Butyrate Acid as a Potential Marker for Diversity of Gut Microbiota

in Colorectal Cancer Patients

Fauzi Yusuf

, Azzaki Abubakar

, Desi Maghfirah

and Siti Adewiah

Division of Gastroenterology-Hepatology, Department of Internal Medicine, Faculty of Medicine University of Syiah

Kuala/ Dr. Zainoel Abidin Centre Hospital, Banda Aceh, Indonesia.

Department of Internal Medicine, Faculty of Medicine, Syiah Kuala University, Banda Aceh, Indonesia.

Keywords: Colorectal Cancer, Butyrate Acids, Biomarker

Abstract: The gut microbiota acts as a real organ and many changes in its composition have been reported in

colorectal cancer (CRC). Short-chain fatty acids (SCFA) mainly produced as microbial metabolites, acetate,

propionate, and butyrate acids. Butyrate is produced by specific bacteria, mainly in the colon, and is taken

up by the host. In our study, we found that CRC patients had lower level of acetate, propionate and butyrate

acids than non-CRC. The mean concentration of acetate 8,55 µg/mL, propionate 5,61 µg/mL and butyrate

acids 3,79 µg/mL respectively. In three of SCFA, the level of butyrate acids had the best diagnostic

properties with area under receiver operating characteristic (ROC) curve of 0.84 higher than acetate (0.71)

and propionate (0.75) (p< 0.05).



1 INTRODUCTION

Colorectal cancer is the fourth most cancer evident

worldwide (Iffrig and Weinber, 2009). The rate of

colorectal cancer is 5-10 times higher in the most

developed country (Ginsberg et al, 2010). In

Indonesia, colorectal cancer disease is caused the

increasing evidences of cancer-related mortality in

recent years Based on epedimiological report during

1996-1999 from Pathology Anatomy Division of

Medical Faculty, Indonesia University, wrote the

colorectal cancer patients with age under 40 years

are around 36.75% ( Sudoyo et al, 2010). Colorectal

cancer is a disease from an accumulation of genetic

mutation, epigenetic, disregulate in their

communication in signaling pathways, and gut

microbial contribution. Microbial involvement in

colorectal cancer (CRC) is now well established

(Sekirov et al, 2010).

SCFA are the main products of anaerobic

microbial fermentation in the large intestine and

affect colonic health. SCFA mainly produced as

microbial metabolites, acetate, propionate, and

butyrate acids. The formation of butyrate and other

SCFA possibly playing a major role as

chemopreventive products of microbial fermentation

in the colon (Faujan et al, 2010; Topping and

Clifton, 2001). Butyrate production is a major source

of energy for colonic epithelial cells and affecting to

the protection from colitis and colorectal cancer (

Rose et al, 2007; Vinolo et al, 2011). The relation

between functional characteristics, such as SCFA

especially butyrate acid and CRC has not been

extensively investigated. In one study showed

butyric acid was significantly higher in the feces of

healthy subject than CRC ( Simpson and Campbell,

2015). In this study, we evaluated the level of

butyrate acid as diagnostic biomarkers for diversity

of gut microbiota in colorectal cancer patients.

2 PATIENTS AND METHODS

2.1 Participants and Sample Collection

The study consists of fourteen subjects with CRC

and 14 non-CRC were from in the Gastroenterology-

Hepatology Department at Dr. Zainoel Abidin

General Teaching Hospital Banda Aceh, Indonesia.

The patients were selected based on the following

inculsion criteria :1) Patients aged 18 years or over ;

2) Indonesian citizen that proved by identity cards;

3) Patients wtih colorectal cancer confirmed by

pathological examination; 4) Patients instead of

CRC; and 5) Patients who are able to cooperate in

Yusuf, F., Abubakar, A., Maghﬁrah, D. and Adewiyah, S.

Butyrate Acid as a Potential Marker for Diversity of Gut Microbiota in Colorectal Cancer Patients.

