Paulownia Organs as Interesting New Sources of Bioactive Compounds

Paulownia spp. is a genus of trees in the Paulowniaceae family. It is native to southeastern Asia (especially China), where it has been cultivated for decorative, cultural, and medicinal purposes for over 2000 years. Depending on taxonomic classification, there are 6 to 17 species of Paulownia; P. tomentosa, P. elongata, P. fortunei, and P. catalpifolia are considered the most popular. Nowadays, Paulownia trees are planted in Asia, Europe, North America, and Australia for commercial, medical, and decorative purposes. Lately, growing interest in Paulownia has led to the development of various hybrids, the best-known being Clone in vitro 112, Shan Tong, Sundsu 11, and Cotevisa 2. Paulownia Clone in vitro 112 is an artificially created hybrid of two species of Paulownia: P. elongata and P. fortunei. The present review of selected papers from electronic databases including PubMed, ScienceDirect, and SCOPUS before 15 November 2022 describes the phytochemical characteristics, biological properties, and economic significance of various organs from different Paulownia species and hybrids, including P. tomentosa, P. elongata, P. fortunei, and Paulownia Clone in vitro 112. Many compounds from Paulownia demonstrate various biological activities and are promising candidates for natural preparations; for example, the leaves of Clone in vitro 112 have anti-radical and anticoagulant potential. However, further in vivo studies are needed to clarify the exact mechanism of action of the active substances and their long-term effects.


Introduction
Paulownia is a genus of trees in the Paulowniaceae family [1]. It is native to southeastern Asia (especially China), where it has been cultivated for decorative, cultural, and medicinal purposes for over 2000 years [1,2]. It is also known as the princess tree, royal tree, Kiri tree, empress tree, and phoenix tree [3,4], while its Chinese name is 泡桐 (pāo tóng) [5]. The genus Paulownia is believed to comprise 6 to 17 species depending on taxonomic classification [6]. Of these, P. tomentosa, P. elongata, P. fortunei, and P. catalpifolia are considered the most popular [4].
Nowadays, Paulownia trees are planted in Asia, Europe, North America, and Australia for commercial, medical, and decorative purposes [7]. Due to their fast growth rate and adaptability, they are considered invasive species in some countries. However, most of the risks could be attenuated by planting hybrids that produce infertile seeds (e.g., Clone in vitro 112) [1,6]. Paulownia can adapt to varied environmental conditions, and it has a fast growth rate and exceptional regenerative abilities; a cut tree trunk can regrow up to 2-4 m in one year [7]. In fact, it is one of the fastest-growing trees in the world, being able to produce several times more biomass in one year than some of the slower-growing species. These properties have led to an increased interest in establishing Paulownia plantations for the purpose of biomass production [1]. The use of Paulownia as a bioenergy crop, i.e., for the production of biofuel and CO 2 sequestration is also being considered [8]. In addition, its ability to withstand high concentrations of heavy metals (e.g., Mn, Pb, or Zn) Paulownia Clone in vitro 112, also known as Oxytree for its large leaves and ability to absorb large amounts of CO2, is an artificially created hybrid of two species of Paulownia: Paulownia elongata and Paulownia fortunei [24]. The first plantations were established in 2014 [7]. The plant is believed to effectively improve air quality. It has a faster growth rate than other species of Paulownia and can tolerate a wide range of temperatures, which makes it a convenient and profitable biomass and bioenergy crop [25]. Since it has infertile seeds, planting Clone in vitro 112 does not carry any risks of it spreading uncontrollably and becoming invasive [6].
The present review describes the phytochemical characteristics, biological properties, and economic value of various Paulownia organs, including Paulownia Clone in vitro 112. This review is based on studies identified in electronic databases, including PubMed, Sci-enceDirect, and SCOPUS. The last search was run on 15 November 2022

Taxonomy of Paulownia
Although Paulownia currently belongs to the Paulowniaceae family, it was previously classified as a member of Scrophulariaceae [16]. The total number of Paulownia species is not universally agreed upon; depending on taxonomical classification, this number can Paulownia Clone in vitro 112, also known as Oxytree for its large leaves and ability to absorb large amounts of CO 2 , is an artificially created hybrid of two species of Paulownia: Paulownia elongata and Paulownia fortunei [24]. The first plantations were established in 2014 [7]. The plant is believed to effectively improve air quality. It has a faster growth rate than other species of Paulownia and can tolerate a wide range of temperatures, which makes it a convenient and profitable biomass and bioenergy crop [25]. Since it has infertile seeds, planting Clone in vitro 112 does not carry any risks of it spreading uncontrollably and becoming invasive [6].
The present review describes the phytochemical characteristics, biological properties, and economic value of various Paulownia organs, including Paulownia Clone in vitro 112. This review is based on studies identified in electronic databases, including PubMed, Sci-enceDirect, and SCOPUS. The last search was run on 15 November 2022

