Totarol


Totarol is a naturally produced diterpene that is bioactive. It was first isolated by McDowell and Easterfield from the heartwood of Podocarpus totara, an endemic conifer species found in New Zealand. Podocarpus totara was investigated for unique molecules due to the tree's increased resistance to rotting. Recent studies have confirmed totarol's unique antimicrobial and therapeutic properties. Consequently, totarol is a candidate for a new source of drugs and has been the goal of numerous syntheses.

Discovery

Totarol was discovered in 1910 by New Zealand scientist Sir Thomas Hill Easterfield. While investigating the properties of Miro, Kahikatea, Rimu, Matai and Totara, Easterfield detected a "crystalline bloom" on totara boards a few hours after leaving the planing machine. After extraction of totarol from Podocarpus totara, Easterfield observed no other compound had been cited in chemical literature before with this formula. Easterfield and his colleague J.C McDowell proposed the name "totarol" in a follow-up paper in 1915, as the crystalline substance was believed to possess a tertiary alcohol group. In 1937 Short and Stromberg continued investigations, publishing Totarol Part 1. In 1951 Short and Wang became the first to identify the chemical structure of totarol with their paper Totarol Part 2.

Occurrence

Although totarol was first isolated in Podocarpus totara, totarol has also been identified in numerous other species of Podocarpaceae and Cupressaceae, with the majority found in the genus Podocarpus of the family Podocarpaceae and the subfamily Cupressoideae of the family Cupressaceae.
Outside Podocarpus and Cupressoideae, totarol is rarely found in the plant kingdom. However, totarol has recently been isolated in Rosmarinus officinalis. The gymnosperms that contain totarol are distributed worldwide but are concentrated in North America, the far-south regions of South America, East Asia and East Africa.

Biological activity

Antimicrobial activity

Totarol motivates research in drug discovery due to its ability to inhibit numerous microorganisms. Totarol exhibits antimicrobial properties in numerous species including gram-positive bacteria, acid-fast bacteria, nematodes, parasitic protozoans, crustaceous foulers. In addition to inhibiting microorganisms by itself, totarol exhibits inhibitory synergy with currently used antimicrobial drugs: totarol potentiates isonicotinic acid hydrazide against various Mycobacteria.; methicillin against Mycobacterium tuberculosis and Staphylococcus aureus; and anacardic acid and erythromycin against Staphylococcus aureus. In nature, totarol is a key player in gymnosperm's defense against harmful microbes: gymnosperms that produce totarol are resistant to rotting.
Table 1. Antibacterial activity of totarol against microorganisms
MicroorganismMIC IC50
Artemia salina-1
Bacterium ammoniagenes0.78-
Bacillus subtilis1.56-
Caenorhabditis elegans-80
Enterococcus faecalis2-
Klebsiella pneumoniae>32-
Mycobacterium aurum27.5
Mycobacterium fortuitum47.5
Mycobacterium phlei47.5
Mycobacterium smegmatis27.5
Mycobacterium tuberculosis H37Rv21.17.5
Leishmania donovani-3.5
Proprionibacterium acnes3.9-
Staphylococcus aureus ATCC 125981.56-
Staphylococcus aureus ATCC 335910.78-
Staphylococcus aureus ATCC 116320.78-
Streptococcus mutans0.78-
Streptococcus pneumoniae2-

Mechanism of antimicrobial inhibition

Although totarol exhibits antimicrobial properties, the mode of action is unclear and various methods of inhibitory action have been proposed. In Staphylococcus aureus strains resistant to penicillin via creation of penicillin binding protein 2', totarol may inhibit the synthesis of PBP2'. Totarol may inhibit effluxing Staphylococcus aureus strains through inhibition of MsrA, although it is unclear if MsrA is an efflux pump. Totarol may also gain its antibacterial properties by inhibiting bacterial respiratory transport but this is very unlikely because totarol is also effective against anaerobic organisms. Recently totarol was also hypothesized to inhibit gram-positive and acid-fast bacteria via inhibition of FtsZ protein, which forms the Z-ring, a polymer necessary for efficient bacterial cell cytokinesis.
Totarol may also function by disrupting the structural integrity of the phospholipid bilayer of bacteria by weakening Van der Waals interactions with its phenolic group, which also results in bacterial cells unable to synthesize ATP. Motivation for totarol functioning via disruption of membrane structure is due to its high phospholipid/water partition coefficient. However, totarol's partitioning capability was only observed at concentrations 10 to 100 fold higher than required for antibacterial activity. Thus it is unlikely that totarol is an uncoupler of bacterial respiration at the low levels observed in antimicrobial studies.

