1H NMR Metabonomic Investigations of N-Benzylhexadecanamide Induced Proliferation and Iestosterone Secretion of Mouse Testicular Leydig Cells†

doi:10.7503/cjcu20190256

Abstract

Abstract:

A metabolomic mechanism was evaluated for inducing proliferation and testosterone secretion of mouse testicular Leydig cells by N-benzylhexadecanamide(NBH) from maca based on ¹H NMR metabolism approach. The cell proliferation rate of mouse testicular Leydig cell sbefore and after administration and the testosterone content in the cell culture medium was determined experimentally. Based on principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA), metabolite differences in mouse testicular Leydig cells before and after administration were studied, and metabolic pathway analysis was performed. The results showed that NBH could increase the proliferation rate and testosterone secretion of mouse testicular Leydig cells. After administration, the profile of the faulty liquid metabolic in mouse testicular Leydig cells was changed significantly, which identified 24distinct metabolites such as leucine, lysine, scyllo-inositol, valine and alanine. Metaboanalyst showed that distinct metabolites mainly involves 10 pathways including alanine, aspartate and glutamate metabolism, glycine, serine and threonine metabolism, glutathione metabolism, pyruvate metabolism, pantothenate and CoA biosynthesis, aminoacyl-tRNA biosynthesis, lysine degradation, Inositol phosphate metabolism, D-glutamine and D-glutamate metabolism as well as glycerophospholipid metabolism. The result indicated that NBH promotes proliferation and secretion of testosterone in mouse Leydig cells by organizing the relevant nodes of the above metabolic pathway.

Key words: Cell metabonomics, ¹H NMR, N-Benzylhexadecanamide, Mouse testicular Leydig cells

CLC Number:

O657.2

TrendMD:

ZHANG Nanxi, LV Jingwei, JIN Ping, LI Jingfeng, BIAN Xuefeng, ZHANG Hui, SUN Jiaming. ¹H NMR Metabonomic Investigations of N-Benzylhexadecanamide Induced Proliferation and Iestosterone Secretion of Mouse Testicular Leydig Cells^†[J]. Chem. J. Chinese Universities, 2019, 40(9): 1832.

Figures/Tables 6

References 31

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Group code	Concentration of NBH/(μg·mL_-1)	Cell viablility rate(%)	Testosteron secretion/(nmol·L_-1)
Control	0	100±1.73_**	15.70±0.85_**
NBH	62.5	121.49±1.59_**	24.50±1.77_**
NBH	125.0	125.82±2.12_*	28.71±2.08_**
NBH	250.0	130.54±1.93_**	31.10±1.93_**

Group code	Concentration of NBH/(μg·mL_-1)	Cell viablility rate(%)	Testosteron secretion/(nmol·L_-1)
Control	0	100±1.73_**	15.70±0.85_**
NBH	62.5	121.49±1.59_**	24.50±1.77_**
NBH	125.0	125.82±2.12_*	28.71±2.08_**
NBH	250.0	130.54±1.93_**	31.10±1.93_**

