Calculus bovis inhibits M2 tumor-associated macrophage polarization via Wnt/β-catenin pathway modulation to suppress liver cancer

Table 1 Primers used for qPCR in this study

Gene name	Forward (5′ to 3′)	Reverse (5′ to 3′)	Product Length
WNT5B	TCTTCATCCTCCACGGTT	CAGTTTAGGGCTTTCCTGAC	84 bp
β-catenin	CAACTAAACAGGAAGGGATGGAAGG	CAGATGACGAAGAGCACAGATGG	239 bp
AXIN2	TACACTCCTTATTGGGCGATCA	TTGGCTACTCGTAAAGTTTTGGT	151 bp
CCL22	ATCGCCTACAGACTGCACTC	GACGGTAACGGACGTAATCAC	129 bp
TGF-β	GCAGGTATTGATGGCACCTCC	GGCATGCTCCAGCACAGAAG	301 bp
Arg-1	CTTGGCAAAAGACTTATCCTTAG	ATGACATGGACACATAGTACCTTTC	170 bp
IL-10	TTTAAGGGTTACCTGGGTTGC	TTGATGTCTGGGTCTTGGTTC	98 bp
GAPDH	GCTGAGAACGGGAAGCTTGT	GCCAGGGGTGCTAAGCAG	299 bp

Table 2 Information analysis of 22 compounds in Calculus bovis

Number	Retention time (min)	Molecular weight	Molecular formula	Name	Serum
1	11.362	222.0905	C13H10N4	1,5-Diphenyltetrazole	√
2	13.144	374.1593	C17H26O9	Deoxyloganin
3	14.277	352.1787	C21H24N2O3	Gelsevirine	√
4	16.133	678.5017	C40H70O8	Campechic acid A
5	16.381	190.1205	C6H13NO6	D-Glucosaminic Acid	√
6	18.253	221.0953	C14H11N3	Isoquinoline
7	21.631	515.2957	C26H45NO7S	Taurocholic acid
8	22.525	408.2876	C24H40O5	Cholic acid	√
9	23.039	584.2634	C33H36N4O6	Bilirubin	√
10	23.716	465.309	C26H43NO6	Glycocholic acid	√
11	24.844	499.2968	C26H45NO6S	Taurodeoxycholic acid	√
12	25.939	256.2402	C16H32O2	Palmitic acid
8	26.684	408.2876	C24H40O5	Cholic acid
13	27.776	449.3141	C26H43NO5	Glycohyodeoxycholic acid	√
14	28.919	392.2927	C24H40O4	Hyodeoxycholic acid	√
15	29.765	390.277	C24H38O4	7-Ketolithocholic acid
16	31.405	356.2715	C24H40O4	Anoxycholic acid
17	32.051	784.5853	C24H40O4	Deoxycholic acid
18	33.541	222.162	C14H22O2	Isobornyl methacrylate
19	37.532	422.3032	C25H42O5	Methyl cholate
20	37.556	280.2402	C24H40O3	Lithocholic acid	√
21	37.701	148.016	C8H4O3	Isobenzofuran-1,3-dione
22	38.046	304.2402	C20H32O2	Arachidonic acid	√

Table 3 Quantitative analysis of active components in Calculus bovis extract

No.	Component	Concentration of standard	Peak area of standard	Peak area of sample	Concentration of sample (mM)
1	Cholic acid	1 mg/mL	303383832.9	111239787.3	0.366663531
2	Glycohyodeoxycholic acid	1 mg/mL	263903105.5	151518620.6	0.574144894
3	Anoxycholic acid	1 mg/mL	255212724.5	84507042.75	0.331123939
4	Deoxycholic acid	1 mg/mL	303655598.9	116148310.7	0.382500145
5	Lithocholic acid	1 mg/mL	377852336.8	133498724.1	0.706618491

Table 4 Quantitative analysis of active components in Calculus bovis-enriched serum

No.	Component	Concentration of standard	Peak area of standard	Peak area of sample	Concentration of sample (mM)
1	Cholic acid	1 mg/mL	18294415.89	4577619.85	0.250219514
2	Glycohyodeoxycholic acid	1 mg/mL	14604541.97	4029834.55	0.275930225
3	Lithocholic acid	1 mg/mL	15655005.77	175178.8	0.022379909