Flavonoids are the most abundant phenol group in nature, present in a wide range of conifers [71]. Flavonoids have a central three-ring structure, but the different subclasses vary due to the centrally located heterocyclic ring structure (C-ring), which connects the two benzene rings [72]. To date, more than 6000 flavonoids have been recorded in several studies from plants. Flavonoids are aglycones in their basic structure, but most of them are glycosides in plants [73]. The subclasses of flavonoids found in the leaves, barks and seeds of conifers are represented in Table 1.

Lignans are phenylpropanoids dimers made up of two coniferyl or sinapyl alcohol units bound together at the tails [74]. Isolariciresinol, taxiresinol, lariciresinol, pinoresinol, and their glycosides are examples of such compounds. There is a growing interest in lignans, especially because of their chemotherapeutic ability [75]. The most commonly present lignin compounds in conifer spp. are shown in Table 1.

Stilbenes are produced by a number of conifer species, including Pinus sylvestris and Picea abies. Briefly, stilbenes are phenolic compounds with a heterologous bridge connecting two aromatic rings [76]. Many other compounds, such as trans-pinosylvin, cis-stilbene, resveratrol and piceatannol, have been isolated from the barks of conifer species (Table 1). For stilbenes, excellent antimicrobial effects have been reported [71].

Tannins are polyphenolic compounds that can be in a wide range of plants. Tannins are colored pigments, astringent and are characterized by a bitter taste [77]. Tannin-rich conifer bark extracts have antimicrobial properties and high potential in preventing lipids from oxidation in the liposome model [78]. As a result, the tannins can be divided into four main classes based on their structural characteristics: gallotannins, complex tannins, ellagitannins and condensed tannins [79]. Condensed tannins (CTs), also well-known as proanthocyanidins, are prevalent in P. abies and P. sylvestris tree bark [77]. Condensed tannins are considered as polymers or oligomers of flavan3-ol units connected by C-C bonds that are hydrolysis resistant [80]. Procyanidins (PCs) and prodelphinidins (PDs) are the most popular PAs. Catechin and other epicatechin units make up PCs. Epigallocatechin units make up PDs [81].

Tannins exhibit antioxidant activity through various pathways, including free radical scavenging, transition metal chelation, and inhibition of pro-oxidative enzymes [82], besides having the capability to bind and form complexes with proteins and other compounds, and being responsible for their biological activity [83]. Tannins also act as antimicrobial agents, inhibiting extracellular microbial enzymes, depriving microbial growth substrates, and exerting a direct action on microbial metabolism, such as the denaturation of cell membrane proteins [84]. In the food industry, they could be used as functional coatings, adhesives, preservatives and as flavor compounds [71]. In a study, pine and spruce bark-derived PA-rich extracts revealed good potential for use in the food industry to develop preservative agents and to prevent lipid peroxidation in food items containing fatty-acids [78].

In chronic obstructive pulmonary diseases (COPD), ROS play an important role [97]. The bark, wood, needle, and cone extracts of the Pinaceae family are high in polyphenolic compounds (Pycnogenol), primarily procyanidins, stilbenes, tannins and phenolic acids, and have significant antioxidant activity against ROS [98]. The antioxidant activity of conifer extracts has been confirmed by DPPH, FRAP and reducing power assays [51].

Flavonoids’ ability to scavenge free radicals can protect the human body from oxidative damage, which accelerates the ageing process [99]. Pycnogenol^®, a polyphenol-rich compound extracted from the bark of P. pinaster, P. glauca, and P. mariana, has shown the ability to boost plasma antioxidant capacity and ameliorate pulmonary function and asthma traits [51,100]. In a study by Senthilmohan et al. (2003), 6–12 weeks of supplementation of Enzogenol^®, and proanthocyanidin-rich flavonoid extracted from Pinus radiata bark in combination with vitamin C reduced DNA and protein oxidative damage in 55–75-year-old people [101]. In vivo studies have reported that the intake of quercetin alone did not protect DNA, but the combination of flavonoids (quercetin and myricetin) and isoflavonoids provides protection against DNA damage [102]. It has been found that the stilbenoid component resveratrol and piceatannol obtained from conifer spp. have more potent biological activities, namely as antioxidants [103]. Terpenoids and phenolic compounds, such as pinene, lycopene, camphene, gallocatechin, lutein, limonene and catechin found in the extract of conifer bark and needles have also been reported for their antioxidant potential by DPPH, FRAP, H₂O₂, ABTS assays, as shown in Table 2 [104].

