Yog 2
Yog 2
Yog 2
Anaerobe
journal homepage: www.elsevier.com/locate/anaerobe
Clinical microbiology
Institute of Food Science & Technology, Fuzhou University, No. 2 Xueyuan Road, Fuzhou 350108, PR China
Almond Board of California, 1150 9th Street, Suite 1500, Modesto, CA 95354, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 7 August 2013
Received in revised form
5 November 2013
Accepted 22 November 2013
Available online 3 December 2013
Almonds and almond skins are rich in ber and other components that have potential prebiotic properties. In this study we investigated the prebiotic effects of almond and almond skin intake in healthy
humans. A total of 48 healthy adult volunteers consumed a daily dose of roasted almonds (56 g), almond
skins (10 g), or commercial fructooligosaccharides (8 g) (as positive control) for 6 weeks. Fecal samples
were collected at dened time points and analyzed for microbiota composition and selected indicators of
microbial activity. Different strains of intestinal bacteria had varying degrees of growth sensitivity to
almonds or almond skins. Signicant increases in the populations of Bidobacterium spp. and Lactobacillus spp. were observed in fecal samples as a consequence of almond or almond skin supplementation.
However, the populations of Escherichia coli did not change signicantly, while the growth of the
pathogen Clostridum perfringens was signicantly repressed. Modication of the intestinal microbiota
composition induced changes in bacterial enzyme activities, specically a signicant increase in fecal bgalactosidase activity and decreases in fecal b-glucuronidase, nitroreductase and azoreductase activities.
Our observations suggest that almond and almond skin ingestion may lead to an improvement in the
intestinal microbiota prole and a modication of the intestinal bacterial activities, which would induce
the promotion of health benecial factors and the inhibition of harmful factors. Thus we believe that
almonds and almond skins possess potential prebiotic properties.
2013 Elsevier Ltd. All rights reserved.
Keywords:
Almond
Almond skins
Prebiotics
Intestinal microbiota
Bacterial enzymes
1. Introduction
It is well established that the colonic microbiota have a profound inuence on health. The human gut contains a large variety
of bacterial genera, species, and strains, which are either benecial
(e.g., Bidobacterium spp. and Lactobacillus spp.) or harmful (e.g.,
Clostridium spp., Shigella spp., and Veillonella spp.) to host health.
The intestinal microbiota play an essential role in inuencing the
health of the host. They can greatly inuence the intestinal environment [1], contributing to the hosts health through a variety of
mechanisms such as activation of the immune response, production of bacteriocins, nutritional and physical competition with
pathogens, and maintenance of an acid environment.
Currently there is great interest in the use of prebiotics as
functional food ingredients to manipulate the composition of
colonic microbiota in order to improve health [2]. Many food oligosaccharides and polysaccharides (including dietary ber), such as
fructooligosaccharides, inulin, galactooligosaccharides, and other
aminobenzoic acid per minute, respectively. One unit of azoreductase activity was dened as1 mmol of amaranth metabolized
per minute. Results were expressed as units per gram (U/g) of wet
feces.
2.7. Statistical analysis
Results were expressed as mean SD (standard deviation) or
mean SEM (standard error of the mean). Paired-sample t-tests
were applied to compare the data among study groups. The statistical analysis of the results was performed by the SPSS 13 software and the signicance threshold was set at 5% (P < 0.05).
3. Results
3.1. Characteristics of the study subjects
Throughout the whole experimental period, there were no
serious adverse reactions to the diets reported by the subjects,
except for 4 subjects in the FOS control group reported mild diarrhea on the rst week of FOS ingestion, however they recovered
naturally a few days later. All of the 16 subjects in each group were
in good health, except for two subjects, one in control group and
the other in the almond group who failed to nish the study for
personal reasons. The weight, BMI, body fat percentage, body water
content and muscle mass were measured at the beginning (Baseline, week 0) and after 6 weeks of treatment (Ingestion, week 6).
Daily water intake and defecation frequency per week were
recorded by the subjects themselves. Details of the study group
members physical characteristics, water intake and defecation are
provided in Table 1. There were no statistically signicant differences in these parameters within and among the groups.
