Halimane diterpenoids: sources, structures, nomenclature and biological activities

Diterpenes with a halimane skeleton constitute a small group of natural products that can be biogenetically considered as being between labdane and clerodane diterpenoids. Some of these compounds show biological activities, such as antitumour, mosquito repellency, germination inhibition and antimicrobial, as well as being biomarkers for tuberculosis. To the best of our knowledge, there are no reviews on these compounds. In this review, halimane skeleton diterpenoids are classified according to their biogenetic origin, characterization and/or the enzymes involved in their biosynthesis. Herein, a review of their synthesis or synthetic approaches is communicated.

Conicts of interest 8.

Introduction and background
Among natural products, terpenes are one of the most numerous and structurally diverse groups. They can be found in nearly all life forms showing a great number of functional activities. Currently, tens of thousands of compounds in this class 1,2 (around 55 000) are known. As soon as new compounds are discovered, from terrestrial or marine origin, 3 they are included in reviews on different terpene classes (monoterpenes, sesquiterpenes, 4 diterpenes, 5,6 sesterterpenes 7 and triterpenes 8 ).
Within terpenes, diterpenes are one of the largest families of natural molecules, with more than 18 000 compounds derived from GGPP (E,E,E-geranylgeranyl diphosphate). Classication of these compounds is done according to their biogenesis, leading to 126 different carbon skeletons known until now. 9 Two big groups can be distinguished among the diterpenes family: those diterpenes in which a pyrophosphate ion is involved in the rst biogenetic step and those diterpenes that are generated as a consequence of cyclizations that do not include pyrophosphate ion in the rst step. [10][11][12][13] Labdanes and related diterpenes (like clerodanes 14,15 ) are included in this second group. According to Peters,16 halimanes are found within the labdane and clerodane group, as indicated in the general biogenetic scheme (Scheme 1). The halimane-type He is co-author of more than 180 papers and he acts as referee for many international scientic journals. Currently he is the Dean of the Chemistry Faculty at the University of Salamanca. His current research interests are focused on the transformation of natural products into biologically active compounds, the chemistry of cyclopropanes, sulfones, tetrahydropyrans, and chiral amides, and recently organocatalysis and nanoparticles.
carbocation can be in position 10 leading to D 1 (10) or D 5 (10) halimanes or in position 5, due to a hydride 1,2-shi, leading to D 5 halimanes. Other halimanes can be understood from these intermediates or derivatives, as reported later on.
The name 'halimane' was introduced in order to provide a simple nomenclature for the isolated ent-halimic acid compound series 17 in Halimium viscosum (Fig. 1). Previously in the literature, these compounds were known by generic names, such as rearranged labdanes, 18,19 isolabdanes 20 or even friedolabdanes. [21][22][23][24][25] Halimium viscosum is a plant of the Cistaceae family that shows a Hispano-Mauritanian distribution and is abundant in the west of the Iberian Peninsula. Ent-halimic acid and acetoxy ent-halimic acid are found in a 1 : 1 ratio in considerable amounts in Halimium viscosum extracts (47%), 0.34% with respect of the dry plant weight. Nowadays, a cheap, quick and efficient methodology for the isolation of ent-halimic acid has been achieved. The study of the components of Halimium viscosum and the need for large amounts of ent-halimic acid for the synthesis of biologically active compounds led us to search for Halimium viscosum plants in new locations with better access. In this manner, we found several places for the study of the Halimium viscosum components, such as Villarino de los Aires (Salamanca, Spain), 17 La Fregeneda (Salamanca, Spain) 26,27 and Valparaiso (Zamora, Spain), 28 localities quite close together. To our surprise, in the rst location rearranged ent-labdanes (ent-halimanes) with an unsaturated side chain were isolated, in the second ent-halimanes with a saturated side chain were found, and, from the plants of Valparaiso, labdanes with the structural strangeness of having a carboxyl function at C-17 were only isolated. At the time of these studies, to nd different chemotypes of the same plant was a novelty. Furthermore, two other chemotypes of Halimium viscosum have been localized in Portugal near the border with Salamanca (Celorico da Beira 29 and San João da Pesqueira 30 ).
In Halimium viscosum, in addition to the referred ent-halimanes and labdanes, a series of compounds that show new carbon skeletons has been isolated, characterized and then synthesized. Among them, tricyclic diterpenoids such as valparanes [31][32][33][34][35] and valparolanes, 36 bicyclic diterpenoids with tormesane skeleton (a new type of sphenolobanes), 37,38 tormesolanes 39 and new rearranged labdanes with an aromatized ring B, such as fregenedanes 40,41 and isofregenedanes, 42,43 are found. The biosynthesis of these new carbon skeletons, as valparanes and tormesanes, contrary to the halimane diterpenoids, starts with a cyclization involving the pyrophosphate, so in this plant two different classes of diterpene cyclases should be present.
Owing to the high number of rearranged ent-labdane derivatives isolated in Halimium viscosum and in order to simplify their nomenclature and to facilitate their classication, the name ent-halimane was proposed for that carbon skeleton. Compounds of this class were isolated previously for the rst time in the roots of Adenochlaena siamensis, chettaphanin I and chettaphanin II (Fig. 1), with unknown absolute conguration.
To the best of our knowledge, there are no reviews of known halimanes. Owing to the important biological properties that some of them show, and their novelty and structural diversity, we considered it to be very interesting to establish a classication and write a review on these compounds.

Sources of halimane diterpenoids
Halimane diterpenoids form a group of secondary metabolites that can be found in different plant species of several families and in other taxonomic group, such as marine organisms and microorganisms ( Table 1).
The majority of known halimanes have been isolated from Magnoliophyta (Dicotyledon) plants and among them halimanes can be found in nine orders and curiously only in eleven families. It looks like, as occurs in the clerodane diterpenoid family, that the orders and families that contain halimanes are not very numerous. That is, they do not follow the usual taxonomic tendency in which a pyramidal form will be found when going from order to genus. 14 Among the families that possess halimane diterpenoids, we can highlight the Compositae, Lamiaceae and Euphorbiaceae families owing to the number of studied species and the number of halimanes found in them. In Cistaceae, it is noteworthy that all the Cistus studied until now, as can be observed in the reviews of diterpenes by Prof. Hanson, 6 halimanes have only been found as minor compounds in one of them (Cistus laurifolius). In contrast, the other studied plant of that family (Halimium viscosum) is the species that has provided a major number of halimanes.
The occurrence of halimanes in non-dicotyledon plants (with only seven species studied), in eleven marine organisms (highlighting the sponges), and in three strains of bacteria is much reduced. However, the interest in halimane diterpenoids is denitively much greater because of the presence of halimane purines with important biological activities among them. Besides the mentioned tuberculosinyl adenosine derivatives isolated from Mycobacterium tuberculosis, eleven other halimane purines, mainly isolated from marine sponges, have been characterized. They could constitute a very interesting area for further biological and chemical research owing to their biological activities.
Herein, an up-to-date review of the 246 isolated and characterized natural halimanes is shown.