When contagious microorganisms such as fungi, bacteria and viruses come into contact with the body, they remain in specific tissues and flow into the bloodstream, causing inflammation [105]. This also occurs as an end result of tissue damage, cancer, cell death, degeneration and ischemia [106,107,108]. In most cases, both the innate and adaptive immune responses are responsible for inflammation development [109]. The primary protection against invading foreign microbodies and cancer cells is the innate immune system, which involves macrophages, dendritic cells, and mast cells [105]. In the adaptive immune system, specialized cells (B and T cells) remove foreign pathogens and cancer cells by generating specific receptors and antibodies [110]. Cytokines such as interleukins, interferons, tumor necrosis factor, eicosanoids (leukotrienes and prostaglandins), modulatory inflammation-transcription nuclear factor (NF-ĸB) and chemokines (monocyte chemoattractant protein 1), are the inflammatory mediators and cellular pathways that have been extensively studied in relation to human pathological conditions [111]. Tumor necrosis factor-α (TNF) is a pro-inflammatory cytokine that is secreted by a variety of cells and has a variety of cellular effects [112]. It has also been linked to a variety of human illnesses, including cancer, mental and skin disorders, immune and inflammatory diseases. IL-1 is another cytokine that primarily has a pro-inflammatory effect [113]. It raises the levels of pro-inflammatory cytokines, including IL-1, TNF and IL-6 [114]. On the other hand, IL-1 has been linked to anti-inflammatory properties. Likewise, IL-1α and IL-6 originating from activated mast cells in the innate immune response also boost acute phase protein synthesis and thus show some anti-inflammatory effects [115]. The cytokine family members, including IL-12, IL-27, IL-23 and IL-35, function as a pro- and anti-inflammatory response [111,116,117]. On the other hand, IL-10 has been recognized as an effective anti-inflammatory cytokine, and helps in preventing several pro-inflammatory mediators from further action [118]. It protects tissue from homeostasis, defends against injury and damage caused by an overactive inflammatory response [118,119,120]. TNF-α accelerates PGE2 synthesis changes caused by edema and the flow of blood [46]. The extraction of plant materials is the first step in deciding the plant biological activities. The is a high probability of synergism between bioactive components when a whole extract is used, which could be lost if each and every component is isolated [121]. This form of synergism has been documented in numerous medical studies, generally for anti-inflammatory function [105]. There are different types of extraction and separation processes, such as:

1. Soxhlet extraction: The Soxhlet extraction method is a more efficient extraction method with high extraction yield and requires less solvent and time. This method requires electricity and solvents such as methanol, petroleum ether, and acetonitrile for the extraction process. However, sometimes high temperature and long extraction time enhance the possibility of thermal degradation and the loss of bioactive compound fraction activity [122].

2. Percolation: Extraction yield is better in percolation than maceration; in this process, pre-soaked plant material is added to a container, which allows the constantly controlled removal of the extract via a valve at the bottom and adding fresh solvent from the top.

3. Maceration: Maceration is carried out at room temperature by soaking the material with the solvent with eventual stirring. It has the advantage of moderate extraction conditions but suffers from high solvent consumption, long extraction times and low extraction yields. It could be used for the extraction of thermo labile components.

