Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 947 of 967

Research Article Open Access

Effects of the consumption of Acacia bark-derived

proanthocyanidins on visceral fat: A randomized, double-blind,
placebo-controlled, parallel-group comparative study
Asami Baba 1*, Sosuke Ogawa2, Kengo Yokosho2, Naoko Suzuki 1, Tsuyoshi Takara3

1
ORTHOMEDICO Inc. 2F Sumitomo Fudosan Korakuen Bldg., 1-4-1 Koishikawa, Bunkyo-ku, Tokyo, 112-0002, Japan; 2Acacia-
No-Ki Co., Ltd. 4291-1 Miyauchi, Hatsukaichi-shi, Hiroshima, 738-0034, Japan; 3 Medical Corporation Seishinkai, Takara Clinic
9F Taisei Bldg., 2-3-2, Higashi-gotanda, Shinagawa-ku, Tokyo, 141-0022, Japan

*Corresponding author: Asami Baba, ORTHOMEDICO Inc. 2F Sumitomo Fudosan Korakuen Bldg., 1-4-1 Koishikawa, Bunkyo-
ku, Tokyo, 112-0002, Japan.

Submission Date: September 27th, 2024, Acceptance Date: December 5th, 2024, Publication Date: December 11th, 2024

Please cite this article as: Baba A., Ogawa S., Yokosho K., Suzuki N., Takara T. Effects of the consumption of Acacia bark-
derived proanthocyanidins on visceral fat: A randomized, double-blind, placebo-controlled, parallel-group comparative
study. Functional Foods in Health and Disease 2024; 14(12): 946-967. DOI: https://doi.org/10.31989/ffhd.v14i12.1420

ABSTRACT
Background: Obesity is an important risk factor for diabetes, kidney, liver, and cardiovascular diseases, and chronic
obstructive pulmonary disease. Factors such as dietary choices, decreased physical activity due to urbanization and
economic expansion, and increased nutritional consumption have made obesity a growing global issue. Acacia bark
extract contains proanthocyanidins, which are believed to inhibit accumulation of fat and promote the metabolism of
body fat.

Objective: This study was designed to investigate the effects of Acacia bark-derived proanthocyanidins on fat
accumulation among healthy Japanese adults with a body mass index (BMI) of 25–30 kg/m2.

Methods: This study was performed as a randomized, placebo-controlled, double-blind, parallel-group study. Out of 199
participants who signed informed consent, 68 were randomized to two groups (n = 34/group) with a computer-generated
allocation table. The participants took six tablets containing either Acacia bark-derived proanthocyanidins (Acacia group)
or placebo (Placebo group) daily for 12 weeks. Their abdominal total, subcutaneous, and visceral fat areas via X-ray
computed tomography, body weight, BMI, fat and muscle mass, and waist, abdominal, and hip circumferences were
measured.

Results: After accounting for dropouts, the Acacia and Placebo groups had 32 and 33 participants, respectively. At the
end of the 12-week intervention, the Acacia group had significantly lower abdominal visceral fat area compared with the
Placebo group.
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 948 of 967

Conclusions: Acacia proanthocyanidins were found to reduce visceral fat.

Keywords: Acacia bark extract, proanthocyanidins, polyphenol, visceral fat, obesity

Trial registration number: UMIN000050201

Graphical Abstract

©FFC 2024. This is an Open Access article distributed under the terms of the Creative Commons Attribution 4.0 License
(http://creativecommons.org/licenses/by/4.0)

INTRODUCTION chronic obstructive pulmonary disease [2–4]. Obesity is

Obesity refers to excessive fat buildup in adipose tissue. now becoming a global issue because of poor dietary
In Japan, obesity is defined as a body mass index (BMI) choices, decreased physical activity, and increased
2
≥25 kg/m [1]. Being obese is a key hazard factor for nutritional consumption [5,6]. According to the World
diabetes; kidney, liver, and cardiovascular diseases; and Health Organization, 43% of male and female older than
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 949 of 967

