Possible mechanisms associated with immune escape and apoptosis on anti-hepatocellular carcinoma effect of Mu Ji Fang granules

Table 1 Four herbs in Mu Ji Fang granules

Herb	Latin name	Family and genus	Plant	Medicinal parts
Sophorae tonkinensis Radix	Sophorae tonkinensis Radix et rhizoma	Leguminosae	Sophora tonkinensis Gapnep	Root
Cuscutae semen	Cuscutae semen	Convolvulaceae	Cuscuta australis R. Br. or Cuscuta chinensis Lam	Seed
Juglans mandshurica Maxim	Juglans mandshurica Maxim	Juglandaceae	Juglans mandshurica	Bark
Coriolus versicolor	Coriolus versicolor	Basidiomycetes	Coriolus versicolor (L. ex Fr.) Quel	Fruit body

Table 2 Primer sequences for Real Time Quantitative PCR of tumor tissue

Name of primers	Sequences
Bax-F	5’-AGGATGCGTCCACCAAGAA-3’
Bax-R	5’-CAAAGTAGAAGAGGGCAACCAC-3’
Bcl-2-F	5’-CAACACTCCCTCTTGACCTATGC-3’
Bcl-2-R	5’-GAAAATGTTCCCAAGTGAGTTAGA-3’
β-actin-F	5’-GTCCCTCACCCTCCCAAAAG-3’
β-actin-R	5’-GCTGCCTCAACACCTCAACCC-3’

Table 3 Primer sequences for Real Time Quantitative PCR of HepG2 cells

Name of primers	Sequences
TNF-α-F	5’-ATCGGTCCCAACAAGGAGGAGAAGT-3’
TNF-α-R	5’-ACGTGGGCTACGGGCTTGTCACTC-3’
IFN-γ-F	5’-ACAACCAGGCCATCAGCAACAACATA-3’
IFN-γ-R	5’-CTGTGGGTTGTTCAGCTCGAACTT-3’
Bax-F	5’-AAACTGGTGCTCAAGGCCC-3’
Bax-R	5’-AAAGTAGGAGAGGAGGCCGTC-3’
Bcl-2-F	5’-CAGGATAACGGAGGCTGGGATG-3’
Bcl-2-R	5’-TTCACTTGTGGCCCAGATAGG-3’
β-actin-F	5’-GGACCTGACTGACTACCTC-3’
β-actin-R	5’-TACTCCTGCTTGCTGAT-3’

TNF-α: Tumor necrosis factor α; INF-γ: Interferon gamma.

Table 4 Compound identification in Mu Ji Fang Granules by High Performance Liquid Chromatography-Electron Spray Ionization-Time of Flight- Mass Spectrometry with reference substances

No.	RT (min)	[M-H/M+FA]-	Actual-M	Formula	Proposed compound
1	3.412	189.1291	189.1290	C11H14N2O	Cytisine
2	3.585	169.0149	169.0145	C7H6O5	Gallic acid
3	3.607	249.2185	249.2179	C15H24N2O	Matrine
4	3.615	247.2023	247.2020	C15H22N2O	Sophocarpine
5	4.997	153.0922	153.0914	C7H6O4	Protocatechuic acid
6	7.109	178.0506	179.0349	C15H24N2O2	Caffeate
7	9.003	463.0878	463.0873	C21H20O12	Hyperoside
8	9.271	303.0512	303.0508	C15H10O7	Quercetin
9	9.832	464.3002	464.2997	C21H20O12	Isoquercitrin
10	10.183	610.3217	610.3213	C27H30O16	Rutin
11	10.452	594.2133	594.2131	C27H35O5	Kaempferol-3-O-rutinoside
12	10.793	476.3361	476.3364	C21H20O11	Astragalin
13	12.033	271.0689	271.0681	C15H12O5	Naringenin
14	15.525	288.3132	288.3128	C15H10O6	Kaempferol
15	17.542	302.3295	302.3287	C14H6O8	Ellagic acid
16	18.413	194.0819	194.0823	C10H10O4	Ferulic acid
17	20.414	318.3255	318.3251	C16H12O7	Isorhamnetin

RT: Real time.