**2. Extracts preparation and iridoids isolated**

**IRIDOIDS SPECIES PLANT MATERIAL REF.**

plumieridin B 1: *Plumeria rubra* 1: flowers 1: [43]; 1: [120]

pulosarioside 14: *Alyxia reinuardtii* 14:bark 14: [40]; 14: [121] scholarein A 15: *Alstonia scholaris* 15:bark 15: [60] scholarein B 15: *Alstonia scholaris* 15:bark 15: [60] scholarein C 15: *Alstonia scholaris* 15:bark 15: [60] scholarein D 15: *Alstonia scholaris* 15:bark 15: [60] secologanin 32: *Catharanthus roseus* 32:leaves 32: [4] sweroside 34: *Alstonia macrophylla* 34:stem 34: [128]

1,3:root; 7:leaves; 7:aerial parts

*Plumeria obtusa* 1:flowers; 7:aerial parts 1: [43]; 1: [118]; 1: [120];

1, 3:root; 10:stem; 10:leaves

13:fruit; 13,18:leaves; 18:root

13:fruit; 13:cortex; 13,18:leaves; 18:root

18: *Thevetia peruviana* 18:leaves; 18:root 18: [125]; 18: [127]

18: *Thevetia peruviana* 18:root 18: [125]

18: *Thevetia peruviana* 18:root 18: [125]

18: *Thevetia peruviana* 18:leaves; 18:root 18: [125]; 18: [127]

10: *Allamanda neriifolia* 10:stem; 10:leaves 10: [39]; 10: [40]

1: [65]; 3: [38]; 3: [40]; 3: [66]; 7: [72]; 7: [119]

7: [72]

1: [65]; 3: [38]; 3: [40]; 3: [66]; 10: [39]

13: [46]; 13: [56]; 13: [61]; 13: [124]; 13,18: [42]; 13,18: [62]; 18: [122]; 18: [123]; 18: [125]

13: [46]; 13: [56]; 13: [61]; 13: [68]; 13: [124]; 13,18: [42]; 13,18: [62]; 18: [125]; 18: [126]

24:bark 24: [87]

*acutifolia;* 7: *P. obtusa*

24: *Himatanthus articulatus*

1: *Plumeria rubra;* 7:

1: *Plumeria rubra*; 3: *P. acutifolia*; 10: *Allamanda neriifolia*

13: *Cerbera manghas*; 18: *Thevetia peruviana*

13: *Cerbera manghas*; 18: *Thevetia peruviana*

plumieride-1<sup>α</sup> 1: *Plumeria rubra;*3: *P.*

plumieride-1β-*O*-β-Dglucopyranosyl

156 Column Chromatography

plumieridin A

protoplumericin A-1α

protoplumericin B (13-*O*-*p*-*O*glucosylcaffeoyl-plumieride)

theveside

theviridoside

theviridoside-10-*O*-β-Dfructofuranosyl

theviridoside-10-O-β-Dglucopyranosyl

theviridoside-3'-*O*-β-Dglucopyranosyl

theviridoside-6'-*O*-β-Dglucopyranosyl

**Table 1.** Iridoids found in Apocynaceae family.

Table 2 illustrates the extraction methods and the iridoids isolated of some genera of Apocynaceae.



**SPECIES / PLANT MATERIAL**

latex *H. sucuuba,* (Spruce ex Müll. Arg.) Woodson / bark

*Nerium indicum* Mill. / root

*Plumeria acutifolia* Poir. */* root

*P. acutifolia* Poir. */* leaves

*P. bicolor* Seem. / bark

*P. rubra* L. var. *acutifolia* (Poir.) Woodson / bark

> *P. rubra* L. / steam bark

> *P. rubra* L. / heartwood

> *P. rubra* L. / stem bark

> *P. rubra* L. / stem bark

*P. rubra* L. var. *acutifolia* (Poir.) Woodson /

**SOLVENT/ EXTRACTION METHOD**

Petrol/ Soxhlet apparatus

> MeOH/ Percolation

> MeOH/ Exhaustive Maceration

> MeOH/ Exhaustive Maceration

> MeOH/ Maceration

CH2Cl2:MeOH (1:1) and MeOH/ Maceration

> EtOAc/ Percolation

> MeOH/ Maceration

> MeOH/ Maceration

**ISOLATED IRIDOIDS REF.**

http://dx.doi.org/10.5772/55784

A General Description of Apocynaceae Iridoids Chromatography

Plumericin [130]