18 years with a BMI of ≥25 kg/m2 are obese in 2022 [7]. This study was designed to evaluate the anti-obesity
In the 2019 National Health and Nutrition Survey, 27.2% effects of Acacia bark-derived proanthocyanidins on
of Japanese both sexes aged older than 20 with a BMI of Japanese adults with a BMI of 25–30 kg/m2 (class I
≥25 kg/m2 are obese [8]. Large quantities of obesity) without health problems.
inflammatory cytokines (e.g., tumor necrosis factor-α
(TNF-α) and interleukin-6) are produced by visceral METHODS
adipose tissues [9], and the expression of these cytokines Study design: A 1:1 allocation ratio was used in this
promotes skeletal muscle degradation [10]. Recently, randomized, placebo-controlled, double-blind, parallel-
several studies have found that increased visceral fat group study. The study protocol had the approval of the
area is linked to lower skeletal muscle mass [11–13]. Institutional Ethical Review Board at Medical Corporation
Notably, the loss of skeletal muscles is associated with Seishinkai, Takara Clinic (approval date: January 18, 2023,
decreased quality of life (QOL), basal metabolic rate, and approval number: 2301-04148-0041-0C-TC). The
bone mineral density and increased fracture risk [14]. In protocol of this trial was registered in the University
Japan, the leading causes of death include cardiac and Hospital Medical Information Network Clinical Trial
cerebrovascular diseases, pneumonia, and renal failure Registry (UMIN-CTR; trial registration number:
[15], wherein obesity is a risk factor. Thus, it is important UMIN000050201) in advance. Moreover, the study
to reduce excess body fat accumulation to maintain the faithfully adhered to the tenets of the Declaration of
QOL and extend the healthy lifespan of the Japanese Helsinki (revised version of 2013) and the Ethical
population. Guidelines for Medical and Health Research Involving
Acacia bark extract is a hydrothermal extract that is Human Subjects.
derived from the bark of Acacia mearnsii De Wild.. Acacia
bark is composed of various compounds, including Eligibility criteria: The following selection criteria were
monomers, dimers, trimers, and condensed polymers of established: (1) Japanese adults of both sexes, (2) BMI
the basic backbone of flavan-3-ols (i.e., gallocatechin and between 25 kg/m2 and 30 kg/m2, and (3) the top 68
robinetinidol) [16,17]. Among them, polyphenols individuals with the largest visceral fat area at screening
comprise approximately 80% of the extract [18,19]. The examination (Scr) among those meeting criteria (1) and
proanthocyanidins derived from grape seeds [20] and (2). The details of the exclusion criteria are registered in
pine bark [21] contain a polymerized structure with two the UMIN-CTR (https://center6.umin.ac.jp/cgi-open-
hydroxy groups each in the A and B rings; by contrast, the bin/ctr_e/ctr_view.cgi?recptno=R000057170).
proanthocyanidins from Acacia bark also have a The participants were gathered through
polymerized structure but with one and three hydroxy GOTOROKU (https://www.go106.jp/), a volunteer
groups in the A and B rings, respectively [22,23]. Acacia recruitment site administered by ORTHOMEDICO Inc.
bark extract prevented fat absorption in the body of mice (Tokyo, Japan). Those who wished to participate in this
by restricting lipase activity [24,25]. According to studies study were fully informed of the study’s content via the
using mouse models of obesity and diabetes (KK-Ay), network, and consent was obtained electronically.
Acacia bark extract can (1) activate AMP kinase (a central Meanwhile, those who were concerned with the lead or
regulator in lipid metabolism), (2) improve lipid sponsor companies of the study were excluded. Medical
metabolism by reducing fat synthesis and accumulation Corporation Seishinkai, Takara Clinic (Tokyo, Japan) was
and promoting hepatic lipolysis, and (3) increase energy the institution responsible for conducting this study,
expenditure in skeletal muscle [26]. Therefore, Acacia evaluating the obtained data and managing the
bark-derived proanthocyanidins may suppress body fat participants’ physical condition. Examinations were
accumulation and promote body fat metabolism and may performed together with Nerima Medical Association,
be considered as functional food [27,28]. Minami-machi Clinic (Tokyo, Japan).
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 950 of 967

Intervention: The nutritional facts per daily intake for Assessments for each item were performed at Scr and
each test food are described in Table 1. The Acacia bark after 4, 8, and 12 weeks of intervention. Moreover, an
extract, one of the test food ingredients, was provided as additional post-intervention examination was conducted
described below: Acacia bark was crushed and extracted at 4 weeks after completing the 12-week intervention.
with boiling water, and the product was spray-dried. The This study's primary outcome was the measured
product is rich in proanthocyanidins, commonly known as
value of abdominal visceral fat area at 12 weeks. The
wattle tannins[25]. The Folin–Ciocalteu method (gallic
secondary outcomes included the abdominal visceral fat
acid equivalence method) was used for measuring the
area; total abdominal fat area; abdominal subcutaneous
total amount of proanthocyanidins in the test food [27].
fat area; body weight; BMI; body fat percentage; fat and
Moreover, the concentration of robinetinidol-(4α,8)-
muscle mass; and waist, abdominal, and hip
catechin concentration of the tablet was analyzed by
high-performance liquid chromatography in accordance circumferences at 4, 8, and 12 weeks and at 4 weeks post-
with a previously described qualitative analysis method intervention.
[29]. This study participants took six pieces per day of (1) Measurement of fat areas: The subcutaneous,
tablet containing Acacia bark-derived proanthocyanidins visceral, and total fat areas of the cross-sectional area of
(Acacia group; dose: 245 mg/day) or placebo (Placebo the umbilical region were determined via X-ray
group). The tables were consumed before meals with computed tomography (NAEOTOM Alpha, Siemens,
water and without chewing. The intervention period was Munich, Germany).
12 weeks. The indistinguishable appearance of the two (2) Physical examination: Body weight, BMI, body
tablets was confirmed by the ethics committee. The fat percentage, and fat and muscle mass were evaluated
Acacia bark-derived proanthocyanidins were produced using a multi-frequency body composition monitor (MC-
by Acacia-No-Ki Co., Ltd. (Hiroshima, Japan), and the
780A-N, TANITA Corporation, Tokyo, Japan). The waist,
intervention foods were provided by Acacia-No-Ki Co., Ltd.
abdominal, and hip circumferences were measured using
Outcomes: The study schedule is shown in Table 2. a cloth measuring tape.

Table 1. Nutritional facts per daily intake (6 tablets)

Acacia bark-derived Placebo tablets
proanthocyanidins tablets
Water 0.0630 mg 0.0504 mg
Proteins 0.0522 mg 0.0468 mg
Fat 0.0558 mg 0.0540 mg
Ash 0.0198 mg 0.0126 mg
Carbohydrates 1.6092 mg 1.6362 mg
Calories 7.1460 mg 7.2180 mg
Sodium 0.3492 mg 0.1548 mg
Salt equivalents 0.0009 mg 0.0004 mg
Proanthocyanidins 0.1980 mg ND

Proanthocyanidins derived from Acacia bark 0.2694 mg ND

ND, not detectable

(3) Confirmation of adverse events and safety abnormalities after the intervention was observed. The
items: The safety assessment included physical group and individual data of these safety assessments
measurements, urinalysis, and blood tests. The were also reviewed for health management.
percentage of participants with previously normal The physical measurements included body weight,
urinalysis and blood test data at Scr who developed BMI, and systolic and diastolic blood pressure. BMI was
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 951 of 967

calculated using the height measured at Scr. Body weight Sample size: Considering that no study has evaluated
was evaluated using the multi-frequency body abdominal visceral fat area after the consumption of
composition monitor (MC-780A-N, TANITA Corporation). Acacia bark-derived proanthocyanidins, we anticipated a
Systolic and diastolic blood pressures were measured large difference between the two groups and used d =
using an automatic blood pressure measuring device 0.80 according to the suggestion of Cohen [31]. The level
(Omron HEM-6022, OMRON Corporation, Kyoto, Japan). of statistical significance (α) and statistical power (1-β)