4. Ultrasound-assisted extraction: In UAE, the plant material, usually in a glass container, is covered by the extraction solvent and put into an ultrasonic bath. It decreases extraction time and improves extraction yields due to mechanical stress, which induces cavitation and cellular breakdown, and has gained increasing popularity [123]. For the isolation of extract from the solvent, the distillation process and many evaporators are used. After isolation to concentrate the extract, many researchers used a rotary evaporator, a normal air-drying process and distillation methods. Generally, to separate different solvent extracts, a separatory funnel is required [124].

Anti-inflammation is one of the main recorded effects of conifer phytochemicals among the numerous biological activities that have been studied so far. Table 3 reported the anti-inflammatory effect of conifer phytochemicals in in vivo and in vitro models.

Cupressuflavone (CUF) isolated from C. macrocarpa has the ability to reduce the levels of several cytokines, including IL-1b, IL-6, TNF-α and PGE2, in plasma dose-dependently, and thus acts as an anti-inflammatory agent [46]. Triterpenoids and abietane type’s compounds extracted from Abies chensiensis show anti-inflammatory effects against NO production in RAW 264.7 macrophage cells [125]. It has been found that tasumatrol, deacetylbaccatin, paclitaxel and many other terpenoids extracted from Taxus spp. are effective in the anti-inflammatory process initiated through the carrageenan and cotton pellets induced edema model [21,22]. Kyung-Jae Cha. (2016) reported that in atopic dermatitis, the Picea wilsonii mast extract is useful and potent only in the inhibition of the production of the inflammatory cytokines IL-6, MCP-1 and IL-13, without significant change in IL-8 production induced in human adult low-calcium high-temperature (HaCaT) cell lines [115]. T. occidentalis mother tincture-containing terpenoids (thujone), polyphenols and flavanoids have potential in reducing ulcerative colitis inflammation in the mouse intestine and rectum by decreasing the stimulation of the pro-inflammatory cytokines IL6 and TNF-α induced by 2,4,6-trinitrobenzenesulfonic acid (TNBS) [126]. THP-1 cell adhesion to TNF was suppressed by enzogenol at a concentration of 5–25 g/mL onto TNF-α-activated human umbilical vein endothelial cells (HUVEC) by reducing integrin β2 induction and inhibiting monocyte trans-endothelial migration [127]. The anti-inflammatory and platelet-inhibitory effects of pycnozenol, extracted from Pinus maritime bark extract, inhibited the activity of cyclooxygenase (COX)-1 and COX-2 present in human plasma [128]. Inflammation has been attributed to cancer and neurodegenerative diseases [46].