3.2. Water content and pH of feces
Changes in the fecal moisture and fecal pH of the subject groups
are shown in Table 2. Overall, no signicant differences were
observed in fecal moisture and fecal pH levels throughout the
experiment, except for the control group, which showed a signicant decrease in fecal pH (P < 0.05) at week 3. Also, in the rst week
of daily consumption of FOS by the control group, the fecal samples
seemed to be moister and 4 subjects reported mild diarrhea. Fecal
moisture in the almond group tended to (P < 0.1) decrease at week
3 and week 6.
3.3. Bacterial populations
The effects of almonds and almond skins intake on the
composition of fecal microbiota in the healthy adult volunteers
are shown in Table 3. Bidobacteria levels increased signicantly
after 6 weeks of ingesting FOS (P < 0.01), almond skins (P < 0.01)
or almonds (P < 0.05). The control and almond skin groups
Table 1
Body composition, water intake and defecation of study volunteers at start and end of 6-week study.a
Parameter
Weight (kg)
BMI (kg/m2)
Body fat percentage (%)
Body water content (%)
Muscle mass (%)
Daily water intake (ml)
Defecation (no.) per week
58.56
21.22
20.61
54.26
43.98
884
6
Baseline, week 0
9.39
2.13
8.07
4.52
8.52
478
2
Ingestion, week 6
57.09
20.70
19.19
55.20
43.63
955
7
8.62
1.99
8.06
4.39
7.87
473
2
Baseline, week 0
57.41
20.83
19.90
54.51
43.66
914
7
7.99
1.74
7.01
4.40
8.26
300
1
Ingestion, week 6
55.98
20.33
18.31
55.62
43.43
1184
7
7.60
1.63
6.71
4.12
8.08
651
2
Baseline, week 0
57.59
20.53
21.06
53.98
43.62
956
6
8.10
2.06
5.66
3.32
8.46
558
2
Ingestion, week 6
56.77
20.25
18.96
55.06
43.69
1491
7
8.14
2.01
5.64
2.84
8.26
1060
3
Table 2
Changes in fecal moisture and pH during the treatment period for all subjects.a
Feces characteristic
Baseline
Ingestion
Week 0
Week 1
Week 3
Week 6
Week 8
Evacuation
80.53 5.60
80.64 6.60
79.53 8.89
83.10 4.47
80.95 5.43
78.67 6.40
82.81 3.86
80.36 6.15
76.60 6.75
80.36 5.87
79.53 7.28
76.73 5.67
80.71 6.77
77.76 8.42
76.69 7.67
6.36 0.58
6.31 0.69
6.16 0.67
6.11 0.61
6.16 0.81
6.24 0.74
5.57 0.55*b
6.06 0.80
6.26 0.52
6.04 0.83
6.25 0.63
6.30 0.54
5.75 0.69
6.12 0.65
6.32 0.40
4. Discussion
Healthy adult humans were used as the subjects in this study to
evaluate the impact of the consumption of almonds and almond
skins on human intestinal microbiota. A total of 48 healthy college
students were involved in an intervention trial including a 2-week
run-in period, a 6-week treatment period, and a 2-week wash-out
period. The 48 subjects were not selected randomly. A screening
procedure was carried out rst. The criteria were the stability of
intestinal bacteria of subjects. In the screening procedure, 80
healthy volunteers recruited from our campus. They all lived in
school dormitories with a similar environment, and were instructed to start dietary restrictions for 2 weeks. During this time, they
were offered the basic diet we mentioned above and not allowed to
have any other food. Following the methods mentioned above, we
collected the fecal samples of all the 80 volunteers on 0th, 7th and
14th day of the 2-week screening procedure period, and then we
enumerated the populations of Bidobacterium spp., Lactobacillus
spp., E. coli and C. perfringens in the stools (data were not shown).
Based on the stability of intestinal bacteria, 48 subjects were
selected from the 80 volunteers. We assumed that the selected 48
subjects would remain stable in the short term (for example, not
more than ten weeks) if they remained in the same environment
and kept the dietary restriction. Based on this assumption, we
dened the populations of the four intestinal bacteria as the
baseline, and considered this 2-week screening procedure as the
Table 3
Effect of almond skin and almond intake on fecal microbiota populations in healthy human subjects.a
Fecal organism
Bidobacterium spp.
FOS control (n 15)
Almond skin (n 16)
Almond (n 15)
Lactobacillus spp.