In urinalysis, protein, glucose, pH, and occult blood were determined to be 0.05 and 0.80, respectively. At

were measured. LSI Medience Corporation (Tokyo, least 52 participants (26 in each group) were needed, and

Japan), the clinical inspection company, was entrusted the sample size was set at 68 (34 in each group) in

with the measurement of each item. expectation of withdrawal and out-of-compliance during

The following items were evaluated in the blood the period of the study.

tests: white and red blood cell counts, hemoglobin,

hematocrit, platelet count, aspartate and alanine Selection, randomization, and blinding: Out of the 199

aminotransferases, γ-glutamyl transpeptidase, total participants who agreed to be part of this study, 68 were

bilirubin, total protein, urea nitrogen, creatinine, uric selected by the principal physician and included. The test

acid, sodium, potassium, chloride, serum amylase, total foods were supplied to the contract clinical trial
organization (CRO) by this study’s sponsor (Acacia-No-Ki
cholesterol, high-density and low-density lipoprotein-
Co., Ltd.). The shipping staff of the CRO, who was
cholesterols, triglyceride, glucose, and hemoglobin A1c
responsible for sending the appropriate study food to the
(NGSP). The standard method of LSI Medience
participants, confirmed that the study foods were
Corporation was used to measure each item.
indistinguishable, entered and confirmed the Scr data,
A count of the number of adverse events was
and provided the identifier to the allocator, who did not
performed. In such cases, the physician was instructed to
have direct involvement in the study.
immediately take the necessary and appropriate
The study participants who conformed to the
measures. They also decided whether the participant
eligibility criteria were randomized to either the Acacia or
could continue in the study or not, and whether the
Placebo groups on a 1:1 basis using a computer-
emergency key should be opened. Furthermore, the
generated allocation table (34 participants per group)
physician was instructed to assess and report the
supplied by the allocator. The allocation table was
association between the adverse event and the
generated by stratified block random assignment using
intervention.
SPSS version 23 (IBM Japan, Ltd.; Tokyo, Japan), with sex
At each examination, surveys and a dietary
(male and female) as the stratification factor. This table
assessment using the Calorie and Nutrition Diary [30]
was only used by the shipping staff of the CRO. All of the
were administered to determine the health conditions of
physicians, study site administrators, monitoring
study participants. The survey was conducted three days
personnel, statistical analysts, ethics committee
prior to each visit. Study participants were also required
constituents, and clinical laboratories were blinded to the
to keep a logbook of their daily living status, including
allocation. Until the statistical analysis methods were
test food consumption, physical condition changes, and
fixed, the allocation table was placed under seal and kept
medication consumption.
by the allocator.
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 952 of 967

Table 2. Schedule of enrolment, interventions, and assessments

Briefing Scr Registration Allocation Study periods
session Start 4 8 12 4 weeks
intervention weeks weeks weeks post-
intervention
Enrollment Eligibility screen X X

Informed consent (include X

other procedures)
Allocation X

Interventions Acacia bark-derived

proanthocyanidins tablets
Placebo tablets

Assessments X-ray CT scan (visceral fat X X X X X

area, subcutaneous fat area,
and total fat area)
Circumference of abdominal, X X X X X
waist, and hip
Physical examination X X X X X

Urinalysis X X X X X

Blood test X X X X X

Interview (usually medical) X X X X X

Dietary Survey* X X X X X

Logbook

Scr, screening examination; 4, 8, and 12 weeks, after 4, 8, and 12 weeks of intervention; 4 weeks post-intervention, additional post-intervention examination done 4 weeks after the end of the 12-week
intervention.
*For the dietary survey, data was obtained for the three days prior to each examination date.
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 953 of 967

