Altered amino acids and acylcarnitines profile in infantile intrahepatic cholestasis liver disease: implication for Traditional Chinese Medicine syndrome differentiation

doi:10.19852/j.cnki.jtcm.2026.04.019

Abstract

Abstract:

OBJECTIVE: To explore the diagnostic efficacy of blood amino acids (AAs) and acylcarnitines (ACs) profiles as metabolic biomarkers for differentiating Traditional Chinese Medicine (TCM) syndromes in infantile intrahepatic cholestatic liver disease (IICLD).

METHODS: Blood samples were collected from a cohort comprising 50 healthy controls and 77 patients diagnosed with IICLD, including 43 cases categorized as cold-damp obstruction and stagnation syndrome (COS) and 34 cases as jaundice due to stasis and accumulation syndrome (JSA) based on TCM diagnostic criteria. Metabolomic analyses targeting AAs and ACs were performed using liquid chromatography-tandem mass spectrometry (LC-MS/MS).

RESULTS: Compared with the healthy controls group, the IICLD group showed 23 upregulated and 2 downregulated metabolites with a screening threshold of variables of importance in projection (VIP) ≥ 1, fold change (FC) ≥ 1.2 or ≤ 0.83, and P <0.05. Six different metabolites were screened between the COS group and JSA group with a screening threshold of VIP ≥ 1, FC ≥ 1.2 or ≤ 0.83, and P <0.05. Ornithine (Orn), Decanedioy lcarnitine (C10DC), Caproylcarnitine (C6) were identified as potential biomarkers for distinguishing between the two syndromes with VIP > 2, FC ≥ 1.2 or ≤ 0.83 and P <0.05; The areas under the receiver operating characteristic curves (AUC) for these biomarkers were Orn (AUC = 0.7394), C10DC (AUC = 0.7777), and C6 (AUC = 0.686). When the levels of these three biomarkers were combined, the AUC increased to 0.922.

CONCLUSION: Distinct alteration of AAs and ACs profile highlighted the oxidative stress and mitochondrial impairment in IICLD. A combined set of metabolites (Orn+ C10DC+ C6) emerged as valuable biomarker for differentiating between the COS syndrome and JSA syndromes, providing a biochemical basis for syndrome differentiation.

Key words: infant, cholestasis, metabolomics, amino acids, carnitine acyltransferases

QIU Yanyan, XIA Yu, LI Weiwei, YAN Suqi, PANG Yusheng. Altered amino acids and acylcarnitines profile in infantile intrahepatic cholestasis liver disease: implication for Traditional Chinese Medicine syndrome differentiation[J]. Journal of Traditional Chinese Medicine, 2026, 46(4): 1000-1008.

Figures/Tables 5

Table 1 Demographic and clinical characteristics of the study cohort ($\bar{x}±s$)

Characteristic	HCs (n = 50)	IICLD (n = 77)	COS (n = 43)	JSA (n = 34)
Gender (M/F, n)	31/19	47/30	27/16	20/14
Age (months)	2.5±0.9	2.3±0.8	2.4±0.7	2.6±0.8
Body weight (kg)	5.0±1.0	4.9±1.0	4.9±1.0	4.9±1.1
Mode of delivery (ND/CS, n)	23/27	30/47	18/25	12/22
TBIL (μmol/L)	10.8±4.4	216.1±51.4^a	204.5±48.9	230.7±51.5^b
DBIL (μmol/L)	3.8±2.1	133.4±35.8^a	126.4±32.6	142.1±38.2^b
ALT (U/L)	34.6±15.0	255.9±187.7^a	202.4±124.6	323.4±230.1^b
AST (U/L)	25.4±8.1	320.6±237.2^a	255.6±177.2	402.8±277.8^b
TBA (μmol/L)	5.7±2.8	165.7±71.1^a	145.7±47.2	190.9±87.3^b
GGT (U/L)	24.4±14.8	225.6±219.1^a	155.5±111.4	314.3±283.3^b
ALP (U/L)	243.8±68.0	478.6±192.5^a	444.4±155.6	521.9±225.9^b