Biological activities of halimanes
The halimane purine derivatives present the most relevant biological activities related to tuberculosis biomarkers, antifouling and antimicrobial activities.
1.2.1. Biomarkers for tuberculosis. Another illness closely related with some halimane type compounds is tuberculosis. Tuberculosis, which is mainly caused by Mycobacterium tuberculosis, is a major source of morbidity and mortality worldwide, with almost two million deaths annually. 60 New drugs for the treatment of tuberculosis are necessary because of the worrying increase in the multidrug and extensively drug resistant strains. 60,64 Nowadays, maximum interest is focused on the development of new therapies that target virulence factor (VF) formation. VFs by denition are not essential for bacterial growth outside the host cell but are involved in processes such as invasion, persistence, lysis and evasion of innate immune system responses. Compounds such as tuberculosinol and isotuberculosinol can be considered VF in M. tuberculosis. 60,61 Currently, two genes (Rv3377c and Rv3378c) found only in virulent species of genus Mycobacterium (such as M. tuberculosis and M. bovis) are known and they are involved in tuberculosinol and isotuberculosinol biosynthesis. Interestingly, these genes could not be found in avirulent species of genus Mycobacterium (such as M. smegmatis and M. avium), 59,65 so they may be involved in the infection processes of these bacteria. However, these genes only seem to be functional in M. tuberculosis and not in M. bovis, so that may explain the lower virulence of M. bovis in comparison with M. tuberculosis. 66,67 It has been observed that tuberculosinol and isotuberculosinol ( Fig. 1), produced in vivo by Mycobacterium tuberculosis 68,69 (in a 1 : 1 ratio), inhibit phagolysosome maturation as well as macrophage phagocytosis, plus a synergistical effect increased by the coexistence of both compounds has been observed. Decrease of the phagocytic capacity could help to explain the pathogenicity of M. tuberculosis. 59 Thus, both tuberculosinol/isotuberculosinol biosynthetic proteins (Rv3377c and Rv3378c) are essential for the bacteria's survival inside the macrophage. 57,59,70 Because of that, both enzymes are likely to be new potential targets for the development of new drugs.
Recently, two new natural tuberculosinol derivatives (having an adenosine unit attached at diterpene C15) have been isolated from M. tuberculosis, 1-TbAd 71 and N 6 -TbAd 62 (Fig. 1). In a comparative lipidomics assay between M. tuberculosis and M. bovis, it has been observed that these two compounds appear in higher amounts in M. tuberculosis and they accumulate to comprise >1% of all M. tuberculosis lipids, so they could serve as an abundant chemical marker of M. tuberculosis. 62,71 In addition, in this study it has been proved that the Rv3378c enzyme is responsible for 1-TbAd formation, so this protein appears to be a tuberculosinyl transferase (prenyl transferase). 71 In fact, they are being evaluated as biomarkers for tuberculosis. 62 Owing to all mentioned above, Rv3377c and Rv3378c are new targets for anti-infective therapies against tuberculosis that block virulence factor (tuberculosinols) formation. 60 1.2.2. Halimane purines bioactivity. The halimane diterpenoids found in marine organisms, such as Porifera, sponges of the genus Agelas and Raspailia, are characterized as halimane purines. Several of these compounds show antibacterial activity. It is interesting to point out that these compounds are structurally similar to the diterpene purines isolated from Mycobacterium tuberculosis (1-TbAd and N 6 -TbAd) although in these ones the purine appears glycosylated and the union with the diterpene is different. It is probable that the biosynthesis of these compounds follows a similar path to 1-TbAd and N 6 -TbAd, in which enzyme homologs to Rv3378c could be involved, probably expressed in the genome of the surrounding microbiome around the macroorganism.
Nosyberkol (isotuberculosinol) was isolated for the rst time from Raspailia sp. and the sponge contains halimane purines too. 72 Recently there has been enormous interest in the bioactivities of diterpenyl purines. 73 From Agelas sp., 74 nine halimane purines have been isolated that show antibacterial, antifungal, antimalarial, cytotoxic activities, inhibition of adenosine transfer rabbit erythrocytes and Ca 2+ channel antagonistic action and a 1 adrenergic blockade, among others. Some of these compounds possess antifouling activity against macroalgae. Natural products with this activity are very useful in the shing industry as an alternative to the antiadherent mixtures that include metals with toxic effects on the marine environment. For this reason, antifouling substances with no or reduced toxicity must be discovered or developed. 75 1.2.3. Other bioactivities. Antitumour halimane diterpenoids against several cell lines 45,48,52,76 (pancreatic adenocarcinoma, human colon carcinoma, bladder, lung and cervix cancer) have been isolated from plants and marine organisms of genus Echinomuricea, Agelas and Raspailia. Several halimane derivatives with antiangiogenic 44 activity and a topoisomerase inhibitor 77 have been described.
Other halimanes show interesting biological activities, such as antibacterial, 50 antiviral, 78 antifungal, 53 and antimalarial. 79 Others exert anti-inammatory, 45 anti-ulcerogenic, 80 antihyperlipidemic 81 or hepatogenic activities. Some halimane diterpenoids act as allelochemicals, regulating the growth of monocotyledon seeds. 54 Others behave as allomones acting against insects as repellents or mosquitocidals. 82 From the Antarctic nudibranch a series of allomones has been isolated, including diterpene glycerides, which seem to be involved in the defense of those nudibranchs. 83 Many of the known halimane diterpenoids have not been biologically evaluated.