Statistical analysis: All statistical analyses were Placebo group) and 95% CI.
performed using the two-tailed method with a 5% level The pharmaceutical guideline (ICH-E9 [32]) was
of significance by SPSS version 23 (IBM Japan, Ltd.). The used to define the datasets in this study.
primary outcome was a between-group comparison of
the visceral fat area at 12 weeks, and no adjustments RESULTS
were made for multiple items or time points in the Participant flow and characteristics: Figure 1 shows a
secondary outcomes. flowchart of the study participants. Recruitment of the
The characteristics of participants were participants was conducted between January 31 and
demographically aggregated for inclusion and analysis of February 24, 2023, and the study period was from
participants. Sex was represented as the number of February 4 to October 1, 2023.
participants and percentage within the group. Age, All participants received their assigned
height, body weight, BMI, and systolic and diastolic blood intervention. Notably, three participants did not undergo
pressure were indicated as the mean and standard any evaluations after Scr (Acacia group, n = 2; Placebo
deviation (SD), minimum (Min), and maximum (Max). group, n = 1) and were excluded from the analysis
The primary outcome, secondary outcomes, and because they did not receive any intervention after
safety assessment items were reported as the mean and assignment. One participant was unable to undergo
SD for measured values. Values at Scr were considered as evaluation at 4 weeks. The efficacy and safety analysis
the baseline and compared across groups with Welch’s t- datasets for this study included the full analysis set and
test. For between-group comparisons of measures of the safety analysis population, both of which included 65
primary and secondary outcomes at other timepoints (4, participants (Acacia group, n = 32; Placebo group, n = 33).
8, and 12 weeks, and 4 weeks post-intervention), a linear Table 3 shows the patient characteristics in each analysis
mixed model with baseline values as covariates was used. dataset, including intention to treat.
The between-group differences (Acacia group minus
Placebo group) and their 95% confidence intervals (CIs) Fat areas in the transverse section of the umbilical
were also shown. Between-group differences were region: No significant intergroup differences in terms of
represented at Scr and post-intervention as the mean visceral, subcutaneous, and total fat areas were observed
and estimated marginal mean (EMM), respectively. at Scr. The primary outcome (visceral fat area at 12
For safety evaluation, the adverse event frequency weeks) was significantly lower in the Acacia group than in
in each group was calculated, and the 95% CIs for the the Placebo group (EMM ± standard error [SE]: 103.5 ±
occurrence of adverse events in each group and the 2.8 vs. 111.6 ± 2.8 cm2; between-group difference
deviation of the occurrence between groups were [95%CI], −8.1 cm2 [−15.9, −0.3]; P = 0.041; Figure 2 and
calculated. The chi-square test was used to provide a Table 4). Conversely, the Acacia group had a significantly
measure of the adverse event frequency in each group higher subcutaneous fat area at 12 weeks and at 4 weeks
and the percentage of participants who developed post-intervention compared with the Placebo group
abnormal urinalysis and peripheral blood test results (EMM ± SE: 12 weeks, 264.1 ± 4.1 vs. 246.8 ± 4.1 cm2;
after the intervention. between-group difference [95%CI], 17.3 cm2 [5.8, 28.9];
Welch’s t-test was used to compare dietary survey P = 0.004; 4 weeks post-intervention, 262.7 ± 4.1 vs.
data, which were indicated as the mean, SD, median 248.5 ± 4.1 cm2; between-group difference [95%CI],
(Med), Min, and Max, with differences between groups 14.3 cm2 [2.7, 25.8]; P = 0.016; Figure 2 and Table 4).
shown as the mean difference (Acacia group minus Total fat area was not significantly different between
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 954 of 967

groups. causally related to the food intervention in the study.

With regard to urinalysis and blood test data,
Body composition and abdominal, waist, and hip significant intergroup differences were observed for
circumferences: No significant intergroup difference in urinary protein, total protein, total cholesterol, and low-
body composition and abdominal, waist, and hip density lipoprotein-cholesterol (Supplementary Table 1).
circumferences was observed at all time points (Table 5). Considering that total cholesterol and low-density
lipoprotein-cholesterol fluctuated outside the reference
Confirmation of adverse events and safety items: Three values after the intervention in the Acacia group, each
participants in the Acacia group experienced adverse blood test was reviewed by the study physician on a
events. One participant had toothache, cough, colds, group basis (Supplementary Table 4), and the
stomach pain, and back pain; another participant had fluctuations were deemed as not medically significant
common colds; and the last participant had periodontal (Supplementary Table 4). Moreover, the study physician
inflammation. Meanwhile, three participants in the reviewed the safety assessment items (physical
Placebo group also reported adverse events. One examination, urinalysis, and blood tests) on a group
participant had common colds, another participant had (Supplementary Tables 2–4) and individual basis (data
dysmenorrhea, and the last participant had dizziness. All not shown). No medically relevant changes related to test
symptoms were deemed by the study physician as not food consumption were observed.

Figure 1. Flow diagram

Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 955 of 967
Table 3. Participant characteristics
ITT FAS, SAF

Acacia group Placebo group Acacia group Placebo group

(n = 34) (n = 34) (n = 32) (n = 33)
Sex Male 17 (50.0%) 17 (50.0%) 17 (53.1%) 17 (51.5%)
Female 17 (50.0%) 17 (50.0%) 15 (46.9%) 16 (48.5%)
Age (years) Mean (SD) 49.3 (8.9) 49.6 (10.4) 49.1 (8.8) 49.8 (10.5)
Min–Max 31–67 29–74 31–67 29–74
Height (cm) Mean (SD) 163.3 (7.2) 163.2 (6.7) 163.6 (7.3) 163.2 (6.8)
Min–Max 149.0–179.5 151.2–174.3 149.0–179.5 151.2–174.3
Body weight (kg) Mean (SD) 72.1 (6.7) 72.8 (7.9) 72.3 (6.8) 72.9 (8.0)
Min–Max 58.8–88.2 57.1–87.9 58.8–88.2 57.1–87.9
Body mass index (kg/m2) Mean (SD) 27.0 (1.5) 27.3 (1.4) 27.0 (1.6) 27.3 (1.4)
Min–Max 25.0–29.8 25.0–29.6 25.0–29.8 25.0–29.6
Systolic blood pressure (mmHg) Mean (SD) 132.3 (19.7) 125.4 (16.9) 132.4 (20.3) 125.6 (17.2)
Min–Max 101–201 96–177 101–201 96–177
Diastolic blood pressure (mmHg) Mean (SD) 86.6 (14.4) 82.3 (13.5) 86.7 (14.8) 82.5 (13.6)
Min–Max 62–126 59–116 62–126 59–116
Sex is indicated by the number of participants and their percentage in the group. Other items are indicated by mean, standard deviation (SD), minimum
(Min), and maximum (Max).
ITT, intention to treat; FAS, full analysis set; SAF, safety analysis population

