Effects of Lentinula Edodes Bar on Lipid Profile and Antioxidant Status in Borderline High Cholesterol Individuals: a Double Blind Randomized Clinical Trial
Phase: NA
Conditions: Dyslipidemias
Sponsor: University of Sorocaba

Effects of Lentinula edodes bar on lipid and antioxidant profiles in borderline 
high cholesterol individuals: A double blind randomized clinical trial 
 
 
NCT 04186780 
 
 
 
 
September 26th, 2018 
 
 

Protocol 
Trial design 
A prospective Phase II, simple randomization, double blind and placebo-
controlled trial was conducted from September to December 2018. The study was 
approved by Human Research Ethics Committee from University of Sorocaba (protocol 
number 2.824.297), in accordance with Resolution 466/2012 of the National Health 
Council and Research Ethics Review Committee (ERC). 
Eligibility Criteria 
The disclosure of the clinical study began in May with the formation of a team 
of collaborators. It was disseminated through folders distributed across all campuses 
where approximately 10,000 students study. The recruitment was also posted on the 
University website. Printed material was prepared for distribution so that students or 
family members could participate. The disclosure containing explanations about the 
research criteria, inclusion and exclusion happened in social networks aimed at 
borderline dyslipidemic individuals living in Sorocaba and cities in the region. 
For interested individuals, it was essential to answer an online questionnaire 
with multiple choice questions related to the inclusion and exclusion criteria, which 
defined the candidate's eligibility. A total of 165 individuals completed the Google 
online survey. 
- Inclusion Criteria 
Individuals aged from 20 to 65 years old, of both gender, who had at least one 
of the following biochemical markers at the borderline high level (total cholesterol 180 
to 239 mg/dL; LDL 130 to 159 mg/dL; triglycerides of 150 to 200 mg/dL), were 
recruited, diagnosed by biochemical exams with dates recent to the day of recruitment. 
The be tolerant to bars ingredients and to Shiitake. Availability to attend the date and 
time of blood collection. 
- Exclusion Criteria  
Some of the individuals had diseases such as cancer, heart disease, 
neurodegenerative disease, diabetes, among others. These diseases could be 
confounders of the study and some recruited people had to be excluded. Pregnant, 
lactating or hormone replacement women could not participate either. 
Individuals were instructed to no change their eating habits and patterns, 
physical activity level or oral contraceptive use during the study, thus identifying only 
the effects of Shiitake added to the diet for each individual on the exposed group.  

Study Interventions. 
The composition of the bars is presented in Table 1. Shiitake bar formulation 
was approved in the sensory analysis in a previous study from our working group. 
Shiitake concentration of 100 mg/Kg/day showed to be safe in a study conducted with 
different Shiitake concentrations intake (100, 400 and 800 mg/Kg/day).  
Table 1 – Ingredients used in the formulation sweet cereal bars in 25g. 
 
Ingredients 
Shiitake Bar 
Placebo Bar  
% 
g 
% 
g 
Dry Shiitake 
14 
3.50 
- 
- 
Flakes oat 
12 
3.0 
23.2 
5.80 
Brown sugar 
8.3 
2.10 
13.6 
3.40 
White sugar 
30 
7.50 
20 
5.00 
Dry plum 
9 
2.25 
10.8 
2.70 
Brazil nut 
7 
1.75 
8 
2.00 
Flaxseed 
3.2 
0.80 
3.44 
0.86 
Glucose 
1.6 
0.4 
1.6 
0.40 
Soy lecithin 
0.4 
0.10 
0.4 
0.10 
Quinoa grains 
3.5 
0.87 
4.8 
1.20 
Peanut 
8 
2.0 
10 
2.50 
Chia seed 
2 
0.5 
3.2 
0.80 
Food glycerin 
0.5 
0.12 
0.5 
0.12 
Coconut oil 
0.5 
0.12 
0.5 
0.12 
Total 
100 
25 
100 
25 
 
Outcomes 
The primary outcome of the study was total cholesterol, triglycerides, low-
density lipoproteins (LDL) or high-density lipoproteins (HDL) levels. The secondary 
outcomes included blood glucose, Body Mass Index (BMI) and oxidative stress 
biomarkers reduced glutathione (GSH), activity of the catalase enzyme and 
Thiobarbituric acid reactive substances (TBARS). 
Sample size 
A total of 165 individuals who completed the online questionnaire between May 
and September 2018, were eligible to participate in this study and were willing to 
participate and had some borderline dyslipidemia. From this number, individuals who 

did not meet the inclusion criteria were excluded. A 95% confidence interval was used 
for analysis. 
Randomization 
The individuals (68) were randomly allocated (using a table of random numbers) 
into 2 groups: I – Placebo group (n = 32) and II – Intervention group (n = 36).  
After nutritional assessment and blood collection, the Individuals received an 
unidentified opaque and sealed, containing bars individually vacuum packed. The bars 
lasted for a period of 33 days. The patients and the data collector were blinded. The 
bars of the intervention and placebo groups were similar in texture, flavor, aroma and 
appearance. 
After 33 days (T33), all individuals returned to the University of Sorocaba for 
new nutritional assessments. They received more food bars and had their blood 
collected with the same double-blind care. At the end of sixty-six days (T66), subjects 
returned for the last blood collection and final nutritional assessment. 
Blinding 
In the blind randomization, a University pharmaceutical and an undergraduate 
student unrelated to the research to keep full impartiality in the assessment contributed 
to the distribution of the intervention and placebo using sealed, opaque envelopes 
which contained specific markings for each group. The volunteers were unaware of 
which group they were participating in, as the envelopes apparently were the same, 
only the two external contributors were aware to the marks on the envelopes, because 
the bars with Shiitake and placebo inside the envelopes had the same size, 
appearance, aroma and taste.  
The researchers did not have access to the identification of participants in the 
groups, after the distribution of the envelopes, the participants were renamed by the 
blinded collaborators sequentially numbered from 1 to 68 until the completion of the 
study.