Altered profiles of fecal metabolites correlate with visceral hypersensitivity and may contribute to symptom severity of diarrhea-predominant irritable bowel syndrome

Table 1 Demographics, clinical characteristics and psychological states of study subjects

Features	IBS-D patients	Controls	P value
n	30	15	NA
Age (yr)	30.0 (28.0, 41.3)	28.0 (25.0, 37.0)	0.299
Gender (male:female)	22:8	11:4	1.000
Body mass index (kg/m2)	23.8 ± 3.9	22.4 ± 2.8	0.219
Duration of disease (yr)	7.5 (1.5, 10.1)	NA	NA
IBS-SSS	261.7 ± 40.4	NA	NA
Severity of abdominal pain	1.0 (1.0, 2.0)b	0.0 (0.0, 0.0)	< 0.001
Frequency of abdominal pain	2.0 (1.0, 4.0)b	0.0 (0.0, 0.0)	< 0.001
BSFS score	6.0 (6.0,6.0)b	4.0 (4.0, 4.0)	< 0.001
HADS anxiety score	6.0 (3.0, 9.0)b	2.0 (1.0, 4.0)	0.002
HADS depression score	4.0 (1.0, 7.3)	2.0 (2.0, 5.0)	0.350
VSI score	34.0 (17.0, 45.3)b	5.0 (2.0, 15.0)	< 0.001

^bP < 0.01 vs controls. The data are presented as the mean ± SD or the median (Q1, Q3). IBS-D: Irritable bowel syndrome with predominant diarrhea; IBS-SSS: IBS symptom severity scale; BSFS: Bristol stool form scale; HADS: Hospital anxiety and depression scale; VSI: Visceral sensitivity index; NA: Not applicable.

Table 2 Levels of partial fecal metabolites in irritable bowel syndrome with predominant diarrhea patients and controls

Metabolites	Prevalence of detection	IBS-D patients (μmol/g)	Controls (μmol/g)	P value
His	93.3%	0.0642 (0.0388, 0.1484)a	0.2636 (0.0780, 0.3966)	0.012
Ala	100.0%	0.5095 (0.2826, 0.9183)a	1.0118 (0.6135, 1.4335)	0.041
Tyr	88.9%	0.1024 (0.0173, 0.4527)a	0.5665 (0.2436, 1.3447)	0.018
Phe	97.8%	0.1511 (0.0775, 0.3248)a	0.3967 (0.1388, 0.7550)	0.028
Trp	77.8%	0.0323 (0.0001, 0.0826)a	0.0834 (0.0170, 0.1759)	0.046
Val	100.0%	0.2710 (0.1208, 0.5729)	0.3661 (0.2960, 1.0122)	0.079
Asp	97.8%	0.3086 (0.1797, 0.6904)	0.4875 (0.1939, 0.6591)	0.647
Fumarate	100.0%	0.0042 (0.0026, 0.0108)	0.0019 (0.0014, 0.0078)	0.051
Pyruvate	100.0%	0.0067 (0.0023, 0.0719)	0.0071 (0.0022, 0.0882)	0.718
Butyrate	100.0%	4.3819 (2.4060, 9.0920)	5.8539 (1.5193, 12.8616)	0.613
Isobutyrate	100.0%	6.8609 (0.5068, 8.9326)	2.1277 (0.5470, 10.4635)	0.962
Valerate	100.0%	0.2024 (0.0331, 0.8245)	0.2059 (0.0497, 1.2489)	0.485
Isovalerate	100.0%	0.1218 (0.0495, 0.3227)	0.1661 (0.0416, 0.2581)	0.866
Hexanoate	100.0%	0.0254 (0.0189, 0.0513)	0.0383 (0.0153, 0.0539)	0.613
Isohexanoate	100.0%	0.0127 (0.0060, 0.0246)a	0.0070 (0.0023, 0.0106)	0.028
Amino acids	NA	7.2873 (3.4177, 10.7709)	8.0603 (6.1393, 12.3225)	0.336
SCFA	NA	25.8082 (18.5826, 38.5067)	26.8452 (5.7182, 70.6763)	0.904
Organic acids	NA	26.6708 (20.0617, 38.5860)	26.9449 (5.8047, 71.4057)	0.866

^aP < 0.05 vs controls. The data are presented as the median (Q1, Q3). IBS-D: Irritable bowel syndrome with predominant diarrhea; SCFA: Short chain fatty acids; NA: Not applicable.