DOI: 10.5220/0008790200270030

In Proceedings of the 2nd Syiah Kuala International Conference on Medicine and Health Sciences (SKIC-MHS 2018), pages 27-30

ISBN: 978-989-758-438-1

the study. None of the patients had active antibiotic

treatment or within the month prior to the

colonoscopy, youghurt consumtion or laxative

medicine for the last five weeks and none were

treated with chemotherapy and/or radiotherapy in

the previous six months. One stool samples was

collected from each participant, where stool samples

will be labelled and stored at -20ºC freezer service

by the researchers. The study was approved by the

Ethical Review Committee of Medical Faculty,

Syiah Kuala University, Banda Aceh, Indonesia.

2.2 Gas Chromatography Analysis of

Fecal SCFA Concentration

Stool samples were analysed for SCFA

concentration with gas chromatography (GC) as

described from a previous methods (Chen et al,

2013; Tangerman and Nagengast, 1996]. The

amounts of acetate, propionate and butyrate acids

have been reported as µg/ml and %.

2.3 Statistical Analysis

The exact chi-square test and Student t-test were

used for the comparisons between the groups, and

nonparametric statistics was used in addition for

variables without normal distribution. The

diagnostic properties were assessed with Receiver

Operating Characteristics (ROC) curves, sensitivity,

specificity, positive- and negative Likelihood Ratio

(LR) and Diagnostic Odds Ratio (DOR). P-values

below 0.05 were judged as statistically significant

3 RESULTS

3.1 Participants

Total 28 participants were included in this study.

Table 1 shows the characteristics of participants.

There were 10 males and 4 females in the CRC

group, and 9 males and 5 female in non-CRC group.

The means (±SD) was 53.8± 13.3 years for CRC and

50.0 ± 17.6 years for non-CRC. Haemoglobin, BMI

and albumin in the CRC group was lower than non-

CRC group. Percentage of cancers based on

location: rectum 79% and colon descending 21%.

Table 1 Characteristic of participants in this study

Variable Colorectal

cancer

(% or SD)

Non – colorectal

cancer

(% or SD)

Gender

Male

Female

10 (72%)

4 (28%)

9 (64%)

5 (36%)

Age (years, mean)

53.8 ± 13.3 50.0 ± 17.6

BMI (kg/m

mean)

20.21 ± 2.65 23.6 ± 1.91

Hemoglobin (g/dl,

mean)

10.6 ± 2.1 12.3 ± 1.2

Albumin (g/dl,

mean)

3.24 ± 0.71 3.91 ±0.53

Colonoscopy



Ca Rectum

11 (79%)

Ca Colon

Descending

3 (21%)

Colitis Infection 10 (72%)

ColitisInfection

+ Hemorrhoid

Interna

4 (28%)

The mean faecal concentrations of acetate,

propionate dan butyrate were significanty lower in

patients with CRC compared non-CRC. From the

table 2, results revealed that the mean concentration

of acetate 8,55 µg/mL, propionate 5,61 µg/mL and

butyrate acids 3,79 µg/mL respectively (all P <

0.05).

Table 2. Fecal short-chain fatty acids in subjects with and

without colorectal cancer.

Variable

Colorectal

cancer

patients

(N=14)

Non-colorectal

cancer patients

(N=14)

P-

value

Acetate

Acids

8.55 ± 3.06 11.78 ± 4.61

0.038

Propionate

Acids

5.61 ± 1.95 8.61 ± 3.40

0.008

Butyrate

Acids

3.79 ± 2.04 6.81 ± 2.59

0.002

The results are given as mean values with SD.

3.2 Diagnostic Properties of SCFA

In three of SCFA, the level of butyrate acids had the

best diagnostic properties with area under receiver

operating characteristic (ROC) curve of 0.84 higher

than acetate (0.71) and propionate (0.75) (p< 0.05)

(Table 3, Figure 1).

SKIC-MHS 2018 - The 2nd Syiah Kuala International Conference on Medicine and Health Sciences

Table 3. Area under curve (AUC) short chain fatty acids.