Biosynthesis overview
Halimane skeleton diterpenoids are formed by cyclization of geranylgeranyl diphosphate (GGPP), catalyzed by class II diterpene cyclases (DTCs). 12,13,16 These enzymes are characterized by having an aspartate-rich DXDD motif and are differentiated between class I terpene synthases (TPS) enzymes by not having the characteristic aspartate-rich DDXXD motif that binds divalent metal ions required for catalysis of diphosphate ionization 11 (Scheme 2). Several studies have been carried out revealing that the 'middle' aspartate in the DXDD motif acts as an acid catalyst. 11,84 The DTCs catalyze GGPP bicyclization by a general acid-base mechanism. The process starts by 14,15 double bond protonation of E,E,E-GGPP followed by carbon-carbon double bond anti addition (C10 on C15, then C6 on C11) to give four possible bicyclic products. Effectively, depending on the different prochiral substrate conformer (1, 2, 3 or 4) operating in the cyclization, the corresponding stereoisomer of labda-13-en-8-yl diphosphate intermediate will appear (5, 6, 7 or 8).
Those halimenyl diphosphate cations 13-16 can evolve by different ways (Scheme 3). Each one can lose a proton providing the halimadienyl diphosphates 17 or 18, or rearrange by hydride shi to form the halimenyl diphosphate cation 19 followed by proton abstraction to give the halimadienyl diphosphate 20. The different diastereoisomer forms (21)(22)(23)(24), (25)(26)(27)(28) and (29)(30)(31)(32) correspond to the halimadienyl diphosphates 17, 18 and 20, respectively (HPP, syn-HPP, ent-HPP, and syn-ent-HPP; where syn prex refers to cis conguration between H8 and C20). These diphosphates are the precursors of nearly all known halimanes. As has been indicated previously, the nal cations 19 usually evolve by deprotonation yielding halimadiene skeleton compounds. However, water addition to the previously cited cations can occur, producing hydroxylated derivatives, 85,86 such as the dihydrohalimene 33, thus leading to formal designation of the relevant class II DTCs as hydratases. 86 Otherwise, although 1,2-hydride and/or methyl shis do not appear to be concerted, 16 the clerodane skeleton is formed from intermediate 19 by methyl group migration (C4 to C5 methyl migration giving trans and cis clerodanes) and later stabilization, thus generating the more than 1300 compounds that make up the clerodane diterpenoids family. 14,15 The formation of 3-secohalimenyl derivatives, such as 35 (Scheme 4), can be explained by oxidation of the C3 position of a halimenyl derivative by some specic oxygenase, such as cytochrome P450 monooxygenase. 87 It is notable that plants have vastly expanded numbers of cytochrome P450 monooxygenases in their genomes 88 providing a ready source of potential downstream acting enzymes. Studies have proved that cytochrome P450 takes part in the biogenesis of a considerable number of tricyclic and tetracyclic diterpenes, 16,89 oxidizing these compounds at C3. So, we cannot discard the oxidation of any biogenetic intermediates of 35, achieving a hydroxy derivative like 34. For the formation of intermediates such as 34, in other families of compounds, it has been speculated with the epoxy derivative participation, 90 as the GGPP oxide, although in diterpene biosynthesis that intermediate has not been detected until now in nature.
Finally, oxidation with rupture of the C3-C4 bond leads to the wide group of 3-secohalimenyl derivatives.
Rearranged halimanes of different types (I-V) are known (Fig. 2). Each one can be formed by rearrangement of some intermediate cations of other halimanes. In fact, rearranged halimanes have been obtained through biomimetic synthesis starting from natural halimanes. 91 Type I rearranged compounds should be formed by ring B expansion (Scheme 5). Oxidation of 27 (5(10),13-ent-HPP) at C11 gives 36, the precursor of an intermediate carbocation 36a that facilitates ring B expansion through an intermediate such as 36b, which nally could be stabilized by double bond formation between C9 and C11.
Type II and III derivatives could be formed by expansion/ contraction of the decaline annular system that arises from an intermediate diketone 37a, which will be formed by oxidation of the double bond D 5(10) through the intermediate 37, as it is shown in Scheme 6.
Type IV derivatives are compounds that can be considered as rearranged halimanes or rearranged clerodanes because C19 is bonded to C4 and C5 forming a cyclopropane ring, and type V derivatives can also be considered rearranged halimanes or clerodanes because C19 has been included in ring A as a consequence of a ring expansion, while other biosynthetic pathways cannot be discarded (Scheme 7). Possibly, the relevant class II diterpene cyclase might promote the cyclopropanation or the ring A expansion as it occurs with the ring contractions in the biosynthesis of permutilin. 92

Enzymatic and genetic experimental studies
A bacterial class II (B type) DTC that produces halima-5,13-dienyl diphosphate (29) has been identied. 16,55 Effectively, Mycobacterium tuberculosis Rv3377c gene has been identied, and the encoded diterpene cyclase has been proved to be responsible for the production of the halimane skeleton (Scheme 8).
Recently the structure of the diterpene tuberculosinol/ isotuberculosinol synthase (Rv3378c) from Mycobacterium tuberculosis has been reported. 60 The biosynthesis of tuberculosinols is catalyzed by two enzymes: Rv3377c, tuberculosinyl (halima-5,13-dien-15-yl) diphosphate synthase, and Rv3378c, tuberculosinol/(R/S)-isotuberculosinol synthase. Both proteins are essential for the bacteria survival inside the macrophage where tuberculosinols inhibit phagolysosome maturation as well as macrophage phagocytosis. 57,59,70 Rv3377c is a DTC classied as class II that transforms GGPP into tuberculosinyl diphosphate (TPP) with a halimane core 55,93 (Scheme 8), while Rv3378c is a diterpene synthase that converts TPP into tuberculosinol or (R/S)-isotuberculosinols acting as a phosphatase/isomerase. 57,59 However, although these two enzymes are sufficient to generate tuberculosinol and isotuberculosinol from GGPP, some studies on the evolution and functional characterization of the biosynthetic operon where these two genes are found have been carried out. 94 Recently, a previously unknown type of diterpene-nucleoside, 1-tuberculosinyl adenosine (1-TbAd), was isolated and characterized (Scheme 8). This discovery leads us to consider a reviewed biosynthetic model in which the Rv3378c protein is not a simple phosphatase as currently believed, but that the enzyme acts with combined phosphatase and tuberculosinyl transferase functions by using adenosine as nucleophilic substrate. 71 Interestingly, the Rv3377c and Rv3378c genes are found only in virulent Mycobacterium species, and not in avirulent ones. 59,66 Another work reported that these genes are only functional in M. tuberculosis, despite being present in other less virulent species, such as M. bovis. 67 Recently the incubation of [16,16,16-2 H 3 ]GGPP with tuberculosinyl diphosphate synthase (Rv3377c) from M. tuberculosis allowed the stereochemical course of the cyclization reaction to tuberculosinyl diphosphate via chair, chair transition state to be followed, conrming the cyclization pathway. 95 New studies with class II DTCs have been carried out where the enzyme has been modied in order to check the fundamental role of the DXDD motif (Scheme 9). 11 The high importance of that motif was conrmed and even a single residue modication in that motif can disrupt the normal activity of the enzyme. If the H501 residue is substituted in the rice (Oryza sativa) syn-copalyl diphosphate synthase OsCPS4 forming the mutant OsCPS4-H501D, the rearrangement of the initially formed bicycle is produced, obtaining the novel compound syn-HPP (Scheme 9), whose dephosphorylated derivative structure was characterized and its conguration established by NMR. 85 Recently, Zerbe and co-workers, 96 in a work guided to achieve bioactive natural products harnessing the plasticity of these class II diTPS, have realized mutagenic experiments with the horehound (Marrubium vulgare) class II diTPS peregrinol diphosphate synthase (MvCPS1) (Scheme 10). Two double mutants based on the combination of F505 and W323 (W323L : F505Y and W323F : F505Y) produce the rearrangement of the labda-13E-en-8-yl + intermediate, yielding the halimane skeleton instead of water capture giving the hydroxylabdane derivative.
Biomimetic rearrangements of simplied labdane diterpenoids have been carried out by treatment with a variety of Lewis and protic acids, demonstrating that those rearrangements involve a series of stereospecic 1,2-alkyl and hydride shis, producing mainly halimanes or a mixture of different dehydration products depending on reaction conditions. However, further rearrangement to clerodane products was not observed, indicating a high degree of enzymatic control for the in vivo formation of these natural products. 97 In this work it was shown that the halimane skeleton appears to be inherently more stable than the clerodane structure, as Peters, Tantillo and co-workers demonstrated in their study of quantum chemical calculations. 85,98 In many of these plants that contain halimanes, labdanes have also been isolated and in other plants they coexist with clerodanes. For example, in a Cistaceae such as Halimium viscosum ent-halimanes and labdanes coexist, 17,26,28,99,100 in Cistus laurifolius labdanes, ent-labdanes, cis-clerodanes and ent-halimanes have been isolated, [101][102][103][104] and in plants of genus Croton 44,105-107 (Euphorbiaceae) labdanes, halimanes and clerodanes coexist. In Haplopappus paucidentatus 25 and Nardophyllum lanatum 19 (Compositae) labdanes, halimanes and clerodanes coexist. It seems that in Halimium viscosum the biosynthetic route to clerodanes is enzymatically interrupted.