15-demethylplumieride [71]

Plumericin, isoplumericin [86]

demetilplumieride, plumieride, isoplumieride [70]

Plumericin, isoplumericin, plumieride, fulvoplumierin [75]

[66]

159

[85]

[44]

[48]

EtOH/ Maceration Plumericin [100]

Plumericin, 13-*O*-coumaroylplumieride, 13-Ocaffeoylplumieride, 13-deoxyplumieride, plumenoside, 1 αplumieride, 1 α-protoplumericin A, plumieride, protoplumericin A, 8-isoplumieride

15-demethylisoplumieride, 15-

Plumericin, isoplumericin, plumieride, l3-*O*-coumaroylplumieride , protoplumericine A

Fulvoplumierin, allamandin, allamcin, plumericin, 15 demethylplumieride, plumieride, α-allamcidin*,* β-allamcidin, 13-*O*-*trans*-p-coumaroylplumieride

Fulvoplumerin, dihydroplumericin, plumieride , plumeridoids A, B and C, isoplumericin, plumericin, allamcin, allamandin, mixture of plumeridoid C and epiplumeridoid C

EtOH/ Maceration Plumieridin A, plumieridin B, plumericin, plumieride [120]


**SPECIES / PLANT MATERIAL**

158 Column Chromatography

*Alyxia reinwardtii* Blume/ leaves

*Cerbera manghas* L./ leaves

*C. manghas* L./ stem bark and root bark

> *C. manghas* L./ leaves

> *C. manghas* L./ leaves

> *C. manghas* L./ leaves

*C. odollam* Gaertn./ leaves

*Himatanthus articulatus* (Vahl) Woodson/ bark

*H. fallax* (Müll. Arg.) Plumel/ stem

*H. sucuuba* (Spruce ex Müll. Arg.) Woodson / latex

*H. sucuuba* (Spruce ex Müll. Arg.) Woodson/ bark

*H. sucuuba* (Spruce ex Müll. Arg.) Woodson / latex

*H. sucuuba* (Spruce ex Müll. Arg.) Woodson /

**SOLVENT/ EXTRACTION METHOD**

MeOH/ Maceration

MeOH/ Not

MeOH/

MeOH/ Percolation

MeOH/ Percolation

MeOH/ Percolation

MeOH/ Percolation

MeOH/ Maceration

EtOAc/

H2O/ Not described

H2O/ Not

**ISOLATED IRIDOIDS REF.**

4-epi-alyxialactone [59]

theviridoside, theveside [61]

[55]

[62]

[87]

Alyxialactone,

Cerberidol, epoxycerberidol, cyclocerberidol, cerberidol-3-*O*β-D-allopyranoside, cerberidol-3,10-bis-*O*-β-Dallopyranoside, epoxycerberidol-3-*O*-β-D-allopyranoside, cyclocerberidol-3-*O*-β-D-allopyranoside

Theviridoside, 10-carboxyloganin, loganin, cyclocerberidol-3-*O*-β-D-glucoside, epoxycerberidol-3-*O*-β-D-glucoside

10-*O*-benzoyltheveside, 10-dehydrogeniposide, loganin,

Cerberidol, cyclocerberidol, cerberidol-3-O-β-Dallopyranoside, cyclocerberidol-3-O-β-D-allopyranoside [55]

1β-*O*- β -D-glucopyranosylplumeric acid, plumeride-1 β -*O*- β -D-glucopyranosyl, plumericin, isoplumericin

