Targeted metabolomics study of fatty-acid metabolism in lean metabolic-associated fatty liver disease patients

Table 1 Ion pairs of analytes and internal standards

ID	Q1	Q3	DP	CE	CXP
AEA-d8	356.100	294.000	64.94	19.09	18.93
AEA	348.000	62.000	56.00	42.00	10.00
2-AG	379.000	287.400	163.86	20.05	13.09
OEA	326.400	309.200	195.32	21.31	29.01
OEA-d4	330.600	66.100	132.66	21.55	12.58
LEA	324.400	62.100	130.26	17.76	11.28
PEA	300.300	62.100	124.00	19.83	10.20
LEA-d4	328.100	66.000	133.80	20.98	8.59
1-LG	355.300	338.400	32.21	10.73	18.49
1-OG	357.300	339.300	145.00	13.20	24.00
1-PG	331.200	313.500	155.00	12.70	19.00
LA	325.200	279.100	-10.11	-8.72	-25.70
OA	327.200	281.300	-47.00	-10.50	-10.00
PA	301.200	255.200	-58.00	-14.00	-15.00
AA	303.100	259.000	-73.90	-16.14	-13.02
OA-d9	336.300	290.300	-14.34	-9.32	-7.94

1-LG: 1-linoleoyl glycerol; 1-OG: 1-oleoyl glycerol; 1-PG: 1-palmitoyl glycerol; 2-AG: 2-arachidonoylglycerol; AA: Arachidonic acid; AEA: Arachidonoylethanolamide; AEA-d8: Deuterated arachidonoylethanolamide; LA: Linoleic acid; LEA: Linoleoylethanolamide; LEA-d4: Deuterated linoleoylethanolamide; OA: Oleic acid; OA-d9: Deuterated oleic acid; OEA: Oleoylethanolamide; OEA-d4: Deuterated oleoylethanolamide; PA: Palmitic acid; PEA: Palmitoylethanolamide.

Table 2 General characteristics and clinical indicators of lean metabolic-associated fatty liver disease patients