Classification
In this review, halimane diterpenoids have been classied according to the endocyclic double bond position and to their corresponding dihydro derivatives. Seco-, nor-and rearranged halimanes have been considered too (Fig. 3).
To the best of our knowledge, all examples of seco-, nor-, and rearranged halimanes (groups 5 and 6) known nowadays belong to the 'enantio' series, except for four tetranorhalimenes and three rearranged halimanes.

Natural halimanes
In this section the structures of natural halimanes can be observed; each one is numbered and accompanied by its trivial name. In addition, we summarize the information in tables that appear in the S.I., including the source of isolation, the plant part or organism from which they were isolated, the biological activities and references (Tables S1-S6 †).
The halimane structure elucidation has been carried out by extensive spectroscopic techniques, mainly NMR and chemical correlation. X-ray structural analysis has been carried out for several of them, as indicated in the comments of the diverse groups that appear aerwards. The absolute conguration has been established by circular dichroism. However, in some cases, mainly due to the scarcity of natural product, it has not been possible to determine the stereochemistry.
The gures and tables will show the natural products as they have been described and characterized; in most cases the carboxylic acids appear as their respective methyl ester derivatives.

Halim-1(10)-enes group
The rst known halimanes are found in the group of halim-1(10)-enes, which is the most numerous class so we start the classication from them.
In literature, the rst halimanes were designated as rearranged labdanes, isolabdanes or friedolabdanes, but in order to classify them in a diterpene group, it was decided to name them as halimanes due to the high number of these compounds that appear in plants of genus Halimium. 17 The study of genus Halimium plants has made possible to determine the presence of ve chemotypes of Halimiun viscosum, known in accordance with the harvesting place: Villarino de los Aires, (Salamanca, Spain), 17 La Fregeneda, (Salamanca, Spain), 26,27 Valparaiso, (Zamora, Spain), 28 Celorico da Beira, (Portugal) 29 and San João da Pesqueira, (Portugal). 30 In this group, the Euphorbiaceae, 78,108,109,[111][112][113][114] Cistaceae, 17,26,27,29,30,110,[115][116][117][118] Leguminoseae, 48,119,120 Compositae, 21,24 Jungermanniaceae, 121 Velloziaceae 52 and Annonaceae 122,123 plant families have been studied together with marine organisms of genus Spurilla 124 and Agelas. 74,125 The most numerous compounds of this group (Fig. 4) are those that show a carboxylic function at C18 and among them the most frequent possess an unsaturated or polyfunctionalized side chain. However, compounds 77 and 78 are the only ones that have an oxygenated function at C19. Compounds with the saturated side chain are abundant too, nding among them those oxidized at C2.
The absolute conguration of 61 was determined by circular dichroism (CD) and the structure of 59 was corroborated by Xray. These two compounds were isolated from Hymenaea courbaril. 48 Only eight compounds 79-86 do not have functionalized C18 or C19. Recently, spurillin B (81), isolated from Spurilla sp (Nudibranchia), has been described, and it is one of the few enthalimanes with a cis double bond in their side chain. 124 8 0 -Oxoagelasine C (86) is a novel purine diterpene that was recently isolated from Agelas nakamurai and it is the only halimane purine presenting a carbonyl group at adenine C8. 125 Compounds of this type, usually known as agelasines, have been isolated from genus Agelas sponges and are very interesting owing to their antimicrobial and antispasmodic activities and their action as Na,K-ATPase enzyme inhibitors. 126 Although this group is quite numerous, only six furohalimane derivatives (38, 63-66 and 75) and a halimanolide (85) are known. These functionalizations are most frequent in the ent-halim-5(10)-enes, as we will explain later. Crassifoliusin A (63) 113 and crassin D (64), 114 isolated from Croton crassifolius, have a tricyclic system formed by cyclization of C1 with C12 of the halimane side chain.
A lot of these compounds have been isolated from plants used in folk medicine, but the biological activity of many of them has not been determined yet. Only 59 48 and 84 52 have proved to be active as antitumour drugs.
4.1.2. Halim-1(10)-enes. Only three compounds of this kind are known: 87, 88 and 89 (Fig. 5, Table S1 †). Charruoic acid (88) is the only halim-1(10)-ene showing a D 7 in ring B and was isolated from Ophryosporus charua. 127 The structure of the natural product agelasine C (89) was established by its enantiomer synthesis 128 correcting, in this manner, the original structure proposed by Nakamura and coworkers. 126 Agelasine C (89) was isolated from Okinawan sea sponge Agelas sp. and Agelas citrina 129 and it exerts antifungal activity.
Echinohalimane A (99), which shows a g-hydroxybutenolide in the side chain, was isolated from a gorgonian identied as Echinomuricea sp. 46,47 It is the rst halimane isolated from a marine organism belonging to the phylum Cnidaria. This compound was found to exhibit cytotoxicity towards various tumour cell lines and displays an inhibitory effect on the release of elastase by human neutrophils.
As chettaphanin II (102), the majority of known ent-halim-5(10)-enes are furo-ent-halimanes. In some of them the furan fragment appears oxidized in the form of 15,16-butenolide. The lactone group can be observed in other positions, for example 20,12-olides. It is usual to nd derivatives of this kind with a carboxylic function at C18 or C19. The side chain C12 position usually appears functionalized and ring A can be oxygenated at C2 or C3.
The structure and absolute conguration of lactone 106 were corroborated by synthesis and it shows a moderate activity against HeLa cells. 44,78,112,141 Crotohalimaneic acid (107) and crotohalimoneic acid (108) show activity against several human cancer cell lines. 140 The structure of crotohalimoneic acid (108) was corroborated by X-ray.