15-demethylisoplumieride, 15-

demethylplumieride, plumieride, isoplumieride [70]

Maceration Isoplumericin, plumericin, plumieride [89]

EtOH/ Maceration Isoplumericin, plumericin [88]

described Plumericin, isoplumericin [92]

BuOH/ Partition Plumieride, isoplumieride, 15-demethylplumieride [73]

described Theviridoside, theveside [124]

Percolation Cerbinal, cerberic acid, cerberinic acid [67]


**3. Chemical structures**

H

O O

**6** *Plumieride* R = H; R1

O H

**1** *Plumericin* R = H; R1

O H O

> <sup>R</sup> <sup>R</sup><sup>1</sup> H

O

**2** *Isoplumericin* R = CH3; R1

**7** *15-demethylplumieride* R=R1

H

O O R1

O-Glucosyl <sup>O</sup> OR

= CH3

O

O

**10** *Plumieride coumarate* 

**Figure 2.** The most isolated iridoids of Apocynaceae family

O-Glucosyl

**4. Chromatographic separation** 

**4.1. Thin layer chromatography** 

OH

Figure 2. The most isolated iridoids of Apocynaceae family

O

O

O-Glucosyl

<sup>O</sup> <sup>O</sup> HO

Preparative chromatography was performed with thin layer chromatography (TLC) aluminum sheets and 8:2 chloroform/ methanol as mobile phase of a fraction from the 95% EtOH extract of stems of *Alstonia macrophylla*. This procedure led to the

Another interesting application of preparative chromatography is described in [47] with ethanol extract of ground stems of *A. schottii*. This extract was fractionated by bioassay monitoring and after successive liquid-liquid partition and flash column chromatography; the authors obtained the iridoids allamandin; allamcin and a mixture of plumericin and isoplumericin. This separation was performed with a Chromatotron rotor (silica gel (Si gel), 2-mm) with 1 % methanol in chloroform as solvent system.

The bioassay-guided fractionation of the extracts of *Plumeria rubra* barks also proved to be a successful strategy, leading to the isolation of eleven substances, of which eight were iridoids. In this context, the aqueous extract of *Plumeria rubra* bark has iridoids such as the epimers, *α*-allamcidin and *β*-allamcidin, which were resolved by preparative TLC on Si-gel G plates (20 20 cm, 250

**11** *Plumieride coumarate glucoside*

OH OH OH

H

O O

O

O H

O

H

O O

**3** *Allamcin* **4** *Allamandin* 

H <sup>O</sup> O-Glucosyl

OH <sup>O</sup>

**8** *Isoplumieride*

O

isolation of the compounds sweroside (2 mg) and naresuanoside (3 mg) [128]

μm, Merck®), using chloroform/ ethyl acetate/ methanol (3:3:1) as solvent system [44].

H

O O

O

O H

O

O

= H

**3. Chemical structures** 

O

= CH3

= H

*Winchia calophylla* A. DC. / stem bark

Table 2. Main extraction methods to obtain iridoids of Apocynaceae family.

O

OH

H

O H O

O H

EtOH/ Hot

O

OH

A General Description of Apocynaceae Iridoids Chromatography

H

O O

O H O

O H

10-*O*- β -D-glucopyranosyltheviridoside, 3'*O*- β -D-glucopyranosyltheviridoside

161

extraction Loganin [129]