Table 3 Correlation of bristol stool form scale scores with the levels of His, Ala, Tyr, Phe, or Trp

Metabolites	IBS-D		Controls		All subjects
	r	P value	r	P value	r	P value
His	0.057	0.764	0.423	0.117	-0.279	0.064
Ala	-0.026	0.892	0.338	0.218	-0.252	0.095
Tyr	-0.208	0.271	0.423	0.117	-0.3471	0.019
Phe	0.002	0.994	0.465	0.081	-0.250	0.098
Trp	-0.114	0.548	0.423	0.117	-0.268	0.075

¹There was association between parameters (P < 0.05). IBS-D: Irritable bowel syndrome with predominant diarrhea.

Table 4 Multiple linear regression model for irritable bowel syndrome symptom severity scale score

Metabolites	Unstandardized coefficients		Standard-ized coefficie-nts β	t	P value	Collinearity statistics
	b	SE				Tolerance	VIF
Hexanoate1	116.194	29.961	0.420	3.878	0.001	0.903	1.107
Fumarate1	-1733.224	490.683	-0.383	-3.532	0.002	0.902	1.108
Pyruvate1	140.381	28.619	0.528	4.905	< 0.001	0.915	1.092
Asp	-15.303	7.943	-0.221	-1.927	0.066	0.807	1.240
Tyr1	60.960	15.019	0.763	4.059	< 0.001	0.300	3.333
Val1	-82.922	26.139	-0.626	-3.172	0.004	0.272	3.671

¹Metabolite which was correlated with irritable bowel syndrome symptom severity scale score (P< 0.01); SE: Standard error; VIF: Variance inflation factor.

Table 5 Collinearity diagnostics of models

Models	Dimensions	Collinearity diagnostics
		Eigenvalues	Condition indices
Multiple linear regression model for IBS-SSS score in IBS-D group	1	3.897	1.000
	2	1.008	1.966
	3	0.751	2.278
	4	0.643	2.462
	5	0.407	3.093
	6	0.222	4.186
	7	0.071	7.394
Multiple linear regression model for maximum tolerable threshold in IBS-D group	1	2.286	1.000
	2	0.675	1.841
	3	0.040	7.580

IBS-D: Irritable bowel syndrome with predominant diarrhea; IBS-SSS: Irritable bowel syndrome symptom severity scale score.

Table 6 Multiple linear regression model for maximum tolerable threshold

Groups	Metabolites	Unstandardized coefficients		Standar-dized coeffici-ents β	t	P value	Collinearity statistics
		b	SE				Tolerance	VIF
IBS-D	Isovalerate2	5.451	1.824	1.525	2.989	0.006	0.099	10.132
	Valerate1	-2.171	0.946	-1.171	-2.295	0.030	0.099	10.132
All subjects	Isohexanoate1	-40.910	18.695	-0.317	-2.188	0.034	NA	NA

¹Metabolite which was correlated with maximum tolerable threshold (P < 0.05).

²Metabolite which was correlated with maximum tolerable threshold (P < 0.01); SE: Standard error; VIF: Variance inflation factor; IBS-D: Irritable bowel syndrome with predominant diarrhea; NA: Not applicable.

Table 7 Correlation of bowel disease questionnaire scores with isovalerate or isohexanoate

Groups	Metabolites	Frequency of abdominal pain		Severity of abdominal pain
		r	P value	r	P value
IBS-D	Isovalerate	0.4752	0.008	0.3771	0.040
	Isohexanoate	0.4141	0.023	0.097	0.612
All subjects	Isovalerate	0.200	0.188	0.143	0.350
	Isohexanoate	0.4732	0.001	0.3711	0.012

¹There was association between parameters (P < 0.05).

²There was association between parameters (P < 0.01). IBS-D: Irritable bowel syndrome with predominant diarrhea.