*3.3.1.1.2. Cythane diterpenes from Cyathus africanus*

*Cyathus africanus* is a medicinal basidiomycete fungus. Diterpenes have been reported to possess multiple bioactivities consisting of antimicrobial and anti-inflammatory properties [86]. The presently reported metabolites in this text are collected by the study of various scientists. Moreover, they have been characterized on the basis of their structural elucidation by spectroscopic methods and discussed in detail in this chapter (**Table 2**). Some of the new metabolites documented by various scientists are isolated from *C. africanus*. Cyathin Q (15) an important metabolite (**Figure 10**) showed autophagy-dependent apoptosis [87]. The gene sequence of this

**Carbon number**

1 **2** **3** **4** **5** **6** **7** **8** **9** **10** **11** **12** **13** **14** **15** **16** **17** **18** **19** **20** **21** **22** **Ref.**

**[85]**

**[85]**

**[85]**

**[85]**

**[85]**

**[85]**

**[85]**

**[87]**

**[88]**

**[88]**

**[88]**

**[88]**

**[88]**

**[89]**

**[89]**

**[89]**

123

23.4

23.4

22.4 172.3

21.1

20.3

21.3

20.8

21.4

22.6 28.0

57.3

57.9

59.3

57.1

22.0

22.6

19.9

19.6

19.8

21.6

23.2

21.2

22.6

23.2

21.9

19.8

20.5

20.7

20.2

21.8

21.9

22.2

19.7

19.4

19.3

20.0

22.5

19.8

http://dx.doi.org/10.5772/intechopen.79186

27.4

27.4

28.4

27.0

26.8

26.3

26.3

28.5

28.2

28.5

29.1

28.1

28.3

27.2

28.4

27.2

19.2

19.3

25.1

20.3

21.0

26.3

22.3

24.8

24.2

24.7

19.9

20.0

19.5

20.4

20.6

23.9

Diterpenes from Different Fungal Sources and Their 13C-NMR Data

17.8

17.3

17.2

17.8

24.9

13.6

13.4

17.7

27.0

16.7

18.9

19.7

17.7

18.0

18.0

16.4

66.6

64.8

169.4

66.3

63.1

60.6

62.2

196.2

172.1

194.7

196.1

194.2

192.9

66.5

66.6

26.9

81.5

76.7

76.2

80.6

81.9

106.4

107.4

75.5

76.5

77.4

74.8

80.2

76.0

80.8

91.3

209.2

69.2

125.9

150.5

69.1

71.4

71.8

70.2

78.5

33.5

159.1

78.2

79.5

157.6

69.3

69.1

127.3

143.1

147.8

129.7

143.7

149.6

55.9

49.3

144.7

130.1

146.8

145.2

142.4

146.0

143.8

143.3

156.5

130.0

72.6

72.0

128.6

118.2

74.6

76.6

160.2

135.8

72.2

159.9

160.5

69.9

128.8

129.8

39.2

29.1

38.5

30.5

28.2

124.4

27.6

32.2

30.3

121.3

32.1

24.8

26.2

25.7

28.4

29.2

27.0

55.8

55.8

50.6

47.7

47.6

43.0

47.1

51.2

50.3

50.9

43.6

42.6

42.2

47.9

50.9

43.6

33.6

34.1

38.4

37.2

35.3

37.3

35.2

36.2

38.0

17.5

31.2

31.7

30.8

37.4

38.1

39.0

29.2

34.1

31.0

28.9

29.7

29.5

29.8

35.8

33.8

35.1

33.7

28.0

32.3

29.1

28.9

35.5

45.7

44.8

45.1

45.0

49.4

46.5

45.3

47.2

48.6

45.1

46.2

43.6

42.8

45.2

43.4

58.2

44.1

41.6

41.7

46.8

142.6

41.0

41.4

38.5

152.7

36.5

36.0

37.6

31.8

47.0

45.8

45.6

146.2

147.3

139.2

176.2

177.0

177.1

172.5

140.6

144.0

139.9

78.5

77.0

76.6

176.4

146.7

179.6

142.3

142.9

140.4

141.8

140.7

145.0

142.6

140.8

144.9

141.8

79.2

77.4

77.5

141.9

140.7

144.4

129.0

129.2

29.4

209.7

209.0

211.1

209.6

29.3

29.7

29.7

23.7

23.2

22.8

209.8

75.3

210.6

145.9

145.9

39.8

83.7

84.3

53.4

84.5

38.6

39.9

38.8

34.9

33.8

33.7

83.8

82.1

53.0

**Compound 8a**

 **9a**

**10a**

**11a**

**12a**

**13a**

**14a**

**15c**

**16c**

**17c**

**18c**

**19c**

**20c**

**21c**

**22c**

**23c**

**Figure 9.** Representative members of abietatetraenes.


[74], *C. striatius* [75], *Strobilurus tenacellus* [76], and *Hericium erinaceus* [77–84]. Some cyathane