Table 4. Visceral fat area, subcutaneous fat area, and total fat area in transverse section of umbilical region
Acacia group Placebo group Group difference P value
n Mean SD n Mean SD [95%CI]
(EMM) [95%CI] (EMM) [95%CI]
Total fat Scr 32 344.8 72.2 33 365.6 62.0 –20.8 0.218
area (cm2) [–54.2, 12.6]
4 weeks 32 348.6 70.4 32 367.8 66.1 –0.4 0.959
(358.5) [348.9, 368.0] (358.8) [349.4, 368.3] [–13.9, 13.2]
8 weeks 32 350.7 68.8 33 364.8 63.1 4.6 0.500
(360.1) [350.6, 369.7] (355.5) [346.1, 364.9] [–8.9, 18.1]
12 weeks 32 357.7 68.5 33 368.3 64.0 7.9 0.248
(367.0) [357.4, 376.6] (359.1) [349.7, 368.5] [–5.6, 21.4]
4 weeks post- 32 359.2 71.2 33 367.1 67.2 11.8 0.087
intervention (369.1) [359.5, 378.7] (357.3) [347.9, 366.7] [–1.7, 25.3]
Visceral fat Scr 32 107.8 20.4 33 110.7 17.8 –2.9 0.542
area (cm2) [–12.4, 6.6]
4 weeks 32 101.6 28.5 32 104.7 24.7 –0.9 0.821
(102.8) [97.3, 108.3] (103.7) [98.2, 109.2] [–8.7, 6.9]
8 weeks 32 103.3 24.8 33 107.1 17.1 –1.3 0.736
(104.5) [99.0, 110.1] (105.9) [100.4, 111.3] [–9.1, 6.4]
12 weeks 32 102.1 21.0 33 113.1 25.0 –8.1 0.041*

(103.5) [98.0, 109.0] (111.6) [106.2, 117.1] [–15.9, –0.3]

4 weeks post- 32 105.0 19.2 33 110.3 22.4 –3.0 0.449

intervention
(106.1) [100.6, 111.6] (109.1) [103.7, 114.5] [–10.8, 4.8]
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 956 of 967
Table 4. Visceral fat area, subcutaneous fat area, and total fat area in transverse section of umbilical region (Continued)
Acacia group Placebo group Group P
difference value
n Mean SD n Mean SD [95%CI]
(EMM) [95%CI] (EMM) [95%CI]

Subcutaneous fat area Scr 32 237.0 70.0 33 254.9 67.2 –17.9 0.297
(cm2)
[–51.9, 16.1]

4 weeks 32 247.0 69.5 32 263.1 77.0 1.6 0.791

(256.2) [248.0, 264.3] (254.6) [246.5, [–10.0, 13.2]

262.7]
8 weeks 32 247.4 68.4 33 257.7 64.2 5.9 0.313

(255.6) [247.4, 263.8] (249.7) [241.6, [–5.6, 17.5]

257.7]
12 weeks 32 255.6 69.0 33 255.1 68.7 17.3 0.004*

(264.1) [255.9, 272.3] (246.8) [238.7, [5.8, 28.9]

254.9]
4 weeks post- 32 254.2 71.1 33 256.8 65.9 14.3 0.016*
intervention
(262.7) [254.5, 270.9] (248.5) [240.4, [2.7, 25.8]
256.5]
Data are indicated as mean, standard deviation (SD), estimated marginal mean (EMM), 95% confidence interval (CI), and between-group difference and
its 95% CI. Scr, screening examination; 4, 8, and 12 weeks, after 4, 8, and 12 weeks of intervention; 4 weeks post-intervention, additional post-intervention
examination done 4 weeks after the end of the 12-week intervention. *P < 0.05

2
(cm )
150

140

130

120

110

100

90 Acacia group

80 Placebo group

70

60
4 weeks
Scr 4 weeks 8 weeks 12 weeks post-intervention

Figure 2. Changes in visceral fat area in transverse section of umbilical region

Data are presented as mean, standard deviation (SD). Scr, screening examination; 4, 8, and 12 weeks, after 4, 8, and 12 weeks of intervention; 4 weeks
post-intervention, additional post-intervention examination done 4 weeks after the end of the 12-week intervention. *P < 0.05
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 957 of 967

Table 5. Body composition and abdominal, waist, and hip circumferences.

Acacia group Placebo group Group difference P value
n Mean SD n Mean SD [95%CI]
(EMM) [95%CI] (EMM) [95%CI]

Body weight (kg) Scr 32 72.3 6.8 33 72.9 8.0 –0.6 0.739
[–4.3, 3.1]
4 weeks 32 72.2 7.2 32 72.7 8.8 0.4 0.330
(72.6) [72.0, 73.2] (72.1) [71.5, 72.8] [–0.4, 1.3]
8 weeks 32 72.0 7.2 33 72.0 8.5 0.6 0.195
(72.3) [71.7, 72.9] (71.7) [71.1, 72.3] [–0.3, 1.4]
12 weeks 32 72.3 7.3 33 72.1 8.7 0.8 0.072
(72.6) [72.0, 73.3] (71.8) [71.2, 72.4] [–0.1, 1.7]
4 weeks post– 32 72.3 6.9 33 72.1 8.8 0.8 0.083
intervention (72.6) [72.0, 73.3] (71.9) [71.2, 72.5] [–0.1, 1.6]
2
Body mass index (kg/m ) Scr 32 27.0 1.6 33 27.3 1.4 –0.3 0.381
[–1.1, 0.4]
4 weeks 32 27.0 1.6 32 27.2 1.8 0.2 0.305
(27.1) [26.9, 27.4] (27.0) [26.7, 27.2] [–0.2, 0.5]
8 weeks 32 26.8 1.5 33 27.0 1.7 0.2 0.250
(27.0) [26.8, 27.2] (26.8) [26.6, 27.0] [–0.1, 0.5]
12 weeks 32 27.0 1.5 33 27.0 1.7 0.3 0.102
(27.1) [26.9, 27.4] (26.8) [26.6, 27.1] [–0.1, 0.6]
4 weeks post– 32 27.0 1.4 33 27.0 1.8 0.3 0.092
intervention (27.1) [26.9, 27.4] (26.8) [26.6, 27.1] [0.0, 0.6]
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 958 of 967

