21. SPSS., 2007. SPSS for Windows, Version 16.0. SPSS Inc.,
Chicago, IL., USA.
22. Ross, S.M., 2010. Introductory Statistics. 3rd Edn.,
Academic Press, Japan, UK., USA., ISBN: 9780080922102,
Pages: 848.
23. Dias, J.S., 2012. Nutritional quality and health benefits of
vegetables: A review. Food Nutr. Sci., 3: 1354-1374.
24. Winarsi, H., 2007. Antioksidan Alami dan Radikal Bebas:
Potensi dan Aplikasinya Dalam Kesehatan. Penerbit Kanisius,
Yogyakarta.
25. Shao, H.B., L.Y. Chu, Z.H. Lu and C.M. Kang, 2008.
Primary antioxidant free radical scavenging and redox
signaling pathways in higher plant cells. Int. J. Biol.
Sci., 4: 8-14.
26. Kumar, N.S., M. Murali, A.M. Nair and A.S. Nair, 2016.
Green blood therapy of wheat grass-nature's finest
medicine-A literature review. IOSR J. Pharm. Biol.
Sci., 11: 57-64.
27. Aminian, B., S.M. Massoompour, A. Sadeghalvaad and
G.H. Omrani, 2003. Unripe grape juice (verjuice) as a
lipid-lowering agent: Fact or fiction. Arch. Iran. Med.,
6: 32-34.
28. Dow, C.A., S.B. Going, H.H.S. Chow, B.S. Patil and
C.A. Thomson, 2012. The effects of daily consumption
of grapefruit on body weight, lipids and blood
pressure in healthy, overweight adults. Metabolism,
61: 1026-1035.
70