A series of ent-halim-5(10)-enes known as crassifolins 112-117, 119-121, 123-124 and crassin C (125) has been isolated from Croton crassifolius. 44,78,144 The absolute conguration of several of them was conrmed by CD. The structure of crassifolin D (114) was corroborated by X-ray; however, its absolute conguration was not determined. As the crassifolins found in Croton crassifolius are included in the ent-halim-5(10)-ene series, the same absolute conguration is proposed for crassifolin D (114) and it was included in this group. Crassin C (125) 114 is characterized by its ring A contraction, showing a carboxylic function at C1. This compound could be the result of an oxidation followed by condensation, of any compound that coexists in that plant with functionalization at C2, forming in this manner a [5.6]    Functionalization at C7 is seen in compounds 128, 134 and 140 and at C8 only on compound 140. 76 Crassifolin A (112), crassifolin B (113), and penduli-aworosin (126) showed anti-angiogenic activity using a wildtype zebrash in vivo model, 44,78 and crassifolin D (114) shows antiviral and anti-angiogenic activities. 44 Crolaevinoids A-B (128)(129) and crothalimene A (130) were recently isolated from Croton laevigatus and Croton dichogamus, respectively. 105,148 They are the only C17 functionalized derivatives known in this class forming a d-lactone with carbon C12. The absolute conguration of compounds 128 and 129 was determined by electronic circular dichroism (ECD). These compounds exhibit pronounced inhibition of nitric oxide (NO) production. 105 3a-Hydroxy-5(10)-didehydrochiliolide (139) was highly active against a human pancreatic adenocarcinoma cell line at micromolar concentrations 45 and 140 shows antitumour activity. 76 Formosin A-C (135-137) isolated from Excoecaria formosana were active as antimicrobials, 147 and, in addition to crassifolius B (138), 150 they are the only compounds that possess a carboxylic acid or methyl ester functionality at C20.
In six of the known ent-halim-5(10)-enes 141-146, no furanic or lactone systems appear in the side chain. Isoscoparin N (143) and isoscoparin M (147), 91,155 together with crolaevinoid A (128), are the only ones that show oxygenated functions at C11. The structure of compound 146 has been proposed but its absolute conguration has not been solved. 153 Compound 145 shows antitumour activity. 154 In this group, the halimane purine agelasine J (148) is included. Agelasine J was isolated from the Solomon Islands marine sponge Agelas cf. mauritiana. It shows antimalarial and antimicrobial activity and MCF7 cell cytotoxicity. 79 isolated from Dysoxylum densiorum 156 and 152 and 153 isolated from Amphiachyris amoena 157 possess functionalization at C3. Allylic groups, as furyl or butenolide derivatives, appear in the unsaturated side chain, except 149, isolated from a Dominican amber, 158 which shows a saturated side chain with a carboxyl group at C15. Derivatives with oxygenated functions at C19 appear esteried with p-hydroxyphenylpropionic acid, amoenolide L (152) and amoenolide M (153). 157 These last compounds include oxygenated functions on ring B at C6. Recently, crassifolius A (154) and crassifolius C (155), from Croton crassifolius, have been characterized and their absolute congurations established by ECD. 150 In this class, two halimane purines isolated from an Okinawan marine sponge Agelas sp. can be found: agelasine O (156) and agelasine S (157). 53 Agelasine O (156), in which C18 is esteried with 2-carboxy-4-bromopyrrole, is biologically active as an antibacterial and antifungal. 53 165 and Marrubium aschersonii 169 166 ) except 158, which was isolated from Stevia gilliesii 167 (Compositae). The most usual derivatives in this series are those that present oxygenated functions on ring B, normally on carbons C6 and C7 (Fig. 9, Table S3 (172), isolated from Cistus laurifolius 102 (Fig. 11, Table S2 †). Their structures were spectroscopically determined and their absolute congurations assigned by chemical correlation by comparison with rearranged products of labdanolic, populifolic or ent-halimic acid. 102,168 The ve compounds of this type are functionalized at C3. Salmantic acid (170), its methyl ester (171) and salmantidiol (172) have the side chain saturated, while leucasperone A (173) and leucasperone B (174), isolated from Leucas aspera, 169 19 and Nardophyllum bryoides 45 ). Acids 177 and 178 were isolated as their methyl esters and their epimers at C-13 separated, although assignment of their stereochemistry at C-13 was not possible. 20 Salicifolic acid (179) regulates the growth of Panicum miliaceum (monocotyledon) seedlings 54 and 3a-hydroxy-5,6didehydrochiliolide (181) was highly active against a human pancreatic adenocarcinoma cell line at micromolar concentrations. 45 4.3.2. Halim-5-enes. This is the biggest halimenes group (182-201) with a double bond at C5 (Fig. 13, Table S3 †) and perhaps the most interesting group considering the bioactivity of its derivatives, because tuberculosinol (182) 55 and isotuberculosinol (also known as nosyberkol; 183) are found among them. [56][57][58]72 Nosyberkol was isolated for the rst time in 2004 from the Nosy Be Islands (Madagascar) sponge Raspailia sp. as a single stereoisomer. 72 However, the stereochemistry at C-13 of natural nosyberkol has not been determined.
It has been proved that compounds tuberculosinol (182) and isotuberculosinol (183) are produced by Mycobacterium tuberculosis. 68,69 None of the studies done detected the presence of 182 or 183 from the cultured cells of 12 nonpathogenic Mycobacterium species. 65 It has been observed that tuberculosinol (182) and isotuberculosinol (183) (in a 1 : 1 ratio, with 183 being a mixture of the diastereomers 13R-isotuberculosinol (183R) and 13S-isotuberculosinol (183S) in a 1 : 3 ratio) inhibit phagolysosome maturation and macrophage phagocytosis in human-like cells. 59 The structures of tuberculosinol (182) and isotuberculosinol (183) have been corroborated and their absolute conguration established by total synthesis. 56,58 Recently, two new natural products derived from tuberculosinol have been isolated and characterized: 1-TbAd (200) 71 and N 6 -TbAd (201). 62 These two tuberculosinol derivatives possess an adenosine unit bonded at C15 by N1 0 or by the nitrogen at C6 0 of the adenosine (Fig. 13). Recently it has been observed that compounds 200 and 201 accumulate to comprise >1% of all M. tuberculosis lipids. These diterpene nucleoside compounds are being investigated as biomarkers for tuberculosis. 62 The structures of 1-TbAd (200) and N 6 -TbAd (201) have been corroborated by total synthesis. 63 In the halim-5-enes series (Fig. 13), the side chain can be saturated or unsaturated and furans or functionalized butanolides can be found on it.
Tuberculosene (184) has been obtained by enzymatic reaction from a mixture of GGPP with tuberculosinyl diphosphate synthase and CYC2 enzyme from the bacteria Kitasatospora griseola. 173,174 In this group, two plants of the Compositae family have been studied. From Koanophyllon conglobatum 175 koanophyllic acids 185, 192, 196 and 197, with carboxylic function at C18, were isolated, and from Haplopappus pulchellus 23 [186][187][188][189][190][191] were isolated, all of them with a saturated side chain. The structure of 195 was spectroscopically determined and its absolute conguration established by ECD 51 of their 3-p-bromobenzoate derivatives. This compound possesses antimicrobial activity. From Acalypha macrostachya the 7-oxo derivatives 193 and 194 were isolated. 22 Micromonohalimanes A and B (198 and 199, from Micromonospora sp. 50 ), which present antibacterial activity, have been characterized. Micromonohalimane B (199) is the only halimane which includes a chlorine atom in its structure.