Table 5. Body composition and abdominal, waist, and hip circumferences (Continued)
Acacia group Placebo group Group difference P value
n Mean SD n Mean SD [95%CI]
(EMM) [95%CI] (EMM) [95%CI]

Body fat percentage Scr 32 31.6 7.7 33 33.0 7.6 –1.4 0.453
(%) [–5.2, 2.4]
4 weeks 32 31.6 8.0 32 32.3 7.2 0.5 0.407
(32.3) [31.4, 33.2] (31.7) [30.8, 32.7] [–0.8, 1.9]
8 weeks 32 31.3 7.5 33 32.2 7.2 0.3 0.617
(31.9) [31.0, 32.8] (31.6) [30.7, 32.5] [–1.0, 1.6]
12 weeks 32 31.6 7.7 33 32.3 7.1 0.7 0.323
(32.3) [31.3, 33.2] (31.6) [30.7, 32.5] [–0.7, 2.0]
4 weeks post-intervention 32 31.8 7.8 33 32.4 7.0 0.7 0.271
(32.4) [31.5, 33.4] (31.7) [30.8, 32.6] [–0.6, 2.0]
Fat mass (kg) Scr 32 22.6 4.9 33 23.9 5.3 –1.3 0.303
[–3.8, 1.2]
4 weeks 32 22.6 5.2 32 23.2 5.2 0.5 0.372
(23.2) [22.4, 24.0] (22.7) [21.9, 23.5] [–0.6, 1.7]
8 weeks 32 22.3 4.8 33 23.1 5.2 0.4 0.536
(22.9) [22.0, 23.7] (22.5) [21.7, 23.3] [–0.8, 1.5]
12 weeks 32 22.6 5.0 33 23.1 5.1 0.7 0.225
(23.2) [22.4, 24.0] (22.5) [21.7, 23.3] [–0.4, 1.8]
4 weeks post-intervention 32 22.8 5.2 33 23.2 5.2 0.8 0.191
(23.4) [22.6, 24.2] (22.6) [21.8, 23.4] [–0.4, 1.9]
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 959 of 967

Table 5. Body composition and abdominal, waist, and hip circumferences (Continued)
Acacia group Placebo group Group difference P value
n Mean SD n Mean SD [95%CI]
(EMM) [95%CI] (EMM) [95%CI]

Muscle mass (kg) Scr 32 46.9 8.5 33 46.3 8.6 0.6 0.770
[–3.6, 4.8]
4 weeks 32 46.9 8.7 32 46.7 8.7 –0.2 0.596
(46.6) [46.0, 47.2] (46.8) [46.3, 47.4] [–1.0, 0.6]
8 weeks 32 47.0 8.5 33 46.3 8.4 0.1 0.833
(46.7) [46.1, 47.2] (46.6) [46.1, 47.1] [–0.7, 0.9]
12 weeks 32 46.9 8.5 33 46.3 8.6 –0.1 0.887
(46.6) [46.1, 47.2] (46.7) [46.1, 47.2] [–0.8, 0.7]
4 weeks post-intervention 32 46.8 8.3 33 46.3 8.4 –0.1 0.867
(46.5) [46.0, 47.1] (46.6) [46.0, 47.1] [–0.8, 0.7]
Waist circumference (cm) Scr 32 88.0 5.2 33 89.2 4.3 –1.2 0.297
[–3.6, 1.1]
4 weeks 32 88.2 5.1 32 90.1 4.9 –0.8 0.315
(88.7) [87.6, 89.8] (89.5) [88.4, 90.6] [–2.3, 0.8]
8 weeks 32 88.1 5.4 33 88.7 4.9 0.4 0.611
(88.6) [87.5, 89.7] (88.2) [87.1, 89.3] [–1.1, 1.9]
12 weeks 32 87.9 5.1 33 88.2 5.1 0.8 0.312
(88.4) [87.4, 89.5] (87.7) [86.6, 88.7] [–0.7, 2.3]
4 weeks post-intervention 32 88.2 4.6 33 89.1 5.4 0.1 0.901
(88.7) [87.6, 89.8] (88.6) [87.5, 89.7] [–1.4, 1.6]
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 960 of 967

Table 5. Body composition and abdominal, waist, and hip circumferences (Continued)
Acacia group Placebo group Group difference P value
n Mean SD n Mean SD [95%CI]
(EMM) [95%CI] (EMM) [95%CI]

Abdominal circumference (cm) Scr 32 94.4 5.3 33 95.9 4.3 –1.5 0.220
[–3.9, 0.9]
4 weeks 32 95.3 4.4 32 95.3 6.0 1.2 0.170
(95.9) [94.7, 97.2] (94.7) [93.5, 95.9] [–0.5, 2.9]
8 weeks 32 95.2 5.2 33 96.3 4.0 –0.1 0.933
(95.7) [94.5, 96.9] (95.8) [94.6, 97.0] [–1.8, 1.6]
12 weeks 32 95.4 4.8 33 95.2 4.8 1.3 0.143
(95.9) [94.7, 97.1] (94.7) [93.5, 95.8] [–0.4, 3.0]
4 weeks post-intervention 32 95.4 5.9 33 95.4 4.0 1.0 0.229
(95.9) [94.7, 97.1] (94.9) [93.7, 96.1] [–0.7, 2.8]
Hip circumference (cm) Scr 32 99.6 4.5 33 99.8 4.0 –0.2 0.874
[–2.3, 2.0]
4 weeks 32 99.7 4.6 32 99.5 4.0 0.4 0.474
(99.8) [99.1, 100.5] (99.4) [98.7, 100.1] [–0.6, 1.4]
8 weeks 32 99.8 4.7 33 99.4 4.4 0.6 0.280
(99.8) [99.1, 100.6] (99.3) [98.6, 100.0] [–0.5, 1.6]
12 weeks 32 99.5 4.6 33 99.4 4.1 0.3 0.586
(99.6) [98.9, 100.3] (99.3) [98.6, 100.0] [–0.7, 1.3]
4 weeks post-intervention 32 99.2 4.5 33 99.1 4.1 0.2 0.629
(99.3) [98.6, 100.0] (99.0) [98.3, 99.7] [–0.8, 1.3]