AUC 95%CI

Acetate acids

Propionate acids

Butyrate acids

0.71

0.75

0.84

0.52 – 0.90

0.57 – 0.94

0.68 – 0.99

Butyrate acid value < 5.4 µg/mL was judged

as a well-suited cut-off for indicating CRC and was

used in the calculation of the diagnostic. According

to the reference range employed in this study, the

sensitivity, specificity, positive and negative

likelihood ratio, and diagnostic odds ratio were 85%,

78%, 4.04, 0.18 and 22.2 respectively.

Figure 1: The diagnostic properties of SCFA presented as

the Receiver Operating Characteristic curve.

4 DISCUSSION

With the increasing evidence linking gut microbiota

and CRC, fecal microbiota has emerged as a

promising candidate to non-invasively screen for

CRC. The present study demonstrated that the level

butyrate acids could be a valuable diagnostic

biomarker for CRC than the other SCFA.

Colorectal cancer (CRC) is a leading cause

of cancer-related mortality worldwide, and its

incidence has increased rapidly in recent years

(Jemal et al, 2011; Ji et al, 1998). In recent years,

the 16S rRNA gene sequencing approach has been

widely used as an effective tool to globally analyze

the microbial community, and multiple studies have

demonstrated that breakdown of the intestinal

microbiota structure can promote carcinogenesis and

development of CRC (Wei et al, 2016). Flanagan et

al, 2014 . demonstrated a significant association

between Fusobacterium nucleatum level and patient

outcome and suggested that F. nucleatum may have

value as a prognostic indicator. Boleij et al, 2015.

found that the detection of Bacteroides fragilis toxin

(BFT), which was produced by Enterotoxigenic

Bacteroides fragilis (ETBF), increased in the

mucosa of later staged CRC.Recently, the others of

our study suggest that the appearance of

Bifidobacterium as one of the indicators of

detections for colorectal cancer (Yusuf et al, 2016).

The microbiota metabolises non-digestible

food constituents into short-chain fatty acids (SCFA)

that have extensive immunological and regulatory

functions and appear to be the link in the host-

microbe interactions Within gut microbiota, several

distinct bacterial communities live at a certain ratio

under steady state condition ( Farup et al, 2016). The

level of SCFA content in faecal samples have been

shown to be related with some diseases such as IBD,

irritable bowel syndrome (IBS), cardiovascular

disease (CVD), diarrhoea, and cancer (Faujan et al,

2010). There is significant association between

levels of SCFAs and composition of the microbiota,

with high luminal concentrations resultant of

fermentation lowering colonic pH (5.5–6.5 in

proximal colon where fermentation is highest,

compared to pH 6.5–7.0 in the distal colon) and

inhibit growth of Gram-negative Enterobacteriaceae

including familiar pathogens Salmonella spp. and

Escherichia coli (Simpson and Campbell, 2015).

Therefore, a greater increase in SCFA production

and potentially a greater delivery of SCFA,

specifically butyrate, to the distal colon may result in

a protective effect (McOrist et al, 2011)

Fecal levels of butyrate in patients with

colonic neoplasia have been investigated by

different studies. Vernia et al, 1995. compared 20

patients with colorectal cancer, 8 patients colon

polyps, and healthy controls. No significant

differences were found, although patients with rectal

cancer showed slightly lower levels of propionate

and butyrate than those with more proximal cancer.

Contrast with the previous study, in our study we

found that butyrate was the best properties

diagnostic for CRC with AUC (0.84) higher than

acetate (0.71) and propionate (0.75).

The results of the present study are limited

by the relatively low number of participants and a

larger study population would provide enhanced

statistical reliability. In addition, we did not study

Butyrate Acid as a Potential Marker for Diversity of Gut Microbiota in Colorectal Cancer Patients

for identifies bacteria as far as we know, SCFA is

the main products of anaerobic microbial

fermentation. Another limitation is the fact that we

cannot eliminate environment factors such as diet

and everything related to microbiota.

5 CONCLUSIONS

In conclusion, this study was the first report

demonstrating of the level butyrate acids as useful

biomarkers to detect the presence of cancerous

lesions. Because the study had an exploratory design

and limited number of participants, these results

need for more validation study.

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