According to the report by the World Health Organization, cancer was a major cause of death in 2018, with a death rate of 9.6 million people [129]. Hippocrates, before 370 B.C., coined the word “cancer” to describe carcinoma tumors [130]. On the basis of evidence, bone cancer was identified in Ancient Egyptian mummies in around 1600 B.C., and cancer of the breast was identified in 1500 B.C., although there is no record of a cure for cancer [131]. Giovanni’s research laid the foundation for scientific cancer techniques in 1761, when he performed the first autopsies on dead human bodies to determine the connection between a patient’s disease and their death, as well as pathologic studies [132]. Cancer has been identified as the chief matter of public health concern around the world [133]. Surgery, radiotherapy, and chemotherapy are some of the conventional cancer treatments [134,135]. On the other hand, despite the use of a variety of synthetic drugs for cancer treatment and the successful completion of different management schedules, current therapies are not able to achieve the desired results, as tumor relapse and metastasis are common [136]. Nature contains various chemicals and pharmacologically active substances which act as anticancer drugs [137]. Recently, many of the phytochemicals and synthetic analogs, such as HS-1793 (resveratrol), have been identified as inhibiting the growth of cells and inducing apoptotic cell death, helping to cure cancer [138]. While only a few phytochemical compounds obtained from natural products have been developed into clinically active drugs, their bioactive components may be used as a basis for the development of more successful analogues and prodrugs using chemical techniques such as metabolomics, complete or combinatorial fabrication, and biosynthetic pathway modification [139]. Many phytochemical compounds are highly efficient in inducing apoptosis and cytotoxicity by modulating different MAPK andAKT/PI3K pathways, and suppress cancer cells line invasion and migration potential along with the stimulation of senescence phenotype, regulation of Bax or p53 genes, cell cycle arrest and modulation of IL-8, IFN-γ, TNF, IL-6 [140]. Many of the compounds derived from conifers’ bark and leaves act as antitumor drugs, such as paclitaxel (PTX) (trade name Pycnogenol^® and Taxol^®), a diterpene found in the crude extract of P. pinaster and Taxus brevifolia bark [141]. Paclitaxel, as well as its analogues docetaxel (taxoteres) and jevtanas (cabazitaxel), are examples of chemotherapeutic synthetic analogues derivative from plants that have been formulated and validated clinically [139]. By binding microtubules, PTX and other microtubule-targeting agents (MTAs) induce cellular death [142]. Microtubules are tubulin heterodimers that play a role in disease and perform numerous cellular functions including transport, force production in cell division, and structural support [143]. During the G2 phase of the cell cycle, tubulin is produced, and microtubules are assembled. Microtubule stabilizing agents, such as PTX, bind to α/β tubulin in order to disassemble microtubules. As a result, they cause cell death and are used as an anti-cancer agent [144]. In general, cells exposed to PTX are stuck in the G2/M phase, resulting in death due to failure to move through the cell cycle [145,146]. Recently, a study published in 2017 found that pycnogenol and PTX at doses of 20 g/mL and 0.5 μM cause DNA and mitochondrial damage in cancer breast cell line (MDA-MB) in 24 h, and concluded that it is possibly a target drug for cancer treatment through DNA and mitochondrial damage mechanisms [141]. From different conifer species crude extracts, the anticancer activities on different cancer cell lines are summarized in Table 4.

Neurodegenerative diseases (NDs) are more common among the elderly and may even lead to death, and so are a major threat in the 21st century [175]. AD, Parkinson’s disease (PD), Huntington’s disease (HD), amyotrophic lateral sclerosis (ALS), frontotemporal dementia, and the spinocerebellar ataxias are examples of ND [176], whose main features include nitrosative/oxidative stress, mitochondrial dysfunction, aggregated proteins accumulation, synapse loss, neuro-inflammation and decreased neuronal survival [177]. The progression of ND is also affected by genetic and surrounding ecological factors [175]. Indeed, it has been stated that the appropriate mechanism behind the cause of ND is mitochondrial dynamics variation, which elevates the oxidative damage, altering the biological activity of respiratory complexes, which results in brain energy dysfunction [178]. These stimuli trigger cellular stress, which leads to the synthesis and release of brain-derived neurotrophic factor (BDNF), as well as the activation of transcription factor CREB (cAMP response element-binding protein), with consequent expression of Arc (synaptic plasticity), PGC-1 (cellular energy metabolism), and APE1 (DNA repair enzyme), as well as the activation of the tropomyosin-related kinase (Trk B) receptor family and other downstream protein kinases [179].

Neurotrophins avoid neuron degeneration by binding to and activating the Trk receptor family, which is located in the plasma membrane [177]. Neurotrophins work by binding to and activating the Trk receptor family, which is found in the plasma membrane, to prevent neuron degeneration. Since neurotrosphins bind to Trk receptors, they create a microenvironment that promotes neuron development [180]. Various intracellular signaling pathways, such as ERK and PI3k/AKT, are regulated as a result of this binding, allowing cells to survive and aiding in the recovery of neurons from neurodegeneration. Additionally to signaling pathway activation, neurotrophins support Bcl-2 gene expression, which inhibits intracellular apoptosis [180]. Thus, early diagnosis of neurodegeneration may allow for early treatment, which may help to prevent the disease from progressing further [181]. Inhibition of the N-methyl-D-aspartate (NMDA) receptor can prevent or postpone AD. The drugs memantine and namzaric, which act as antagonists for the NMDA receptor, are used to treat AD patients [182].