Indicator	Lean MAFLD group, n = 20	Control group, n = 20	Statistical value	P value
Male/female sex	5/15	0/20	χ2 = 3.657	0.056
Age in years	52.75 ± 15.17	44.95 ± 12.11	t = 1.798	0.08
Weight in kg	60 ± 6.12	54.8 ± 5.64	t = 2.793	0.008b
BMI in kg/m²	22.65 (21.52, 22.80)	21.25 (20.05, 21.80)	Z = 3.196	0.001b
White blood cells as × 109/L	7.25 ± 2.08	5.31 ± 1.06	t = 3.715	0.001b
Red blood cells as × 1012/L	4.49 ± 0.63	4.23 ± 0.34	t = 1.616	0.117
Hemoglobin in g/L	134.20 ± 20.78	125.70 ± 13.58	t = 1.531	0.134
Hematocrit in %	39.22 ± 5.84	36.67 ± 3.27	t = 1.703	0.097
Platelets as × 109/L	261.25 ± 69.41	228.80 ± 45.42	t = 1.749	0.088
Mean platelet volume in fL	10.35 ± 1.21	10.96 ± 1.06	t = -1.693	0.099
Neutrophils in %	55.03 ± 11.49	51.53 ± 6.33	t = 1.192	0.243
Lymphocytes in %	35.01 ± 11.49	38.25 ± 6.39	t = -1.104	0.278
Monocytes in %	7.53 ± 1.8773	6.71 ± 0.93	t = 1.75	0.091
Mean red cell volume in fL	87.90 (84.5, 90.58)	87.85 (86.08, 89.95)	Z = -0.298	0.766
Mean red cell hemoglobin in pg	30.25 (28.63, 31.13)	30.55 (29.25, 31.28)	Z = -0.69	0.49
Mean red cell hemoglobin concentration in g/L	341.00 (337.00, 349.75)	346.00 (337.50, 351.00)	Z = -0.895	0.371
Red cell volume distribution width in %	12.90 (12.33, 13.63)	12.50 (12.13, 13.08)	Z = -1.178	0.239
Eosinophils in %	1.80 (0.75, 2.40)	1.95 (1.53, 3.33)	Z = -0.934	0.35
Basophils in %	0.50 (0.40, 0.70)	0.60 (0.40, 0.70)	Z = -0.849	0.396
Creatinine in μmol/L	66.25 ± 17.65	57.20 ± 6.99	t = 2.132	0.043a
Urea nitrogen in mmol/L	5.55 (4.35, 6.98)	4.65 (3.78, 5.38)	Z = -2.274	0.023a
Uric acid in μmol/L	350.50 (296.25, 450.00)	269.00 (236.75, 310.00)	Z = -3.044	0.002b
Total bilirubin in μmol/L	9.00 (11.00, 13.00)	9.00 (10.00, 11.00)	Z = -1.186	0.236
Direct bilirubin in μmol/L	1.66 (1.88, 2.30)	1.56 (1.62, 1.92)	Z = -1.556	0.120
Alkaline phosphatase in U/L	69.00 (59.25, 89.75)	61.50 (51.00, 82.50)	Z = -1.448	0.148
Cholinesterase in U/L	9214.95 ± 1232.29	7059.20 ± 1260.51	t = 5.469	0.001b
Alanine aminotransferase in U/L	17.00 (11.00, 35.75)	9.50 (6.25, 16.00)	Z = -3.022	0.003b
Aspartate aminotransferase in U/L	24.00 (20.00, 33.75)	20.50 (17.25, 26.75)	Z = -1.64	0.101
γ-glutamyl transferase in U/L	36.00 (21.50, 45.50)	20.50 (16.50, 34.75)	Z = -1.719	0.086
Total protein in g/L	73.05 ± 5.93	72.15 ± 4.04	t = 0.561	0.578
Albumin in μmol/L	42.10 ± 3.37	41.55 ± 2.98	t = 0.547	0.588
High-density lipoprotein in mmol/L	1.07 (0.98, 1.26)	1.40 (1.27, 1.59)	Z = -3.882	0.001b
Low-density lipoprotein in mmol/L	3.48 ± 0.72	3.40 ± 1.10	t = 0.255	0.800
Triglycerides in mmol/L	2.68 (1.37, 4.20)	1.09 (0.94, 1.39)	Z = -3.598	0.001b
Total cholesterol in mmol/L	5.19 ± 1.07	5.06 ± 1.28	t = 0.366	0.716
Apolipoprotein A1 in g/L	1.35 (1.22, 1.56)	1.55 (1.36, 1.74)	Z = -2.153	0.031a
Apolipoprotein B in g/L	0.99 ± 0.22	0.94 ± 0.32	t = 0.559	0.579
Glycated albumin in %	12.60 (10.90, 14.45)	13.00 (12.53, 14.18)	Z = -0.717	0.473
Fasting blood glucose in mmol/L	5.40 (5.03, 6.05)	4.90 (4.63, 5.15)	Z = -3.052	0.002b

^aP < 0.05, lean metabolic-associated fatty liver disease group vs control group.

^bP < 0.01, lean metabolic-associated fatty liver disease group vs control group.

MAFLD: Metabolic-associated fatty liver disease.

Table 3 Fatty-acid standard curve and linear range

No.	Name	Linear	Range in μg/mL	r
1	Arachidonic acid	y = 0.0087x - 0.0053	0.1-5	0.9963
2	Oleic acid	y = 0.0272x - 0.0425	0.1-5	0.9914
3	Palmitic acid	y = 0.0074x + 0.0036	0.1-5	0.9940
4	Linoleic acid	y = 0.0341x - 0.0821	0.1-5	0.9919

Table 4 Changes in fatty-acid content in lean metabolic-associated fatty liver disease patients, n = 6

No.	Name	Lean MAFLD patients	Normal individuals
1	Palmitic acid	3.41 ± 0.84a	1.53 ± 0.85
2	Oleic acid	2.63 ± 1.45a	1.26 ± 0.65
3	Linoleic acid	2.42 ± 1.18a	1.30 ± 0.49
4	Arachidonic acid	2.45 ± 1.21a	1.24 ± 0.56

^aP < 0.05.

Data are mean ± SD in μg/mL. MAFLD: Metabolic-associated fatty liver disease.