*Cyathus gansuensis* was reported in 2002 and produced valuable bioactive metabolites from fermented grains of barley and rice [13] by transformation. Recently, seven new [85] metabolites (8–14) named have been isolated from fruiting body of *C. gansuensis* as presented in

Biological activity: NO inhibition activity was tested on mouse monocyte, macrophages. Seven newly discovered cyathane diterpene derivatives showed inhibitory activity against the NO production in LPS-activated macrophages. The fungus can be a good choice for a

13C-NMR structural elucidation: The detail of 13C-NMR is presented in **Table 2**. 13C-NMR data for compounds 8–14 revealed 20 carbons ascribable for 4 methyls, 4 methylenes (one oxygen-

to NMR and HRTOFMS at m/z 341.2079, [M + Na] + presented molecular formula of 8 and 9

(seven degrees of unsaturation) [85].

*Cyathus africanus* is a medicinal basidiomycete fungus. Diterpenes have been reported to possess multiple bioactivities consisting of antimicrobial and anti-inflammatory properties [86]. The presently reported metabolites in this text are collected by the study of various scientists. Moreover, they have been characterized on the basis of their structural elucidation by spectroscopic methods and discussed in detail in this chapter (**Table 2**). Some of the new metabolites documented by various scientists are isolated from *C. africanus*. Cyathin Q (15) an important metabolite (**Figure 10**) showed autophagy-dependent apoptosis [87]. The gene sequence of this

(six degrees of unsaturation), 10 (cyathin L) C22H32O5

carbons. According

(six degrees of unsaturation),

(six degrees of unsaturation), 12 (cyathin N)

(seven degrees

transformation on a large scale to acquire enough pure metabolites for the future [85].

diterpenoids represented interesting and significant biological activities.

**Figure 9**. The L69 fungal strain was used to isolate these compounds (8–14).

ated), 4 methines (two oxygenated), two quaternary carbons, and six sp<sup>2</sup>

(seven degrees of unsaturation), 13 (cyathin O) C20H30O5

*3.3.1.1.1. Cythane diterpenes from Cyathus gansuensis*

of unsaturation degrees), 11 (cyathin M) C20H30O5

*3.3.1.1.2. Cythane diterpenes from Cyathus africanus*

**Figure 9.** Representative members of abietatetraenes.

(cyathin J and K) as C20H30O3

and 14 (cyathin P) C20H28O5

C20H28O5

122 Terpenes and Terpenoids


**Carbon number**

**2'** **3'** **4'** **5'** **Ref.** a13C NMR at 125

b13C NMR at 75

c13C NMR at 150

MHz CD3OD.

d13C-NMR spectroscopic data for compounds 24–33

e13C-NMR at 125

f13C-NMR 175

**Table 2.**

13C-NMR data of cyathane diterpenoids.

Diterpenes from Different Fungal Sources and Their 13C-NMR Data

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125

MHz in acetone-d6.

 MHz.

in MeOH at 200

 MHz.

MHz in CDCl3.

 MHz.

**[90]**

**[90]**

**[90]**

**[90]**

**[90]**

**[90]**

**[90]**

**[90]**

**[90]**

**[90]**

**[96]**

**[96]**

**Compound 8a**

 **9a**

**10a**

**11a**

**12a**

**13a**

**14a**

**15c**

**16c**

**17c**

**18c**

**19c**

**20c**

**21c** 73.5 69.6 75.5 65.1

65.0

73.3

70.3

74.6

**22c**

**23c**


**Carbon number**

**Carbon number**

**1** **2** **3** **4** **5** **6** **7** **8** **9** **10** **11** **12** **13** **14** **15** **16** **17** **18** **19** **20** **21**