Data are presented as mean, standard deviation (SD), estimated marginal mean (EMM), 95% confidence interval (CI), and between–group difference and its 95% CI. Scr, screening examination; 4, 8, and 12
weeks, after 4, 8, and 12 weeks of intervention; 4 weeks post–intervention, additional post–intervention examination done 4 weeks after the end of the 12–week intervention.
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 961 of 967

DISCUSSION Americans found that visceral fat, along with BMI and
This study aimed to determine whether Acacia bark- waist circumference, was positively associated with type
derived proanthocyanidins can reduce the visceral fat 2 diabetes risk, but not subcutaneous fat in the abdomen
2
area in Japanese adults with a BMI of 25−30 kg/m and a and thighs [36]. This finding suggests the importance of
relatively large baseline visceral fat area. reducing visceral fat rather than subcutaneous fat to
The Japan Society for the Study of Obesity defines lower the risk of obesity complications. Thus, the
visceral adiposity as having a visceral fat area of ≥100 cm2 consumption of Acacia bark-derived proanthocyanidins
on an abdominal computed tomography scan [1]. In the may contribute to risk reduction of obesity-related
present study, the selection criteria included participants diseases in the future.
2
with a BMI of 25−30 kg/m and a large visceral fat area. Previous in vitro and in vivo studies suggest that the
The mean visceral fat area decreased from 107.8 cm 2 at following mechanisms may be responsible for the visceral
2
Scr to 102.1 cm at 12 weeks in the Acacia group, whereas fat area reduction effect of Acacia bark-derived
it increased from 110.7 to 113.1 cm2 in the Placebo group proanthocyanidins: (1) inhibitory activities of lipase,
2
(EMM group difference: −8.1 cm ). Diet and lifestyle have glucosidase, and α-amylase, (2) increased expression of
been thought to influence fat accumulation [33-34]. heat production–related genes in the skeletal muscle and
Previous studies have reported that (1) dietary liver, (3) suppressed synthesis and uptake of fat in the
components (e.g., soluble dietary fiber, minerals, and liver, and (4) suppression of TNF-α secretion and
vitamins) decrease the visceral fat area [33], (2) enhancement of adiponectin secretion [24–26,37].
monounsaturated fatty acids increase the visceral fat Stomach acid degrades procyanidins to epicatechins and
area [33], and (3) physical activity decreases the visceral catechins, which are then absorbed through the small
fat area [34]. In the present study, dietary assessments intestine [38]. Catechins exert an anti-obesity effect by
conducted three days before each examination period inhibiting digestive activity and macronutrient
revealed no significant intergroup differences regarding absorption in the digestive tract; suppressing anabolism
the aforementioned nutrients (Supplementary Table 5). and promoting catabolism in the liver, muscle, and
With regard to physical activity, the participants were adipose tissue; and inhibiting glycogenesis, synthesis of
asked by the physician to avoid binge eating and drinking, fatty acid, insulation, and heterotopic fat deposition in
as well as retain their usual lifestyle. Taking these factors the muscle and liver [39]. In fact, previous studies in
into account, the effect of diet and lifestyle changes on healthy humans have reported that the ingestion of
visceral fat area in the participants was considerably catechins resulted in significant reductions in body
small. Therefore, the significantly lower abdominal weight and waist circumference [40], besides significant
visceral fat area in the Acacia group than in the Placebo increases in energy expenditure [41]. Therefore,
group was considered as a visceral fat reduction effect catechins, which are metabolites of procyanidins in
caused by proanthocyanidins. Furthermore, the Acacia bark, may contribute to the molecular mechanism
2
abdominal visceral fat area approached 100 cm , which is behind the anti-obesity effect.
the cutoff value proposed by the Japan Society for the Conversely, visceral fat reduction could also be
Study of Obesity [1]. Asian people, including the caused by changes in intestinal microbiota after
Japanese, are more prone to visceral fat accumulation consuming proanthocyanidins. The relationship between
than other races [35]. A cohort study of Japanese obesity and gut microbiota has been demonstrated in
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 962 of 967