Taxonomy of Paulownia
Although Paulownia currently belongs to the Paulowniaceae family, it was previously classified as a member of Scrophulariaceae [16]. The total number of Paulownia species is not universally agreed upon; depending on taxonomical classification, this number can range from 6 to 17 [6]. Li et al. [26] define eight species: P. tomentosa, P. coreana, P. kawakamii, P. fortunei, P. elongata, P. catalpifolia, P. australis, and P. fargessi. The Chinese Flora Editorial

Commercial Uses and Economic Value of Paulownia
Fast growth rate and good adaptability are the reasons for increased interest in Paulownia as a biomass source. For example, it is a good candidate for short-rotation forestry (SRF). SRF is a type of tree cultivation where crops reach their optimal size and are ready to be harvested in 8 to 20 years [35]. Unlike Paulownia, most of the tree species commonly used in SRF plantations (e.g., poplar, willow, black locust, or alder) employ the less efficient C3 photosynthesis [36]. The rotation cycle of SPF is usually three to six years; trees that are planted more densely have shorter rotation times. It is often possible to plant up to 10,000 units per 1 ha [37]. In the past, the biomass collected from SRF plantations was used primarily to produce cellulose pulp; however, nowadays it is mainly utilized as a source of thermal or electrical energy, i.e., as a bioenergy crop [38]. In this aspect, biomass acquired from SRF could help replace fossil fuels as a source of energy and reduce the emissions of greenhouse gasses [39]. However, this form of tree cultivation has its disadvantages. SRF crops release a large quantity of volatile organic compounds (VOC) that contribute to tropospheric ozone production [37]. Tropospheric ozone is toxic-it can increase the production of reactive oxygen species in cells and impair CO 2 absorption in plants [40,41].
Paulownia leaves have high nutritional value and are a good source of bioactive substances, which makes them a valuable animal feed component (Table 1) [10,42,43]. They are rich in minerals, proteins, nitrogen, and crude fiber. For example, P. tomentosa leaves have higher levels of manganese, zinc, tyrosine, and methionine than lucerne [42,44]. The addition of Paulownia leaves to rabbit feed (up to 15%) increased the blood concentration of high-density lipoprotein (HDL) and decreased the amount of low-density lipoprotein (LDL). Furthermore, it reduced the number of pathogenic bacteria in the caecum. However, at high concentrations, the rabbits demonstrated slower growth [44]. The leaves are also a good fertilizer and can enrich the soil with valuable organic matter and microorganisms. Fallen leaves support the growth of bacteria, which stimulate the production of various phytohormones, enzymes, biosurfactants, and precursors of secondary metabolites, which enhance plant growth and improve immunity to pathogens. These bacteria contribute to the circulation of minerals in the soil, bind atmospheric nitrogen, and decompose organic matter; however, they can also promote the occurrence of various diseases [7,45].
Tricin-7-O-β-Dglucopyranoside Antihypoxic [46,55]   Paulownin [10,96] Lately, interest in Paulownia has increased following the discovery of multiple geranylated flavonoids in P. tomentosa, many of which have never been isolated from any other plant. C-geranylated flavonoids, a group of flavonoid derivatives, are relatively rare and occur only in a small number of plant families [5,14]. They consist of a flavonoid skeleton and a monoterpenoid side chain [97]. The geranyl part is synthesized by the mevalonate pathway, while the flavonoid part is the product of the shikimic acid pathway. The two components are linked together by prenyltransferases [5]. C-geranylated flavonoids exhibit a wide range of activities, including antioxidant, anti-inflammatory, antibacterial, antiviral, antiparasitic, and cytotoxic activities [5,22,97,98]. Recent studies have shown that they could have the potential to be developed into anti-inflammatory drugs [17].
Several C-geranylated flavonoids isolated from P. tomentosa fruit have demonstrated antiproliferative and cytotoxic effects against the THP-1 cell line. Diplacone demonstrated the strongest activity in both regards, while 3 ′ -O-methyl-5 ′ -hydroxydiplacone exhibited a relatively strong antiproliferative, but weaker cytotoxic effect [14]. These results are in line with those of Zima et al., where diplacone and 3 ′ -O-methyl-5 ′ -hydroxydiplacone were found to be the most active C-geranylated flavonoids isolated from the fruits of P. tomentosa [57]. In addition, another geranylated flavonoid (CJK-7) upregulated autophagy and induced caspase-dependent cell death in the HCT-116 human colon carcinoma cell line [99].
A fruit extract inhibited the activity of protein tyrosine phosphatase 1B (PTP1B) and α-glucosidase, which are important targets in the treatment of obesity and diabetes. Geranylated flavonoids isolated from the extract also showed potent inhibitory activity. The most effective compound turned out to be mimulone [63].
Kim et al. reported that methanol extract from P. tomentosa flowers had neuroprotective properties. The extract reduced glutamate-induced toxicity in primary cultured rat cortical cells in a dose-dependent manner. Protection from glutamate-induced damage plays a crucial role in preventing neurodegenerative diseases. Glutamate is an endogenous amino acid that acts as an excitatory neurotransmitter. Although it plays a significant role in the nervous system by facilitating neuroplasticity, neuronal survival, and learning processes, it can also promote the development of neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, or epilepsy. Among five flavanones isolated from the extract (5,4 ′ -dihydroxy-7,3 ′ -dimenthoxy-flavanone, 5-hydroxy-7,3 ′ ,4 ′ -trimenth oxyflavanone, diplacone, mimulone, and isoatriplicolide tiglate), isoatriplicolide tiglate had the most potent neuroprotective ability. Its incubation with rat cortical cells at concentrations of 1 µM and 10 µM improved cell viability to 43% and 78%, respectively [19].
A furanquinone (methyl 5-hydroxy-dinaphtho [1,2-2 ′ 3 ′ ]furan-7,12-dione-6carboxylate) isolated from the stem of P. tomentosa had an inhibitory effect on cathepsin K [90]. Cathepsin K is a protease expressed mainly in bone marrow, although small amounts can be also found in other tissues. It is involved in the process of bone matrix degradation by osteoclasts. Cathepsin K inhibitors are currently under evaluation as potential drugs for osteoporosis treatment [102].