**1'**

25.3

20.1

22.2

21.0

31.2

26.7

14.5

24.3

17.4

17.1

64.4

65.0

209.7

76.2

122.6

127.3

157.5

145.9

73.0

71.6

34.8

36.6

54.0

43.5

34.5

39.3

33.1

32.0

52.9

45.8

43.7

43.2

84.0

181.7

188.5

144.2

127.0

211.1

213.7

53.1

**Compound 24e**

 **25d**

**Compound 8a**

 **9a**

**10a**

**11a**

**12a** **26d** 88.5 83.5 140.2 141.4

39.0 56.6 34.8 30.2 48.7 37.0 72.4 157.0 123.2 210.6

64.5 15.6 23.6 28.2 24.3 19.6

20.4

22.6

21.2

24.6

21.0

23.3

20.6

21.8 56.6 105.3

106.3

\*\*\*

22.3

20.5

21.2

20.2

19.3

23.4

21.0

20.5

21.5

22.0

29.8

27.5

26.3

27.2

33.0

33.0

26.2

27.0

27.7

20.7

17.3

22.4

19.6

20.5

22.4

26.0

24.5

24.7

17.9

12.0

12.2

12.2

14.6

16.1

13.2

16.4

16.4

64.4

58.9

58.9

58.9

59.1

62.1

62.0

192.9

194.2

210.1

111.2

110.8

111.1

104.8

108.2

107.4

85.4

85.3

123.2

126.6

126.9

126.6

57.0

70.6

70.2

158.2

154.6

157.3

149.0

149.2

149.2

64.0

49.7

49.8

138.6

148.2

72.3

80.0

79.9

79.9

74.6

76.9

76.3

72.4

62.1

32.3

28.3

27.6

28.2

29.9

33.3

31.5

30.1

35.0

42.6

49.9

47.3

47.2

50.2

50.4

42.9

49.2

50.4

33.7

36.2

35.3

36.1

30.3

30.8

37.1

37.0

37.1

34.7

31.0

30.5

30.9

29.7

29.9

29.7

30.4

34.3

54.6

42.6

44.9

42.8

42.7

41.9

46.0

40.6

44.5

37.1

41.6

43.3

41.3

38.9

37.5

40.5

40.4

36.1

78.2

137.8

174.0

140.1

53.3

53.5

177.0

136.6

139.8

77.8

137.2

141.6

139.6

192.3

193.2

144.9

139.9

140.7

72.8

37.2

209.5

83.3

125.8

126.4

210.9

28.4

29.1

47.0

82.8

84.5

90.5

215.4

216.0

53.3

38.5

38.3

124 Terpenes and Terpenoids

**27d**

**28d**

**29d**

**30d**

**31d**

**32d**

**33d**

**34f**

**33a**

**13a**

**14a**

**15c**

**16c**

**17c**

**18c**

**19c**

**20c**

**21c**

**22c**

**23c**

**Table 2.** 13C-NMR data of cyathane diterpenoids.

**Figure 10.** Newly (8–14) isolated metabolites from *C. gansuensis* [85].

strain has also been reported and submitted to GenBank with an accession numbers JX103204. Sequences of analysis exhibited 100% homology with that of fungus *C. africanus*. Compounds 16–20 (D–H) structurally represented new group of metabolites, while neosarcodonin O (21), cyathatriol (22), and 11-O-acetylcyathatriol (23) are also known cyathane diterpenes. Five novel compounds are isolated from *C. africanus* and show potential NO inhibition and cytotoxicity against HeLa cell line in vitro [88]. The structural elucidation is also described in **Figure 11**.

superoxide, and transition metals like iron or zinc-sulfur clusters, and overproduction of NO is involved in many pathogenic diseases, including inflammation and cancer. The inhibition of NO overproduction in cells may prevent the occurrence of inflammatory diseases and cancer. The inhibition capacity (IC50) was more pronounced for 16, 17, and 19 by exhibiting NO inhibition 79.44, 89.2 and 84.33% reduction, respectively [91]. Moreover, inhibition of NO is concentration dependent as compounds 16–23 showed no NO inhibition at concentration 100 μM [88, 89]. COX-2 and iNOS are two major inflammatory mediators in brain neurodegeneration [92, 93]. Compounds isolated from *C. africanus* [90] showed strong COX-2 and iNOS capacities. Western blot analysis demonstrated that compounds 24 and 28 significantly suppressed LPS-induced COX-2 expression, whereas compounds 27, 28, 30, 31, and 33 markedly inhibited LPS-induced iNOS expression. Among these compounds, 28 showed strong inhibitory effects on both COX-2 and iNOS. Interestingly, 30 abolished LPS-induced iNOS expression but did not affect LPS-induced COX-2 expression. In addition, we also assayed

Diterpenes from Different Fungal Sources and Their 13C-NMR Data

http://dx.doi.org/10.5772/intechopen.79186

127

13C-NMR structural elucidation: The 13C-NMR spectrum of some of the compounds isolated

*Hericium* genus is among the most blessed medicinal and eatable mushrooms and known to produce secondary metabolites with the potential to treat neurodegenerative diseases. It enables improvement of many brain-related disorders [94]. In this regard, neurotrophins are nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) involved

the activities of iNOS enzyme [90].

from *C. africanus* presented in **Table 2**.