Dihydrohalimenes group
A structural characteristic of this group of compounds (208)(209)(210)(211)(212)(213)(214)(215)(216)(217)(218)(219) is that all of them show oxygenated functions at C5, except diasin (219, from Croton diasii), 176 which we include in this group precisely for not having any unsaturation in the decalin system (Fig. 15, Table S4 †). Although some members possess acyclic chains (208 and 209, isolated from Pleurozia gigantea 18 and Jungermannia truncata, 177 and Baccharis salicifolia, 54 respectively), the most usual functionalization is the furan (210 and 211 from Dysidea amblia, 178 178 being the only occasion that halimanes and 8epi-halimanes coexist in the same organism. Originally, a cisfused bicyclic ring system was assigned to ambliol B (210), but nally the structure was revised by X-ray analysis. 178,181 Compound 209 is a germination inhibitor 54 and the structure of compound 214 was corroborated by synthesis. 182 Cracroson D (212) shows a new pentacyclic scaffold. It is chemically related to chettaphanin I (38) because 212 is generated from 38 by an intramolecular [2 + 2]-photocycloaddition. The existence of this compound in the extract as a natural product is conrmed by HPLC-MS. Cracroson D (212) exerts moderate cytotoxicity against T24 and A549 cell lines (bladder and lung cancer respectively).
In this group, halimane-purines such as agelasimines A and B (217 and 218, respectively), isolated from Agelas mauritiana, 183 are included. In this case, halimane C15 is bonded to purine N7. Both compounds show a wide range of interesting biological activities, such as cytotoxicity, inhibition of adenosine transfer into rabbit erythrocytes, Ca 2+ channel antagonistic action and a 1 adrenergic blockade.

Secohalimenes and norhalimenes group
All secohalimenes known 220-232 (Fig. 16, Table S5 †) are included in the ent-halimenes series, and are formed by cleavage of the C3-C4 bond. All of them show a furan unit in the side chain, except for 231 179 that contains a butenolide in that chain. Frequently C20 is a carboxylic acid that lactonize with a hydroxyl group at C12, 225-230, 19,45 or with a hydroxyl group at C5, 231. C3 is always a carboxylic acid (free, lactonized or esteried) except for 229 and 230. C4 is part of a disubstituted olen (D 4(18) ) 231-232 or tetrasubstituted one (D 4 ) 220-230.

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Compounds such as isoteuin (233; isolated from Croton crassifolius 144 and Teucrium canadense 185 ), teupolin VIII (234; Teucrium polium 186 ), teucvin (mallotucin A; 235; Teucrium viscidum, 187,188 Mallotus repandus, 136 Teucrium chamaedrys, 189 and Croton jatrophoides 190 ) and crotoeurin B (236; Croton euryphyllus 191 ) belong to a norderivatives group and have always been considered as clerodanes. However, these compounds could be considered norhalimanes as well, since up to now there is no biosynthetic evidence regarding the exact moment at which C19 (or C18) is lost. In any case, a table collecting all norclerodanes (norhalimanes) known to date has been elaborated as a ESI † (Fig. S15-S16, Table S7 †). In the table the natural sources of these compounds are presented, as well as their bioactivities and the concerning references.

Rearranged halimane group
The known rearranged halimanes are shown in Fig. 17 (Table  S6 †). They can be classied into four different groups.
Scopariusins A-C (246-248), isolated from Isodon scoparius, 91,155 are characterized for ring B expansion, by incorporation of C11. The structure and absolute conguration of scopariusin A (246) were conrmed by biomimetic synthesis 91 from isoscoparin N (143). These compounds were not active as antitumour agents.
Compounds 255-257 have butenolides or hydroxybutenolides in the side chain. Methyl dodonates A-C (258-260) have C18 functionalized with a methoxycarbonyl group, and they show a furan at the side chain, plus a double bond D 2 . Asmarines I-J (261 and 262) are diterpene-adenine compounds with an 8-epient-halimane skeleton bonded by C15 and C13 to adenine nitrogens N9 and N10. These compounds show cytotoxic activity.
Compounds 263-283 can also be considered rearranged halimanes or clerodanes because C19 has been included in ring A as consequence of a ring expansion. All of these compounds belong to the 'enantio' series except viterofolins A-B (253-254) and scapanialide B (283), which belong to the 'normal' series.

Halimane diterpenoids: synthesis and transformation into bioactive compounds
In the following paragraphs, several natural halimane diterpenoids synthesis and transformations of some of them into bioactive compounds are described.