Bioactive molecules have been recognized for their valuable biological effects, including neuroprotective properties, such as the ability to regulate mitochondria in a way that is distinct from TMs [183]. Branco et al. (2018) found that the flavonoid-rich A. angustifolia bracts extract (AAE) has neuroprotective properties by restoring rotenone-induced mitochondrial complex I, inhibiting the formation of lipid peroxidation and neuronal ROS, and through over expression of NDUFS7 protein and NDUFV2 gene levels in human dopaminergic SH-SY5Y cells [43]. Bark extract of P. pinaster shows protective effects against oxidative hemolysis induced by H₂O₂, the formation of thiobarbituric acid reactive products and lipid peroxidation [184]. In addition, it prevents oxidative damage to many proteins aggregation and may lessen the risk of several NDs, such as AD, PD and HD [185]. The neuro-protective potential of various conifer spp. crude extracts is summarized in Table 5.

AD is a common neurodegenerative disease that affects 80% of the elderly population, accounting for about half of all dementia cases and ultimately results in death [186]. Its symptoms include failure to learn, gradual memory loss, and deterioration in behavior and neuronal function [187]. Regarding treatment, only five approved treatment options are licensed in the European Union for the treatment of AD, including rivastigmine, donepezil (cholinesterase inhibitors (ChEIs), galantamine, and memantine (NMDAR antagonist) [188]. An antimitotic agent paclitaxel widely used for the treatment of lung, ovarian and breast cancer has also been investigated as a possible treatment for AD [189]. It is mainly effective in the treatment of tauopathies, which are disorders caused by mutations in the tau protein, which is abundant in central nervous system (CNS) cells and acts by stabilizing microtubules [190]. The consumption of polyphenol-rich foods or beverages has been related to the prevention of AD in distinct studies [191]. The accumulation of amyloid-(A) in brain and leptomeningeal vessels causes cerebral amyloid angiopathy (CAA), which is also a central component of neuritic plaques in AD amyloid-(A) and has been related to the pathogenesis of two of the most common forms of dementia: AD and CAA. As a result, Aβ should be a top priority in the treatment of these diseases, which currently have no effective therapies [192]. Taxifolin, an antioxidant and anti-glycation flavonoid, reduces Aβ aggregation and its accumulation in the cerebrovascular system. In vitro studies have shown that taxifolin facilitates Aβ clearance in the brain, prevents Aβ fibril formation and CAA cognitive loss, and increases cerebral blood flow [193]. The methanol extract of P. roxburghii bark contains bioactive compounds, such as quercetin and gallic acid, which play important roles in neuroprotection by reversing mitochondrial dysfunction, free radical formation, and improving memory and cognition in rats, as well as reducing oxidative stress by improving acetylcholine levels. Furthermore, anti-AD activity has been documented in Pinus species, such as P. halepensis and P. massoniana [194]. Piceatannol, a compound derived from pine bark, has proven to be effective in preventing AD [195]. Resveratrol (RV), a stilbenoid, protects neurons from oxidative damage in a variety of ways, such as lowering lipid peroxidation and increasing intracellular antioxidant levels including antioxidant enzymes catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and heme oxygenase 1 (HO-1) [196]. In this way, RV acts as an anti-AD agent by reducing neuroinflammation, inhibiting Aβ-plaque formation and tauopathy, and as a result inhibits neuronal death and improves memory [197]. Pycnogenol derived from P. pinaster bark has antioxidant, anti-inflammatory, and neuroprotective properties, including inhibition of amyloid-induced neuron apoptosis [198]. When the effect of pycnogenol was investigated in AD-related pathology in a β-amyloidosis mouse model, a decline in plaque numbers was found, while no changes were reported in the soluble β-amyloidosis levels, astrocytes, neurons, microglia, myelination pattern, morphology of axons and the gene expression of APP-processing enzymes [199]. Hence, it is suggested that pycnogenol has potential use in the prevention or in early stages of AD and mild cognitive impairment (MCI) [200]. Table 5 summarizes the neuroprotective potential of different conifers’ phytochemicals in AD.

