Cytotoxic furanosesquiterpenoids and steroids from Ircinia mutans sponges

Figures & data

Figure 1. Structures of furodysin (1), furodysinin (2), and furoircin (3) isolated from Ircinia mutans.

Figure 2. Structures of steroids identified from Ircinia mutans.

Figure 3. HPLC chromatograms of the subfraction FS6-10- F25-29 (steroids fraction) isolated from I. mutans (summer sample).

Figure 4. GC-MS chromatograms of FH2-FH5 of the HPLC semi-purified compounds 4–13 from FS6-10- F25-29 (steroids fraction) isolated from I. mutans (summer sample).

Table 1. ¹H NMR (500 MHz) and ¹³C NMR (125 MHz) data of furoircin (3) in CDCl₃.

position	δH (J in Hz)	^1H-^1H COSY	δC (HSQC)	HMBC
1	7.34 brt (J = 1.8)	H-2	142.46	C-2, C-3, C-4
2	6.24 brd (J = 1.7)	–	111.49	C-1, C-3, C-4, C-12
3	–	–	123.82	–
4	7.17 brs	H-2, H-11	139.40	C-1, C-2, C-3, C-11, C-12
5	–	–	147.63	–
6	2.24 ddd (J = 11.9, 3.8, 3.8)	H-7	43.80	C-5, C-7, C-8, C-11, C-12, C-13
7	1.68 m	–	22.50	C-5, C-6, C-8, C-9, C-11
8	2.00 m	H-7	30.78	C-6, C-7, C-9, C-10
9	–	–	133.85	–
10	5.39 brdd (J = 1.8, 1.8)	H-11, H-12′	124.94	C-6, C-8, C-11, C-15
11	2.41 m	H-12′	36.77	C-2, C-4, C-9, C-10, C-12, C-15
12	2.38 dd (J = 12.6, 3.9)	–	25.97	C-2, C-3, C-4, C-6, C-11
12′	2.07 ddd (J = 12.7, 12.7, 2.6)	–	25.97	C-2, C-3, C-4, C-6, C-11
13	4.87 brs	H-14	110.14	C-5, C-6, C-11(w*), C-14
13′	4.72 brs	H-14, H-6	110.14	C-5, C-6, C-11(w), C-14
14	1.77 s	–	22.81	C-5, C-6, C-13
15	1.63 s	–	23.55	C-8, C-9, C-10, C-11, C-14

*Weak cross peak.

Table 2. Sterol composition of I. mutans detected by GC-MS.

Compound	Retention time (min)	Kovats retention index (KI) Column type (HP-5 MS)	Kovats retention index standard (Colum stationary type)	Base peak (m/z)	Molecular weight	Reference for identification
cholesta-5, 22-dien-3β-ol (7)	6.18	3087	3070 (active phase: methyl Silicone*)	55	384	webbook.nist.gov
cholest-5-en-3β-ol (cholesterol) (4)	6.67	3121	3192 (active phase: DB-5*)	43	386	Pherobase.com
cholesta-5, 7-dien-3β-ol (5)	7.27	3158	3160 (active phase: methyl Silicone *)	351	384	webbook.nist.gov
ergosta-5, 7, 22-trien-3β-ol (ergosterol) (6)	8.01	3204	3152 (active phase: DB-5*)	363	396	webbook.nist.gov
ergosta-5, 24-dien-3β-ol (24-methyl-desmosterol) (8)	8.22	3214	3230 (active phase: OV-1*)	314	398	webbook.nist.gov
ergost-5-en-3β-ol (campesterol) (9)	8.26	3216	3131 (active phase: HP-5 MS*)	43	400	webbook.nist.gov
stigmasta-5,22-dien-3β-ol (stigmasterol) (10)	8.97	3252	3248 (active phase: VF-5MS*) 3170 (active phase: HP-5 MS*)	412	412	webbook.nist.gov
γ-ergostenol (fungisterol) (11)	9.24	3265	3220 (active phase: Methyl Silicone*)	400	400	webbook.nist.gov
5α-stigmasta-7,22-dien-3β-ol (chondrillasterol) (12)	9.69	3288	3295 (active phase: DB-5MS*)	271	412	webbook.nist.gov
γ-sitosterol (clionasterol) (13)	10.13	3308	3351 (active phase: HP-5 MS*)	43	414	webbook.nist.gov

*Column type: VF-5MS; DB-5; HP-5 MS: 5% phenylmethyl polysiloxane, OV-1, DB1 = 100% methylsiloxane

Table 3. Sterol composition of the HPLC Purified fractions from FS6-10- F25-29, characterised by GC-MS.

Fraction	Compounds
FH2	Cholesta-5, 22-dien-3β-ol (7) (10.2%)*
	Cholesta-5, 7-dien-3β-ol (5) (13.1%)*
	Ergosta-5, 7, 22-trien-3β-ol (ergosterol) (6) (49.3%)*
	Ergosta-5, 24-dien-3β-ol (24-methyldesmosterol) (8) (12.9%)*
FH3	Cholest-5-en-3β-ol (cholesterol) (4) (60.8%)*
FH4	Cholest-5-en-3β-ol (cholesterol) (4) (6.3%)*
	Ergost-5-en-3β-ol (campesterol) (9) (6.3%)*
	Stigmasta-5,22-dien-3β-ol (stigmasterol) (10) (45.7%)*
	γ-Ergostenol (fungisterol) (11) (4.4%)*
	5α-Stigmasta-7,22-dien-3β-ol (chondrillasterol) (12) (15.9%)*
FH5	γ-Sitosterol (Clionasterol) (13) (65.8 %)*

*Area percentage (%) of the detected compounds using GC-MS.

Figure 5. Cytotoxic activity of compounds isolated from I. mutans collected in winter (A) and summer (B) against three human cancer cell lines. Values are presented as mean ± S.E.M. of 3–5 experiments. *Compound 1w was not active against MOLT-4 and MCF-7 cell lines; IC₅₀ > 100 µg/mL. **Compounds 4–13 were not tested against MCF-7 and HT-29 cell lines.

Figure 6. Cytotoxic activity of RP4-HPLC purified steroids isolated from I. mutans (summer sample) against human lymphoblastic leukaemia cell line. Values are presented as mean ± S.E.M. of 3-5 experiments. FH2: Compounds: 5, 6, 7 and 8, FH3: compound 4, FH4: compounds 4, 9, 10, 11 and 12; FH5: compound 13. Based on statistical analysis, cytotoxic activity of FH2 has significant difference with other fractions (p < 0.05) while FH4 and FH5 were not significantly different compared to that of cisplatin as a positive control (p > 0.05).