ent-Halimic acid as precursor of biologically active compounds and other derivatives of interest
ent-Halimic acid, characterized as its methyl ester (39), is very abundant in Halimium viscosum extract (0.34% with respect to the dry plant weight). We have developed a very quick and efficient method to isolate the natural product in multigram quantities by chromatographic separation of the ethyl acetate extract. This compound has an unsaturated side chain, D 13 , and a hydroxyl group at C-15, a carboxyl group at C-18 and a double bond D 1(10) in the decalin system. All these functionalities make ent-halimic acid a versatile molecule and a very appropriate starting material for the synthesis of natural halimanes, biologically active compounds and other interesting compounds. In Fig. 18 some of the compounds obtained from ent-halimic acid are shown: (1) ent-Halimanolides. 123,141,207,208 (2) Chettaphanin I and II. 109,209 (3) Bioactive sesterterpenolides. 123,210-212 (4) Sesterterpenolides and glycerophospholipids hybrid compounds. 213 (5) Rearranged derivatives: ent-labdanes, 214 abeopicrasanes 215 and propellanes. 216 (6) Sesquiterpene-quinone/hydroquinone. 217 (7) Sesqui-and diterpene-alkaloids. 128,[218][219][220][221][222] Synthesis from ent-halimic acid methyl ester (39) of several natural halimanes, such as ent-halimanolides, chettaphanin I and II and agelasine C (a diterpene-purine derivative), will be commented on in the following points. These syntheses have made it possible to corroborate their structures.
The synthesis of intermediate 285 requires the reduction of the C18 methoxycarbonyl to a methyl and two carbon degradation of the side chain. First of all, the C15 hydroxy group is protected as its methoxy derivative and the resulting compound is treated with LAH followed by TPAP oxidation to obtain 284. Huang-Minlon reduction, followed by chemoselective oxidation and cleavage of the D 13 double bond with m-CPBA and periodic acid gives the required ketone 285 in very good yield (57% six steps).
Bestmann methodology 223 has been used for the synthesis of butenolides in similar systems. In order to apply it, functionalization of C16 as a hydroxy group is necessary. The synthesis of the g-hydroxybutenolide 132 has been done by Boukouvalas methodology. 224 Treatment of 85 with LDA and TBDMSTf followed by reaction of the intermediate 2-trialkylsilyloxyfuran with m-CPBA afforded 287 in good yield, aer column chromatography. Compound 132 was obtained in quantitative yield by acidic isomerization of 287 using HI in benzene at 85 C. The physical and spectroscopic data of the synthetic product 132 are identical with those reported for the natural product 16-hydroxy-ent-halima-5(10),13-dien-15,16olide. This synthesis conrms the structures and absolute congurations of the natural products obtained.
Biological assays have been carried out on these compounds and conrmed that compound 85 exhibits cytotoxic and antiviral activity [HeLaM cells (IC 50 ¼ 5.0), MDCK (IC 50 ¼ 5.1) and inuenza virus (IC 50 ¼ 6.8)]. 123 In the same way, an efficient synthesis of ent-halimanolide 106 (15,16-epoxy-12-oxo-ent-halima-5(10),13(16),14-trien-18,2bolide) has been achieved from ent-halimic acid methyl ester 39, corroborating the structure of the natural compound and establishing its absolute conguration (Scheme 12). 141 A new route employing the dinorderivatives 289 and 291 as intermediates allowed the tetranorderivative 292 to be obtained in multigram scale (53% from 39). ent-Halimic acid methyl ester (39) oxidation with OsO 4 was regioselective. The resulting triol was oxidized with Pb(OAc) 4 , giving ketone 288 in a 94% global yield for the two steps. reaction of 299 in acidic media led to isomerization of the double bond to the tetrasubsituted position, which, followed by deprotection of the primary hydroxyl group and oxidation gave 292 in three steps. Reaction of 292 with 3-furyllithium followed by oxidation gave the key intermediate 300 (36%, global). Chettaphanin II (102) was obtained by reaction of 300 in acidic media (92%) and chettaphanin I (38) was synthesized from 300 in two steps: epoxidation followed by reaction in acidic media (28%, two steps).
Agelasines are diterpene alkaloid 7,9-dialkylpurine salts, isolated from marine sponges of the genus Agelas. 225 Agelasine C is one of the rst four agelasines to be isolated by Nakamura and co-workers in 1984 126 from the Okinawan sea sponge Agelas sp. (À)-agelasine C showed powerful inhibitory effects on Na,K-ATPase and antimicrobial activities (Fig. 19). In their work, Nakamura and co-workers proposed the structural formula 303 for (À)-agelasine C. epi-Agelasine C (304) was isolated in 1997 by Hattori and co-workers 75 from the marine sponge Agelas mauritiana as an antifouling substance active against macroalgae.
Due to the interest of epi-Agelasine C as an antifouling agent 75 and to establish the absolute conguration of this compound, the synthesis of 302 (Scheme 14) was carried out. 128 The synthesis of 302 was planned following an analogue design for other agelasines, which consists of coupling the terpenic fragment 306 with a purine derivative such as 307. 128 The synthesis of the bromoderivative 306 was achieved in six steps starting from ent-halimic acid methyl ester (39) using the tetrahydropyranyl derivative 305 as an intermediate (Scheme 14) (46%, six steps). Reaction of 39 with DHP in p-TsOH followed by reduction with LAH leads to a hydroxy derivative that was oxidized with TPAP to the carbonyl function and reduced using Huang-Minlon methodology to give 305. Deprotection of the primary hydroxyl group followed by treatment with CBr 4 in the presence of PPh 3 gave the required bromoderivative 306 (46%, six steps). Alkylation of methoxyadenine 307 with the bromoderivative 306 by heating in dimethylacetamide and subsequent reduction with Zn/AcOH gave compound 302 (13%, two steps).
The physical properties of the synthesized product 302 were very different to those of the natural product epi-agelasine C, thus its proposed structure 304 (Fig. 19) should be revised. On the other hand, when the 1 H and 13 C NMR spectra of 302 were compared with those for the proposed structure of (À)-agelasine C (303), the two pairs of spectra were identical. However, the optical rotatory power of 302 and natural (À)-agelasine C were similar in absolute value but had a different sign. So, it should be concluded that the structure of the natural product (À)-agelasine C should be corrected to structure 89 (Fig. 20), which is the enantiomer of the synthesized product (+)-agelasine C (302, Scheme 14).
Spectroscopic considerations made when comparing the spectra of 302 with those of epi-agelasine C and their specic rotations permitted structure 100 to be suggested for the natural epi-agelasine C, as shown in Fig. 20.

Synthesis of 3a-hydroxy-5b,10b-epoxychiliolide (214)
The natural product structure of 3a-hydroxy-5b,10b-epoxychiliolide (214) was corroborated by the synthesis of the racemic diterpene (Scheme 15). 182 The synthesis starts with the available tetralone 308. Intermediate 309 was obtained in high yield by Mannich reaction, followed by hydrogenation of the resulting exo-methylene bond. In order to introduce the carboxyl group and achieve 310, an epoxidation followed by epoxide isomerization with BF 3 $Et 2 O was done. In this manner the corresponding aldehydes, epimers at C9, were obtained, which via Jones oxidation led to 310. Birch reduction of 310 followed by hydrolysis, esterication and protection as 1,3dioxolane, alkylation with LDA and allyl bromide and deprotection with diluted sulfuric acid on silica gel gave ketone 311 in 72% overall yield. Reduction of 311 with NaBH 4 followed by oxidation with osmium tetroxide/N-methyl morpholine-N-oxide and cleavage with sodium periodate gave aldehydes 312a and 312b, which were separated by column chromatography. From ketone 311, the derivative 313 was also obtained by oxidative degradation. Its X-ray analysis allowed the conguration of C8 and C9, quaternary centres, to be established for all compounds. Reaction of epimer 312 with 3-lithiofuran and subsequent lactonization afforded the intermediates 314a-d, and the epoxidation of each led to the isomeric diterpenes 315a-d. Conguration at C12 was established by nOe experiments, but the relative stereochemistry at C3, C5, and C10 still had to be solved. The NMR spectra for 315a was identical to the natural product one. Only one epoxide was obtained with both isomers, so a b-orientation of the epoxy group was more likely. PCC oxidation of 315a gives ketone 316, which gave one isomer by reduction with sodium borohydride, the 3b-hydroxy derivative 315c, so the natural product 214 is the 3a-hydroxy derivative 315a. Through inspection of a model, it was determined that the most favorable entry of the hydride is by the a-face of the molecule, so the relative conguration of all compounds was established in this manner.

Synthesis of (+)-agelasimine A (217) and (+)-agelasimine B (218)
Ohba and co-workers 226 synthesized the diterpene-adenine derivatives (+)-agelasimine A (217) and (+)-agelasimine B (218) from (+)-trans-dihydrocarvone and, in this manner, they established the absolute conguration of the natural products isolated from the orange sponge Agelas mauritiana (Scheme 16). Previously, Ohba and co-workers followed a similar reaction sequence when they communicated the racemic synthesis of (AE)-agelasimine A and (AE)-agelasimine B. 227,228 In the asymmetric synthesis, the authors used (+)-trans-dihydrocarvone (317) as the starting material. Ozonolysis in methanol of 317 and subsequent treatment with FeSO 4 -Cu(OAc) 2 (ref. 229 and 230) led to (+)-318, which was transformed into (+)-319 by MeLi treatment followed by PCC oxidation. 231 Reaction of (+)-319 with vinylmagnesium bromide in the presence of CuBr and Me 3 SiCl gave stereoselectively a silylenol ether that reacts with formaldehyde to give 320 as a diastereoisomeric mixture. Cyclohexanone 320 was transformed into 321 following the previously described procedure. 231 Methylation of 321 and subsequent Huang-Minlon reduction led to 322. Reaction of 322 with 9-BBN followed by Suzuki cross-coupling reaction with E-3-iodo-2-  butenoic acid ethyl ester (323) afforded the a,b-unsaturated ester (324). Treatment of 324 with m-CPBA, DIBAL reduction and subsequent treatment with LAH in boiling THF for epoxide reduction led to 325. Reaction of 325 with PBr 3 achieved bromination of the primary hydroxy group and subsequent alkylation with 3-methyladenine led to compound 326 aer neutralizing the hydrobromide salt. Methylation of (+)-326 with MeI followed by neutralization gave the desired compound (+)-217, which was identical to the natural product (+)-agelasimine A. Reaction of (+)-326 with NaBH 4 followed by methylation and neutralization led to (+)-218, which was identical to the natural product agelasimine B. In this manner, the two structures (217 and 218) were corroborated and their absolute congurations established.