*3.3.1.1.3. Cythane diterpene from Hericium erinaceus and H. flagellum*

**Figure 12.** Structures of metabolites isolated from *C. africanus* [88, 89].

Ten new polyoxygenated cyathane diterpenoids, named as neocyathins (24–33), together with four known diterpenes are isolated from fungus *Cyathus africanus* (**Figure 12**). These compounds were isolated and identified by <sup>13</sup>C-NMR technique [90] (**Figure 13**).

Biological activity: Diterpenes with diverse bioactivities have been identified from plants and fungi [91]. Cyathin Q has the capacity to induce the apoptosis in HCT116 cells in a time- and dose-dependent manner. It was observed, when HCT116 cells exposed to 10 mM cyathin Q for 24 h exhibited apoptotic cells 82.07% [87]. This compound induced hallmarks of apoptotic events in HCT116 cells, including caspase activation, cytochrome c release, poly (ADP-ribose) polymerase (PARP) cleavage, and depolarization of the mitochondrial inner transmembrane potential. Nitric oxide has the capacity to react with aqueous oxygen,

**Figure 11.** Structures of cyathane Q (15) isolated from *C. africanus* [87].

**Figure 12.** Structures of metabolites isolated from *C. africanus* [88, 89].

strain has also been reported and submitted to GenBank with an accession numbers JX103204. Sequences of analysis exhibited 100% homology with that of fungus *C. africanus*. Compounds 16–20 (D–H) structurally represented new group of metabolites, while neosarcodonin O (21), cyathatriol (22), and 11-O-acetylcyathatriol (23) are also known cyathane diterpenes. Five novel compounds are isolated from *C. africanus* and show potential NO inhibition and cytotoxicity against HeLa cell line in vitro [88]. The structural elucidation is also described in **Figure 11**.

Ten new polyoxygenated cyathane diterpenoids, named as neocyathins (24–33), together with four known diterpenes are isolated from fungus *Cyathus africanus* (**Figure 12**). These com-

Biological activity: Diterpenes with diverse bioactivities have been identified from plants and fungi [91]. Cyathin Q has the capacity to induce the apoptosis in HCT116 cells in a time- and dose-dependent manner. It was observed, when HCT116 cells exposed to 10 mM cyathin Q for 24 h exhibited apoptotic cells 82.07% [87]. This compound induced hallmarks of apoptotic events in HCT116 cells, including caspase activation, cytochrome c release, poly (ADP-ribose) polymerase (PARP) cleavage, and depolarization of the mitochondrial inner transmembrane potential. Nitric oxide has the capacity to react with aqueous oxygen,

pounds were isolated and identified by <sup>13</sup>C-NMR technique [90] (**Figure 13**).

**Figure 10.** Newly (8–14) isolated metabolites from *C. gansuensis* [85].

126 Terpenes and Terpenoids

**Figure 11.** Structures of cyathane Q (15) isolated from *C. africanus* [87].

superoxide, and transition metals like iron or zinc-sulfur clusters, and overproduction of NO is involved in many pathogenic diseases, including inflammation and cancer. The inhibition of NO overproduction in cells may prevent the occurrence of inflammatory diseases and cancer. The inhibition capacity (IC50) was more pronounced for 16, 17, and 19 by exhibiting NO inhibition 79.44, 89.2 and 84.33% reduction, respectively [91]. Moreover, inhibition of NO is concentration dependent as compounds 16–23 showed no NO inhibition at concentration 100 μM [88, 89]. COX-2 and iNOS are two major inflammatory mediators in brain neurodegeneration [92, 93]. Compounds isolated from *C. africanus* [90] showed strong COX-2 and iNOS capacities. Western blot analysis demonstrated that compounds 24 and 28 significantly suppressed LPS-induced COX-2 expression, whereas compounds 27, 28, 30, 31, and 33 markedly inhibited LPS-induced iNOS expression. Among these compounds, 28 showed strong inhibitory effects on both COX-2 and iNOS. Interestingly, 30 abolished LPS-induced iNOS expression but did not affect LPS-induced COX-2 expression. In addition, we also assayed the activities of iNOS enzyme [90].

13C-NMR structural elucidation: The 13C-NMR spectrum of some of the compounds isolated from *C. africanus* presented in **Table 2**.