PD is second to AD in terms of the most prevalent progressive ND, with an estimated global prevalence of over 10,000,000 cases [201]. The selective loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) leads to PD. Briefly, PD occurs due to oxidative stress, dysfunction of mitochondrial complex-1, oxidative cell damage, neuronal excitotoxicity, calcium homeostasis, apoptosis, distressed energy metabolism, inflammation and protein aggregation, such as a-synuclein, apoptosis, and interaction between genetic and environmental causes [202]. Due to uncoordinated mouth and throat movements, PD causes bradykinesia, muscle rigidity, rest tremor, and the loss of postural control, as well as certain secondary symptoms, such as dementia, sialorrhea, soft voice, and trouble swallowing [203,204]. Oxidative stress generates ROS that causes oxidative damage, such as 4-hydroxynonenal (HNE), 26S proteasome and interferes with dopamine metabolism leading to PD [85]. Changes in protein ubiquitination and degradation have recently been related to dopaminergic cell death in PD [205]. Presynaptic protein α-synuclein (α-syn) influences the release of neurotransmitters from synaptic vesicles in the brain [206]. Currently, the treatment of PD includes drugs such as L-DOPA, which is catalyzed primarily by dopa decarboxylase in the brain, and some others such as ropinirole, selegiline, and rasagiline. Ropinirole has some adverse effects, including ankle oedema, vomiting, nausea, hypotension, insomnia, weight loss, hallucinations, psychosis, arrhythmia, dry mouth, nightmares, persistent diarrhea, somnolence and constipation, limiting their clinical applications [207]. As a result, the focus of rising interest in alternative treatments for ND, such as PD, has turned to natural products, which can provide alternatives due to their high effectiveness and few side effects [208]. Many plant extracts tend to stop α-syn from oligomerization and fibrillation, which is an emerging therapeutic mechanism in PD [183]. Methanol extract of J. communis at doses of 100 and 200 mg/kg was found to be effective in reducing catalepsy, enhancing locomotor activity (actophotometer), and increasing the level of reduced glutathione (GSH), protein level and muscle activity in rats [209]. In an in vitro study on Fisher F344 rats, Zhang et al. (2010) discovered that RV protect dopaminergic neurons from damage caused by MPP+, 6-OHDA, and also show efficacy against lipopolysaccharide-induced neurotoxicity by inhibiting nuclear factor kappa B (NF-κB) signaling and microglial activation [210].

Insomnia is a chronically debilitating disease that has become increasingly common, posing immense health and economic challenges for both individuals and the community [211]. Trouble falling asleep, staying asleep, fragmented sleep (repeatedly waking up at night or waking up early in the morning) are all symptoms of this condition [212]. While behavioral therapy, psychotherapy and light therapy have all been used to treat insomnia, the most common medications for insomnia are hypnotic drugs that target GABAA-benzodiazepine (BZD) receptors, such as diazepam and zolpidem [213]. However, several side effects have been identified, including cognitive impairment, resistance, headaches, nausea, and rebound insomnia [214,215]. Methanol extract of A. webbiana leaves showed potent synergistic effect in mice at dose of 100, 150, and 200 mg/kg, with sleep-inducing sedative drugs, diazepam (6 mg/kg), pentobarbitone sodium (50 mg/kg) and propylene glycol [157]. In addition, the major monoterpenoid components present in Pinus spp., α-pinene and 3-carene, have been reported to have hypnotic effects through GABAA-BZD receptors. 3-carene increases the length of sleep in mice given pentobarbital-induced sleep drugs by binding to the BZD site of the GABAA-BZD receptor α1 and ϒ2 [216].