Biomimetic synthesis of scopariusin A (246) and isoscoparin N (143)
Scopariusin A (246) was isolated from Isodon scoparius. Its structure was spectroscopically determined and conrmed by biomimetic synthesis from the clerodane isoscoparin O (327) found in the same plant (Scheme 17). 91 The structure and absolute conguration of isoscoparin O (327) has been conrmed previously by X-ray analysis and for scopariusin A (246).
Treatment of 327 with BF 3 $Et 2 O leads mainly to a halimane derivative, which leads to the hydroxy acid 328 by hydrolysis with NaOH in MeOH. Esterication of 328 with MeI in the presence of potassium carbonate gives 143, whose properties were identical to isoscoparin N isolated from the same extract of Isodon scoparius. Reaction of 328 with p-toluensulfonic acid (PTSA) in toluene and posterior methylation with MeI in KOHacetone led to 246, whose properties were identical to those of the natural product scopariusin A. In this manner, the structures and absolute congurations of isoscoparin N (143) and rearranged ent-halimane scopariusin A (246) were conrmed. (182) and isotuberculosinol (183) Tuberculosis, caused by Mycobacterium tuberculosis, is one of the biggest causes of morbidity and mortality worldwide. Developing new drugs effective against those bacteria and new therapies directed to inhibit virulence factor (VF) formation have a great interest. 60,63 The tuberculosinols tuberculosinol (182) and isotuberculosinol (183; 13R and 13S) are VFs from M. tuberculosis. 57,59,93 However, without any doubt the most interesting and promising halimane is tuberculosinol (182). The original proposed structures have been conrmed by synthesis, as can be seen in the following points. 56,58 Snider and co-workers' and Sorensen and co-workers' syntheses of tuberculosinol and isotuberculosinol (Schemes 18 and 19, respectively) were published simultaneously and made possible a structural revision of the diterpene obtained from M. tuberculosis, to which the edaxadiene structure was originally assigned, and nally it was revised and reassigned to the same structure of nosyberkol (183; isolated from the Red Sea sponge Raspailia sp. 72 extracts), also known as isotuberculosinol.

Synthesis of tuberculosinol
5.5.1. Snider's synthesis of tuberculosinol (182) and isotuberculosinol (183). The synthesis by Snider and co-workers 56 uses an exo-cycloaddition as a key step in the syntheses of isotuberculosinol (183) and tuberculosinol (182; Scheme 18). In this case, the cycloaddition step was done with 329 in the presence of N-tigloylisoxazolidinone (330) and Me 2 AlCl to afford a mixture of the desired exo Diels-Alder adduct 331 and the endo adduct (54%, $10 : 1 exo/endo). Reduction of 331 followed by oxidation with Dess-Martin periodinane gives exo aldehyde 332. Reaction of 332 with acetone in the presence of NaOMe leads to 333, which gives ketone 334 by reduction with Li in NH 3 /THF/EtOH, then by addition of vinylmagnesium bromide provides 183 as a mixture of stereoisomers. The spectroscopic data obtained from the synthetic compound 183 were identical with those reported for both natural nosyberkol and isotuberculosinol.
Reaction of ketone 334 with triethylphosphonoacetate in the presence of NaH leads to 335 in good yield. DIBAL reduction of 335 leads to 182, which is identical to the natural tuberculosinol. 55 5.5.2. Sorensen's synthesis of tuberculosinol (182) and isotuberculosinol (183). In the synthesis of tuberculosinol (182) and isotuberculosinol (183; Scheme 19) achieved by Sorensen and co-workers, 58 the key step is an exo-selective Diels-Alder reaction. Cycloaddition of the known diene 329 and ethyl tiglate followed by ester reduction and primary alcohols separation through purication by supercritical uid chromatography provides the enantioenriched material (>99% ee), that by Parikh and Doering 232 oxidation conditions leads to 332a. An aldol condensation with acetone sodium enolate and conjugate reduction with Wilkinson's catalyst 233 allowed them to achieve ketone 336. Vinylmagnesium bromide addition provides 183 as a tertiary alcohol mixture. The spectroscopic data obtained from the synthetic compound 183 were identical with those reported for both natural nosyberkol and isotuberculosinol.

Conclusion
In this article, the rst natural halimane skeleton diterpenoids review is reported. We have classied them into six different groups according to their biogenetic origin. Herein, 246 natural halimanes have been collected, summarizing their structure, natural source and bioactivity.
Among the halimane family, the major group corresponds to the 'antipode or enantio' series, as also happens in the labdane skeleton diterpenoids. In this manner, ent-HPP and syn-ent-HPP derivatives represent 70% (taking into account nor-, seco-, dihydro-, and rearranged halimanes of these series too) of all known halimanes.
The most interesting reported halimane with the most potential is tuberculosinol (182). The production of isotuberculosinol, tuberculosinol and analogues by M. tuberculosis inhibits the phagocytosis of human macrophage-like cells, thus they can be considered virulence factors (VFs). Recently, two tuberculosinyl adenosines have been isolated. These two natural halimanes, derived from tuberculosinol and isotuberculosinol, are being evaluated as possible biomarkers for early diagnosis of tuberculosis.
Some nor-and rearranged halimanes can be considered as 19norclerodanes or rearranged clerodanes, respectively, and vice versa. These structures can be classied indistinctly as clerodanes or halimanes without further information on their biosynthetic origin. For this reason, halimanes with a 19-norclerodane structure have been also reviewed and included in the ESI. † Different halimanes, such as ent-halimic acid (39) and 11Racetoxy-ent-halima-5,13E-dien-15-oic acid (175), have been used as starting materials for the syntheses of bioactive compounds, for example antibacterial or antitumoral compounds.

Conflicts of interest
There are no conicts to declare.

Acknowledgements
The authors would like to thank the Spanish Ministry of Economy and Competitiveness, MINECO (SAF2014-59716-R, CTQ2015-68175-R), European Funds for Regional Development, FEDER and Junta de Castilla y León (BIO/SA59/15, UIC21) and Universidad de Salamanca for nancial support. AMR and IET are grateful for their fellowships from the FSE (European Social Fund), MINECO and Junta de Castilla y León, respectively. The authors thank Profs. J. G. Urones and P. Basabe for their dedication as teachers, and their guidance and friendship over many years.