Figure 1 Amino acid- and acylcarnitine-targeted metabolomics profiling analysis in healthy controls group and infant intrahepatic cholestasis liver disease group A: PLS-DA score plots; B: validation plots obtained from 200 permutation tests; C: volcano map of 25 differential amino acid and acylcarnitine screening with a screening threshold of variables of importance in projection ≥ 1, FC ≥ 1.2 or ≤ 0.83, and P < 0.05. HCs: healthy controls (n = 50); IICLD: infantile intrahepatic cholestatic liver disease (n = 77). FC: fold change; PLS-DA: partial least squares discriminant analysis; PC: principal component; C0: free carnitine; C2: acetylcarnitine; C3: propionylcarnitine; Thr: threonine; C16DC: hexadecanedioylcarnitine; C18-OH: 3-hydroxysteroylcarnitine; C14: myristoylcarnitine; C16: palmitoylcarnitine; C16-OH: 3-hydroxy-hexadecanoylcarnitine; C12-DC: dodecanedioylcarnitine; C8DC: suberylcarnitine; C18:2: octadecadienylcarnitine; C18DC: octadecanedioylcarnitine; C14-DC: tetracanedioylcarnitine; C16:1: hexadecenoylcarnitine; C16:1-OH: 3-hydroxy-hexadecenoylcarnitine; C18:1-OH: 3-hydroxyoleylcarnitine; C8:2: octadienoylcarnitine; C8:1: octenoylcarnitine; C6-OH: 3-hydroxyhexanoylcarnitine; Orn: ornithine; Ser: serine; Met: methionine. Statistical analyses were measured by using 2-tailed unpaired Student’s t-test.

Figure 2 Amino acids- and acylcarnitines-targeted metabolomics profiling analysis in COS group and JSA group A: PLS-DA score plots; B: validation plots were obtained from 200 permutation tests; C: variable importance in projection analysis. The green and red boxes on the right indicate whether metabolite level is increased (red) or decreased (green). COS: cold-damp obstruction and stagnation syndrome (n = 43); JSA: jaundice due to stasis and accumulation syndrome (n = 34). PLS-DA: partial least squares discriminant analysis; PC: principal component; C10DC: decanedioylcarnitine; Orn: ornithine; C6: caproylcarnitine; His: histidine; Glu: glutamic acid; Trp: tryptophan; C18-OH: 3-hydroxysteroylcarnitine; C18:1-OH: 3-hydroxyoleylcarnitine; Cit: citrulline; C2: acetylcarnitine; Gly: glycine; C5DC: glutarylcarnitine; C14-DC: tetracanedioylcarnitine; C3: propionylcarnitine; Arg: arginine; Ala: alanine; Leu: leucine; Met: methionine; C18:1: octadecenoylcarnitine; C3DC: malonylcarnitine; VIP: variables of importance in projection.

Figure 3 Differential metabolites analysis between COS group and JSA group A: volcano map of six differential amino acids and acylcarnitines with a screening threshold of VIP ≥ 1, FC ≥ 1.2 or ≤ 0.83, and P < 0.05; B: comparison of differential metabolites between HCs group, COS group and JSA group; B1: C6; B2: C10DC; B3: C18:1-OH; B4: C18-OH; B5: His; B6: Orn. HCs: healthy controls (n = 50); COS: cold-damp obstruction and stagnation syndrome (n = 43); JSA: jaundice due to stasis and accumulation syndrome (n = 34). Orn: ornithine; His: histidine; C6: caproylcarnitine; C10DC: decanedioylcarnitine; C18-OH: 3-hydroxysteroylcarnitine; C18:1-OH: 3-hydroxyoleylcarnitine. Data are presented as the median (P25, P75). Statistical analyses were measured by using 2-tailed unpaired Student’s t-test. aP < 0.1, bP < 0.01 vs HCs group.

Figure 4 ROC curve based on potential biomarkers and biomedical indexes A: ROC curves for Orn, C10DC and C6, respectively; B: ROC curves for Orn + C10DC + C6; C: ROC curves for TBIL, DBIL, ALT, AST, TBA, GGT and ALP, respectively; D: ROC curves for TBIL + DBIL + ALT + AST + TBA + GGT + ALP. ROC: receiver operating characteristic; AUC: areas under the receiver operating characteristic curves; Orn: ornithine; C10DC: decanedioylcarnitine; C6: caproylcarnitine; TBIL: total bilirubin; DBIL: direct bilirubin; ALT: alanine aminotransferase; AST: aspartate aminotransferase; TBA: total bile acid; GGT: γ-glutamyl transpeptidase; ALP: alkaline phosphatase.

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