Traditional use

The use of Podocarpus totara extract in Māori medicines for treatment of fevers, asthma, coughs, cholera, distemper, chest complaints and venereal disease dates back to over 100 years. The timber of Podocarpus totara is renowned for its resilience against rotting, which made it valuable to Māori for housing, waka, fencing, pā stockades, drinking vessels, shovels and carvings in chiefs houses. Tōtara bark was used to cover kelp bags containing preserved muttonbirds known as pōhā. The tōtara tree was accorded divine status by Māori based on its immense size and its use in making waka employed on long, dangerous voyages. Carved tōtara can be viewed as a representation of ancestry and historical and mythological events. The tōtara can also be seen to represent a connection between the past and the present, the secular and the spiritual. It has a strong link to Māori creation stories as it reaches down to Papatūānuku  and up towards Ranginui. The roots in the symbolism of the tōtara mark out a genealogical reference point which Māori believe ties them to the natural world, the whenua and their ancestral rohe. In Māori tradition tōtara is the first born child of Tāne-Mahuta and the forest goddess Mumuwhango and is considered a noble tree. According to tradition, felling a tōtara wasn't possible without seeking permission from Tāne-Mahuta. This involved performing specific rituals and chanting karakia. European settlers of New Zealand used the wood for wharf piles, bridges, railway sleepers, telegraph poles, lighthouses, mining equipment, fence posts and foundation blocks. The durability stems from the anti-bacterial activity of totarol. Houses, churches, grave markers, and even cobbles and kerbs were made of tōtara; strips of the bark were used as a roofing material. The presence of totarol means tōtara wood resists decay and insect attack in the heart timber.
Totarol has been found in organic matter in the embankments of a Neolithic site in Northern Sweden. Scientists have suggested that totarol from Cupressaceae resin at the site was used for its antibacterial and antifungal properties to preserve meat, as well as for its ability to repel insects. Totarol has been observed in the endemic Mexican plant Buddleja perfoliata, used in traditional medicine as a topical antiseptic and diuretic against headaches, colds, tuberculosis, heart disease, dizziness and nervousness. Totarol has also been identified in the leaves of Algerian grown ethnomedicinal shrub Myrtus Communis. Leaf extract from the flowering plant has been used historically as a medicine known as "El Rayhan". It was used as a decoction, infusion and health remedy for bathing newborns with inflamed skin and washing sores. Additionally, it was used to treat oral wounds, disorders of the digestive and urinary systems, diarrhea, peptic ulcers, hemorrhoids and inflammations. Totarol was detected in a 2024 Myrtus Communis leaf extract study and described as having considerable anti-inflammatory potential. Totarol from juniper or cypress trees was used in ancient Egyptian mummification. In 2023 totarol was found in an embalming workshop dated to around 664-525BC a few meters south of the pyramid of King Unas at Saqqara. Despite totarol's demonstrated antimicrobial efficacy, its commercial application remains primarily confined to the cosmetic industry. For totarol to be considered for clinical use, a comprehensive understanding of its mode of action is essential.

Biochemical properties

Totarol decreases the plasma levels of estrogens and can also effectively reduce pathogenic hepatic cells in vitro. Totarol's anti-cancer activity is hypothesized to be due to the natural product's ability to form an o-quinone methide in vivo. Totarol also prevents cells from undergoing oxidative stress in vitro by acting as a hydrogen donor to peroxy radicals or reacting with other peroxy radicals to terminate undesirable radical reactions.

Biosynthesis

Totarol is a precursor to the formation of nagilactones that possess antifungal properties not possessed by totarol. Consequently, gymnosperms that produce totarol and nagilactones are able to defend themselves against bacteria and fungi.
Image:Totarol scheme 1.svg|thumb|Scheme 1. The initially proposed biosynthetic route of totarol.
The biosynthesis of totarol was difficult to determine. The main reason for the challenge in determining how the secondary metabolite is produced is because totarol does not follow the isoprene rule: the isopropyl group of totarol is in the "wrong" place at C14. Initially, it was hypothesized that totarol and the "normal" diterpene ferruginol, also found in Podocarpaceae, were derived by a precursor 2 that would be dehydrated and have its isopropyl group migrate to produce totarol 1 and ferruginol 3. This hypothesis was motivated by the well known santonin-desmotroposantonin rearrangement of steroid dienones into aromatic compounds. It is now accepted that totarol is synthesized biologically from ferruginol. Geranyl geranyl pyrophosphate 4 undergoes typical diterpene cyclization to form -abietadiene 5, which is oxidized to form ferruginol 3, which proceeds through a spiro intermediate to form totarol.
Image:Totarol scheme 2.svg|thumb|Scheme 2. The currently accepted biosynthetic route of totarol