No.	Name	δ
1	Pantothenic acid	0.90(s), 0.94(s), 3.96(s)
2	Leucine	0.96(d), 0.97(d), 1.68(m), 3.74(m)
3	Valine	0.99(d, 7.0), 1.01(d, 7.0), 1.04(d)
4	3-Hydroxybutyrate	1.15(d, 6.0)
5	Ethanol	1.19(t, 7.2), 3.65(q)
6	Lipid	1.27(m), 2.0(m), 4.24(m)
7	Lactate	1.33(d), 4.11(q)
8	Alanine	1.48(d, 7.2), 3.78(m)
9	Lysine	1.72(m), 1.89(m), 3.04(t, J=7.57 Hz), 3.76(t, J=6.11 Hz)
10	Acetic acid	1.92(s)
11	Pyroglutamate	2.06(m), 2.35(m)
12	Glutamate	2.09(m), 2.13(m), 2.36(m), 2.46(m)
13	Methionine	2.14(s), 2.65(t)
14	Citrate	2.53(d), 2.69(d)
15	Dimethylamine	2.73(s)
16	Sarcosine	2.76(s), 3.60(s)
17	Aspartate	2.85(dd, 17.3, 3.7), 3.91(m)
No.	Name	δ
18	Glutathione	2.16(m), 2.56(m), 2.94(m)
19	Creatine	3.03s, 3.23(s), 3.93s
20	Ethanolamine	3.12(t, 5.5 Hz), 3.15(t, J=7.2 Hz), 3.86(t)
21	Malonate	3.15(s)
22	O-Phosphocholine	3.21(s)
23	Phosphocholine	3.22(s), 3.59(m), 4.17(m)
24	Histidine	3.25(dd), 7.08(s), 7.82(s)
25	Betaine	3.27(s)
26	Inositol	3.28(t, J=9.0 Hz), 3.63(t, J=9.6 Hz), 4.07(t, J=3.0 Hz)
27	Scyllo-inositol	3.37(s)
28	Taurine	3.44(t)
29	Methyl phosphate	3.47(d, 10.2)
30	Glycero	3.55(dd, 4.2, 9.6), 3.66(dd, 4.2, 11.4)
31	Glycine	3.57(s), 3.68(s)
32	Threonine	1.33(d), 3.58(d), 4.25(m)
33	Hippurate	3.97(d, 6.0), 7.54(t)
34	Adenosine monophosphate	4.01(m), 4.37(m), 4.51(m), 6.14(d), 8.27(s), 8.61(s)
35	Adenosine	4.30(dd), 8.24(s), 8.35(s)
36	Inosine	4.28(m), 4.44(m), 6.11(d), 8.23(s), 8.34(s)
37	Lactose	4.46(d), 4.66(d, J=7.8 Hz)
38	β-Glucose	4.50(d, 7.9)
39	α-Glucose	5.24(d, 3.6)
40	Urea	5.81(s)
41	Uracil	5.81(d, J=9.0 Hz)
42	Guanosine monophosphate	5.94(d), 8.20(s)
43	Uridine monophosphate	6.00(m), 8.11(d)
44	Cytidine	6.08(d), 7.85(d)
45	Fumaric acid	6.52(s)
46	Tyrosine	6.89(d), 7.19(d)
47	Tryptophan	7.33(d)
48	Phenylacetylglycine	3.68(s), 7.37(m), 7.43(m)
49	Uridine	7.92(d, 8.2)
50	Inosine monophosphate	8.25(s), 8.58(s)
51	Formate	8.46(s)

No.	Name	δ
1	Pantothenic acid	0.90(s), 0.94(s), 3.96(s)
2	Leucine	0.96(d), 0.97(d), 1.68(m), 3.74(m)
3	Valine	0.99(d, 7.0), 1.01(d, 7.0), 1.04(d)
4	3-Hydroxybutyrate	1.15(d, 6.0)
5	Ethanol	1.19(t, 7.2), 3.65(q)
6	Lipid	1.27(m), 2.0(m), 4.24(m)
7	Lactate	1.33(d), 4.11(q)
8	Alanine	1.48(d, 7.2), 3.78(m)
9	Lysine	1.72(m), 1.89(m), 3.04(t, J=7.57 Hz), 3.76(t, J=6.11 Hz)
10	Acetic acid	1.92(s)
11	Pyroglutamate	2.06(m), 2.35(m)
12	Glutamate	2.09(m), 2.13(m), 2.36(m), 2.46(m)
13	Methionine	2.14(s), 2.65(t)
14	Citrate	2.53(d), 2.69(d)
15	Dimethylamine	2.73(s)
16	Sarcosine	2.76(s), 3.60(s)
17	Aspartate	2.85(dd, 17.3, 3.7), 3.91(m)
No.	Name	δ
18	Glutathione	2.16(m), 2.56(m), 2.94(m)
19	Creatine	3.03s, 3.23(s), 3.93s
20	Ethanolamine	3.12(t, 5.5 Hz), 3.15(t, J=7.2 Hz), 3.86(t)
21	Malonate	3.15(s)
22	O-Phosphocholine	3.21(s)
23	Phosphocholine	3.22(s), 3.59(m), 4.17(m)
24	Histidine	3.25(dd), 7.08(s), 7.82(s)
25	Betaine	3.27(s)
26	Inositol	3.28(t, J=9.0 Hz), 3.63(t, J=9.6 Hz), 4.07(t, J=3.0 Hz)
27	Scyllo-inositol	3.37(s)
28	Taurine	3.44(t)
29	Methyl phosphate	3.47(d, 10.2)
30	Glycero	3.55(dd, 4.2, 9.6), 3.66(dd, 4.2, 11.4)
31	Glycine	3.57(s), 3.68(s)
32	Threonine	1.33(d), 3.58(d), 4.25(m)
33	Hippurate	3.97(d, 6.0), 7.54(t)
34	Adenosine monophosphate	4.01(m), 4.37(m), 4.51(m), 6.14(d), 8.27(s), 8.61(s)
35	Adenosine	4.30(dd), 8.24(s), 8.35(s)
36	Inosine	4.28(m), 4.44(m), 6.11(d), 8.23(s), 8.34(s)
37	Lactose	4.46(d), 4.66(d, J=7.8 Hz)
38	β-Glucose	4.50(d, 7.9)
39	α-Glucose	5.24(d, 3.6)
40	Urea	5.81(s)
41	Uracil	5.81(d, J=9.0 Hz)
42	Guanosine monophosphate	5.94(d), 8.20(s)
43	Uridine monophosphate	6.00(m), 8.11(d)
44	Cytidine	6.08(d), 7.85(d)
45	Fumaric acid	6.52(s)
46	Tyrosine	6.89(d), 7.19(d)
47	Tryptophan	7.33(d)
48	Phenylacetylglycine	3.68(s), 7.37(m), 7.43(m)
49	Uridine	7.92(d, 8.2)
50	Inosine monophosphate	8.25(s), 8.58(s)
51	Formate	8.46(s)