http://dx.doi.org/10.5772/55784

O O

O O

**5** *Fulvoplumierin*

O

<sup>H</sup> O-Glucosyl

H

**9** *Loganin*

HO

O O

**Table 2.** Main extraction methods to obtain iridoids of Apocynaceae family.

#### **3. Chemical structures** Table 2. Main extraction methods to obtain iridoids of Apocynaceae family.

**3. Chemical structures** 

**SPECIES / PLANT MATERIAL**

160 Column Chromatography

flowers *P. rubra* L. var. *acutifolia* (Poir.) Woodson/ flowers

*P. obtusa* L. / flower

*P. obtusa* L. / aerial parts

*P. obtusa* L. / leaves

*P. obtusa* L. / leaves

*P. inodora* Jacq. / stems

*P. bicolor* Seem. / bark

*Rauwolfia grandiflora* Mart. Ex A. DC. / bark

*Thevetia peruviana* (Pers.) K. Schum. / Leaves

*T. peruviana* (Pers.) K. Schum. / root

*Winchia calophylla* A. DC. / stem bark

**SOLVENT/ EXTRACTION METHOD**

MeOH/ Maceration

MeOH/ Maceration

MeOH/

MeOH/ Maceration

MeOH/ Percolation

MeOH/ Percolation

EtOH/ Hot

**Table 2.** Main extraction methods to obtain iridoids of Apocynaceae family.

**ISOLATED IRIDOIDS REF.**

Plumieride [114]

[72]

[37]

[127]

EtOH/ Maceration Plumericidine [97]

EtOH/ Maceration Plumieride coumarate glucoside [118]

Obtusadoids A and B, plumieridin A, plumieridine, 1α-plumieride, 15-demethylplumieride

6"-*O*-acetylplumieride *p*-E-coumarate, 6"-*O*acetylplumieride,*p*-Z-coumarate, plumieride, plumieride *p*-Z-coumarate, plumieride *p*-E-coumarate

Maceration 1α-plumieride [119]

H2O/ Maceration Plumieride [113]

EtOH/ Maceration Loganic acid, loganin, boonein, isoboonein [1]

10-*O*- β -D-fructofuranosyltheviridoside,

Theveside, theviridoside, 10-*O*- β -D-fructofuranosyltheviridoside, 6'*O*- β -D-glucopyranosyltheviridoside, 10-*O*- β -D-glucopyranosyltheviridoside, 3'*O*- β -D-glucopyranosyltheviridoside

extraction Loganin [129]

6'*O*- β -D-glucopyranosyltheviridoside [125]

10-*O*- β -D-glucopyranosyltheviridoside, 3'*O*- β -D-glucopyranosyltheviridoside

**1** *Plumericin* R = H; R1 = CH3

**3** *Allamcin* **4** *Allamandin* 

**5** *Fulvoplumierin*

**2** *Isoplumericin* R = CH3; R1 = H

O O

**6** *Plumieride* R = H; R1 = CH3

**7** *15-demethylplumieride* R=R1 = H

**9** *Loganin*

isolation of the compounds sweroside (2 mg) and naresuanoside (3 mg) [128]

μm, Merck®), using chloroform/ ethyl acetate/ methanol (3:3:1) as solvent system [44].

**10** *Plumieride coumarate* 

**11** *Plumieride coumarate glucoside*

Preparative chromatography was performed with thin layer chromatography (TLC) aluminum sheets and 8:2 chloroform/ methanol as mobile phase of a fraction from the 95% EtOH extract of stems of *Alstonia macrophylla*. This procedure led to the

Another interesting application of preparative chromatography is described in [47] with ethanol extract of ground stems of *A. schottii*. This extract was fractionated by bioassay monitoring and after successive liquid-liquid partition and flash column chromatography; the authors obtained the iridoids allamandin; allamcin and a mixture of plumericin and isoplumericin. This separation was performed with a Chromatotron rotor (silica gel (Si gel), 2-mm) with 1 % methanol in chloroform as solvent system.

The bioassay-guided fractionation of the extracts of *Plumeria rubra* barks also proved to be a successful strategy, leading to the isolation of eleven substances, of which eight were iridoids. In this context, the aqueous extract of *Plumeria rubra* bark has iridoids such as the epimers, *α*-allamcidin and *β*-allamcidin, which were resolved by preparative TLC on Si-gel G plates (20 20 cm, 250

Figure 2. The most isolated iridoids of Apocynaceae family **Figure 2.** The most isolated iridoids of Apocynaceae family

**4. Chromatographic separation** 

**4.1. Thin layer chromatography** 