FOS control (n 15)
Almond skin (n 16)
Almond (n 15)
E. coli
FOS control (n 15)
Almond skin (n 16)
Almond (n 15)
C. perfringens
FOS control (n 15)
Almond skin (n 16)
Almond (n 15)
Ingestion
Week 0
Week 1
Week 3
Week 6
Week 8
Evacuation
9.84 0.10
9.64 0.10
9.75 0.16
11.01 0.11**b
10.16 0.10**
9.75 0.12
11.04 0.20**
10.40 0.11**
9.96 0.06
10.90 0.15**
10.15 0.12**
10.29 0.14*
10.38 0.12**^
^
10.22 0.15**
10.47 0.17**^
^
8.99 0.19
8.77 0.14
8.92 0.12
9.18 0.15
9.19 0.16*
9.05 0.11
9.35 0.13
9.38 0.12**
8.91 0.17
9.54 0.19*
9.57 0.12**
9.39 0.12**
9.53 0.14*
9.34 0.12**
9.13 0.23
9.28 0.18
9.23 0.16
9.38 0.13
9.19 0.15
9.34 0.11
9.28 0.09
9.38 0.12
9.24 0.11
9.35 0.16
9.30 0.13
9.30 0.12
9.41 0.09
9.39 0.12
9.31 0.13
9.56 0.12
6.24 0.21
6.27 0.25
6.64 0.19
5.77 1.13
6.03 0.42
6.24 0.35
5.89 0.33
6.14 0.31
6.36 0.37
5.44 0.37*
5.66 0.28*
6.19 0.24
6.33 0.28^
6.13 0.23^
6.27 0.22
Within rows, symbol (*) indicates signicant difference from the value on week 0 (baseline) at P < 0.05, (**) indicates signicant difference from the value on week 0 (baseline)
at P < 0.01, and (^) indicates signicant difference from the value on week 6 (Ingestion) at P < 0.01.
a
Values are expressed as the mean SEM.
15.00
**
15.00
14.50
14.00
13.50
14.50
14.00
13.50
13.00
13.00
FOS Control
Almond Skin
FOS Control
Almond
A
**
Almond
15.00
*
14.00
52.00
**
Almond Skin
50.00
48.00
46.00
44.00
42.00
13.00
12.00
11.00
10.00
9.00
40.00
FOS Control Almond Skin
Almond
8.00
FOS Control
Almond Skin
Almond
Fig. 1. Effects of almond skin and almond consumption on fecal bacterial enzyme activities in healthy human subjects. Fecal samples were examined at the beginning (week 0) and
after 6 weeks (week 6) of FOS (control), almond skin or almond ingestion. Values are means SD; symbol (*) indicates signicantly different means at P < 0.05; symbol (*) indicates
signicantly different means at P < 0.01.
fecal pH and water content did not change signicantly for any
subject. Fecal pH may not accurately reect the pH in the colon and
depends on absorption of short chain fatty acids (SCFAs) and bicarbonate secretion. Bouhnik et al. [17], who studied the prolonged
administration of low-dose inulin that stimulates the growth of
bidobacteria in humans, found no change in fecal pH and SCFA
levels.
After six weeks of almond skin or almond ingestion, the populations of Bidobacterium spp. and Lactobacillus spp. increased
signicantly. Bidobacteria and lactobacilliare are predominant
members of the intestinal microbiota of humans. Bidobacteria are
thought to stimulate the immune system, produce B vitamins,
inhibit pathogen growth, reduce blood ammonia and blood
cholesterol levels, and help to restore the normal ora after antibiotic therapy [18]. Lactobacilli may aid digestion of lactose in
lactose-intolerant individuals, reduce constipation and infantile
diarrhea, help resist infections such as salmonellae and help to
relieve irritable bowel syndrome [19]. The results of the present
study indicated that almond skins and almonds possess bidobacteria and lactobacili stimulation effects. Also, we found that the
stimulation effects of almond skin and almond intake on bidobacteria and lactobacilli were different. Almond skin intake induced
a prompt increase of bidobacteria and lactobacilli, similar to FOS
intake, although the bidogenic effect was not as great as FOS
intake (P < 0.05), and a high level of viable bidobacteria and
lactobacilli remained for 2 weeks after the ingestion of the almond
skins. By contrast, the stimulation effect of almond intake was not
obvious until the end of 6 weeks. However, eventually (week 6) the
populations of bidobacteria or lactobacilli in almond skin group
and almond group reached a similar level (no signicant difference). Almond skin or almond ingestion for 6 weeks also induced a
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