many studies [42–44]. A previous study investigated the previous study reported a weak correlation between
intestinal microbiota across healthy Japanese subjects visceral and subcutaneous fat areas [52]. Thus, it may be
stratified by BMI (lean, BMI < 18.5; normal, 18.5 ≤ BMI < possible that some participants in this study had larger
25; obese, BMI ≥ 25) and found that obese subjects had abdominal visceral fat area but smaller abdominal
different microbiota compared to those with lean and subcutaneous fat area. In fact, the maximum and
normal BMI [45]. In addition, Acacia bark-derived minimum abdominal subcutaneous fat areas at Scr were
proanthocyanidins were also reported to modify the 399.6 and 115.0 cm2, respectively, whereas the SD of the
intestinal microbiota in the feces of dermatitis mouse abdominal visceral fat area was 20 cm2. Thus, the anti-
models [46]. Notably, intestinal microflora is reportedly obesity effect of Acacia bark-derived proanthocyanidins
involved in the host’s lipid metabolism [47,48]. was not observed in the abdominal subcutaneous fat
Therefore, the consumption of Acacia bark-derived area, possibly because of variations in the abdominal
proanthocyanidins could have changed the intestinal subcutaneous fat area at Scr.
microflora and enhanced lipid metabolism, thereby The secondary outcomes in this study (i.e., weight,
reducing visceral fat. BMI, and waist circumference) have been reported to be
Visceral and subcutaneous fat are aerobically strongly associated with visceral and subcutaneous fat
metabolized by being broken down into free fatty acids areas [53–55]. Although these parameters displayed no
[49]. Although visceral fat reportedly has a higher changes after the intervention, the data from each study
capacity to release free fatty acids than subcutaneous fat showed that the values of the correlated items differed
[50], they have identical breakdown processes. Visceral across individuals, even among those with similar levels
and subcutaneous fat are collectively referred to as body of visceral and subcutaneous fat areas. This finding
fat, which increases because of the combined effects of suggests that these parameters were not necessarily
excess energy intake and timing of food intake, as well as reduced in conjunction with significant reductions in
increased fat synthesis [51]. Moreover, there is no visceral or subcutaneous fat area. With regard to the
difference in the accumulation process of visceral and body fat percentage, the results were hypothesized to be
subcutaneous fat. As previously reported, Acacia bark- linked to a significant reduction in visceral or
derived proanthocyanidins can reduce fat synthesis and subcutaneous fat area, since the bioelectrical impedance
uptake in the liver [26]. Therefore, we hypothesized that analysis method reflects total body fat [56]. However, no
Acacia bark-derived proanthocyanidins could also have differences were found between groups in the present
an anti-obesity effect by suppressing visceral and study, because the bioelectrical impedance analysis
subcutaneous fat accumulation. However, the method is affected by the amount of water in the body
subcutaneous fat area at 12 weeks and 4 weeks post- due to its principle [56]. Therefore, the effect of the
intervention was significantly lower in the Placebo group Acacia bark-derived proanthocyanidins on body fat
than in the Acacia group. Furthermore, we could not percentage could not be confirmed due to the effect of
confirm the effect of Acacia bark-derived the amount of water. Nevertheless, the primary outcome
proanthocyanidins on subcutaneous fat area because of in this study (i.e., abdominal visceral fat area) was
variations in subcutaneous fat area at Scr. Notably, the significantly smaller in the Acacia group than in the
selection criteria and allocation of this study did not Placebo group at 12 weeks, and the abdominal visceral
consider the abdominal subcutaneous fat area. A fat area increased at 4 weeks post-intervention. Our
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 963 of 967

results suggest that Acacia bark-derived [65]. No studies reported associations between UCP1
proanthocyanidins have an anti-obesity effect on healthy genetic polymorphisms and polyphenols and visceral fat,
Japanese with a BMI between 25 kg/m2 and 30 kg/m2. as far as we know. However, diets rich in polyphenols and
This study has two limitations. One is the lack of anthocyanins have been shown to modulate the activity
consideration of hypertension risk and gender effects. and/or expression of the high-density lipoprotein-related
Although abdominal subcutaneous fat area has no formal enzyme paraoxonase 1 (PON1) [66–69]. Another study

reference value, increased subcutaneous fat increases determined the role of PON1 as a susceptibility gene to

the risk of the occurrence of diabetes and metabolic high antioxidant intake on cardiovascular health and

syndrome [57–59], and subcutaneous and visceral fat are identified four independent polymorphic protective

important risk factors of obesity-associated diseases, genotypes that were significantly associated with

including hypertension [60]. In fact, the risk of developing elevated high-density lipoprotein levels under high
polyphenol and anthocyanin intake [70]. In a study that
hypertension among obese individuals has increased by
investigated the effects of fucoxanthin (a type of
1.5-fold from 1980 to 2010 [61]. Thus, the risk of
carotenoid that is not a polyphenol) on blood glucose
hypertension should be considered in obesity
levels in Japanese people, those with the G/G allele of
management. In a study of Japanese patients (1,046
UCP1 had significantly lower blood glucose levels than
males and 853 females) attempting to determine cutoff
those with A/A and A/G alleles [71]. In these studies,
values for abdominal subcutaneous fat area based on
genetic polymorphisms can cause different high-density
hypertension prevalence, cutoff values of 114.7 and
lipoprotein levels and different responses to blood
169.3 cm2 for subcutaneous fat area for males and
glucose. Thus, the degree of visceral fat burning by
females, respectively, were suggested [62]. Sex
consuming Acacia bark-derived proanthocyanidins can
differences have been described in body fat
also vary in this study. However, genetic polymorphisms,
accumulation and reduction, with men and women more including UCP1 and β3 Adrenoceptor (ADRB3), which are
likely to accumulate visceral and subcutaneous fat, involved in fat burning, were not included in this study.
respectively [63]. In particular, visceral fat is more easily Thus, it remains unclear which gene polymorphisms were
reduced than subcutaneous fat in both sexes, with men actually incorporated and strongly affected by Acacia
having a higher fat resolution than women [64]. bark-derived proanthocyanidins. Future research should
Accordingly, one of our future tasks is to verify the anti- investigate genetic polymorphisms to determine the
obesity effect of Acacia bark-derived proanthocyanidins attributes of individuals who strongly display the visceral
after considering the effects of abdominal subcutaneous fat–reducing effects of Acacia bark-derived

fat area and sex, particularly in light of the cutoff value proanthocyanidins.

calculated based on hypertension risk.

Another limitation is that we did not take into CONCLUSIONS

account the genetic background. Differences in visceral This study evaluated the effect of Acacia bark-derived

fat mass have been observed depending on the genetic proanthocyanidins on fat accumulation among healthy

polymorphism of Uncoupling protein 1 (UCP1; Japanese adults with a BMI of 25–30 kg/m2 and a large

[−3826A/G]), which involves the burning of visceral fat visceral fat area. The abdominal visceral fat area at 12
Functional Foods in Health and Disease 2024; 14(12): 947-967 FFHD Page 964 of 967

weeks was significantly reduced in the Acacia group than the study participants and measurement staff for their
in the Placebo group. Therefore, the consumption of cooperation in this study.
Acacia bark-derived proanthocyanidins at 375 mg/day
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