No.	Metabolites	VIP	FC	P-value	Trend_a	Related pathway
1	Pantothenic acid	1.70	4.20	3.65×10_-5	↑_**	Pantothenate and CoA biosynthesis
2	Leucine	3.36	9.11	3.57×10_-4	↑_**	Aminoacyl-tRNA biosynthesis
3	Valine	2.27	9.52	1.89×10_-3	↑_**	Aminoacyl-tRNA biosynthesis, pantothenate
						and CoA biosynthesis
6	Lipid	1.67	9.06	3.10×10_-5	↑_**	-
7	Lactate	1.42	4.06	3.86×10_-3	↑_**	Pyruvate metabolism
8	Alanine	2.25	9.68	1.14×10_-3	↑_**	Aminoacyl-tRNA biosynthesis, alanine, aspar-
						tate and glutamate metabolism
9	Lysine	2.46	11.12	2.61×10_-5	↑_**	Aminoacyl-tRNA biosynthesis
						Lysine degradation
10	Acetic acid	2.21	12.07	2.37×10_-4	↑_**	Pyruvate metabolism
11	Pyroglutamate	1.97	5.44	1.63×10_-5	↑_**	Glutathione metabolism
12	Glutamate	1.78	6.95	1.04×10_-5	↑_**	Aminoacyl-tRNA biosynthesis, glutathione metabolism, alanine, aspartate and glutamate metabolism, D-glutamine and D-glutamate metabolism
13	Methionine	1.68	6.89	2.25×10_-4	↑_**	Aminoacyl-tRNA biosynthesis
17	Aspartate	1.42	4.01	3.64×10_-6	↑_**	Aminoacyl-tRNA biosynthesis, alanine, aspartate and glutamate metabolism, pantothenate and CoA biosynthesis, glycine, serine and threonine metabolism
18	Glutathione	1.08	18.44	1.87×10_-5	↑_**	Glutathione metabolism
20	Ethanolamine	1.09	4.45	1.08×10_-5	↑_**	Glycerophospholipid metabolism
21	Malonate	1.64	3.84	1.21×10_-3	↑_**	-
23	Phosphocholine	2.02	2.86	3.41×10_-5	↑_**	Glycerophospholipid metabolism
26	Inositol	1.66	6.59	2.38×10_-4	↑_**	Inositol phosphate metabolism
27	Scyllo-inositol	2.36	0.17	4.76×10_-4	↓_**	-
31	Glycine	1.14	0.67	4.67×10_-2	↓_*	Aminoacyl-tRNA biosynthesis, glutathione metabolism, glycine, serine and threonine metabolism, lysine degradation
No.	Metabolites	VIP	FC	P-value	Trend_a	Related pathway
32	Threonine	1.42	4.06	3.86×10_-3	↑_**	Aminoacyl-tRNA biosynthesis, glycine, serine
						and threonine metabolism
34	Adenosine monophosphate	1.16	4.30	2.00×10_-5	↑_**
35	Adenosine	1.23	4.37	2.27×10_-5	↑_**
36	Inosine	1.06	4.62	4.67×10_-5	↑_**
48	Phenylacetylglycine	1.14	0.67	4.67×10_-2	↓_*

¹H NMR Metabonomic Investigations of N-Benzylhexadecanamide Induced Proliferation and Iestosterone Secretion of Mouse Testicular Leydig Cells^†

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