#### *3.3.1.1.3. Cythane diterpene from Hericium erinaceus and H. flagellum*

*Hericium* genus is among the most blessed medicinal and eatable mushrooms and known to produce secondary metabolites with the potential to treat neurodegenerative diseases. It enables improvement of many brain-related disorders [94]. In this regard, neurotrophins are nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) involved

Biological activity and 13C-NMR analysis: All of the metabolites isolated from *H. erinaceus* and *H. flagellum* exhibited strong neutrotrophin capacity [95, 96]. Metabolites were also studied through 13C-NMR; compound 34 exhibited the presence of five non-proton-bearing carbons, including three olefinic (*δ*C 139.9, 136.6, 138.6) and two aliphatic carbons (*δ*C 40.6, 49.2). Furthermore, five methylene groups with corresponding carbons between *δ*C 28.4 and 38.5 ppm, a further oxygenated methylene group at *δ*C 65.1, vicinal to two aliphatic methines at *δ*C 40.4, and six methines at *δ*C 69.6–105.3 ppm were observed. 13C shifts and correlations of the HSQC-DEPT spectrum showed high similarity to **35** which was a derivative of the cyathane diterpenoid **34**. The major difference between the two compound spectra was the missing methoxy group at C-11 in **35** (**Figure 13**). The detail of 13C-NMR data is described in

Diterpenes from Different Fungal Sources and Their 13C-NMR Data

http://dx.doi.org/10.5772/intechopen.79186

Indole diterpenes are the broad class of secondary metabolites with enormous structural and functional diversity. They mostly occur in filamentous fungal members having most abundance in *Penicillium*, *Aspergillus*, *Neotyphodium*, and *Claviceps* [97, 98]. This class of diterpenes is generally divided into two main groups, paxilline type and non-paxilline type [98], though it mainly consists of cyclic diterpenoid backbone in addition to an indole

The marine fungi *A. nidulans* was reported to be the source of 19-hydroxypenitrem A (1) and 19-hydroxypenitrem E (2). The 13C-NMR spectrum of 19-hydroxypenitrem A (C37H44ClNO7

An endophytic fungi *Drechmeria* sp. was found to be the reservoir of diverse indole diterpenes including drechmerin A (38), drechmerin B (39), drechmerin C (40), drechmerin D (41),

)

129

and 7 sp3

) carbon

terminal), 1 sp2

) lack chlorine atom but have one

and 9 sp2

**Table 2**.

moiety.

*3.3.2. Indole diterpenes*

*3.3.2.1. Diversity of indole diterpenes*

*3.3.2.1.1. Indole diterpenes from Aspergillus nidulans*

additional hydrogen atom [98] (**Figure 15**).

**Figure 15.** Indole diterpenes from *A. nidulans* [98].

provided 37 resonance states from 5 methyl, 8 methylene (with 2 sp2

atoms. In comparison, 19-hydroxypenitrem E (C37H45NO7

*3.3.2.1.2. Drechmeria sp.: a rich source of indole diterpenes*

methines (with 5 oxygenated), and 16 quaternary (with 5 oxygenated sp<sup>3</sup>

**Figure 13.** Structures of metabolites isolated from *C. africanus* [90].

in survival, maintenance, and regeneration of specific neuronal populations in the adult brain [95]. Therefore, the metabolites extracted from *Hericium* are important source of metabolites and source as remedy in the fight against neurodegenerative diseases such as Parkinson's, Alzheimer's, and Huntington's diseases, which are accompanied by decreased neurotrophic factor expression [102]. Two new potential metabolites have been isolated from *H. erinaceus* (strain STMA 06157B) and *H. flagellum* (strain CBS 103681) [96] (**Figure 14**).

**Figure 14.** Structural elucidation of metabolites isolated from *H. erinaceus* and *H. flagellum* [96].

Biological activity and 13C-NMR analysis: All of the metabolites isolated from *H. erinaceus* and *H. flagellum* exhibited strong neutrotrophin capacity [95, 96]. Metabolites were also studied through 13C-NMR; compound 34 exhibited the presence of five non-proton-bearing carbons, including three olefinic (*δ*C 139.9, 136.6, 138.6) and two aliphatic carbons (*δ*C 40.6, 49.2). Furthermore, five methylene groups with corresponding carbons between *δ*C 28.4 and 38.5 ppm, a further oxygenated methylene group at *δ*C 65.1, vicinal to two aliphatic methines at *δ*C 40.4, and six methines at *δ*C 69.6–105.3 ppm were observed. 13C shifts and correlations of the HSQC-DEPT spectrum showed high similarity to **35** which was a derivative of the cyathane diterpenoid **34**. The major difference between the two compound spectra was the missing methoxy group at C-11 in **35** (**Figure 13**). The detail of 13C-NMR data is described in **Table 2**.