P. fortunei and P. elongate-Predecessors of Paulownia Clone In Vitro 112
P. fortunei is also known as the Chinese parasol tree [97]. Its flowers are edible and can be used to make a dish called Zheng Cai. Extracts from different organs of the tree have also been used to treat bacterial infections such as dysentery, tonsillitis, and bronchitis, as well as enteritis and hypertension [20]. P. elongata demonstrates an above-average growth rate. In its second year, it can reach a height of 4 m and have a diameter of 5-6 cm [103].
Liu et al. report that P. fortunei flower extract decreased total cholesterol concentration in plasma, prevented hepatic lipid accumulation, and facilitated weight loss in mice fed with high-fat diets. HDL levels were increased, while plasma insulin and glucose concentrations were reduced. These effects can be attributed to the upregulation of 5 ′ AMPactivated protein kinase (AMPK) pathway and the activation of insulin receptor substrates (IRS1). AMPK plays an important role in regulating lipid metabolism, while IRS1 activates the insulin signaling cascade. The phosphorylation levels of both AMPK and IRS1 were significantly increased in mice supplemented with the extract [20].
Extracts from fresh and fermented leaves of P. fortunei also had antibacterial properties. They inhibited the growth of bacteria (Salmonella enterica, Streptococcus pyogenes, Staphylococcus aureus, Pseudomonas aeruginosa, Paenibacillus alvei) and fungi (Candida albicans), although this inhibitory effect was more pronounced against Gram-negative bacteria [11].
Sheep fed with P. elongata leaves had lower leukocyte and erythrocyte counts and demonstrated lower plasma glucose concentrations [109].
Extracts from fresh and dry leaves of P. fortunei and P. elongata also appear to have antioxidant activity, as indicated by studies based on the TREAC assay (TROLOX Equivalent Antioxidant Capacity), in which the reactivity of an antioxidant is compared to the activity of TROLOX-a water-soluble vitamin E analog. The results are expressed as percentage inhibition of the ABTS •+ radical cation in comparison to TROLOX. Extracts from P. fortunei leaves showed a mean inhibition of 61.03% (fresh leaves) and 95.09% (dry leaves), while for P. elongata, these values were 50.21% and 60.88%, respectively [28,110,111]. The total flavonoid contents of the fresh leaf extracts were 157.53 µg/mL for P. fortunei and 102.58 µg/mL for P. elongata [28].

Paulownia Clone In Vitro 112-Characterization
Paulownia Clone in vitro 112 (also known as Oxytree and Biotree) is a hybrid of P. elongata and P. fortunei. It can withstand a wide range of temperatures (−25 • C to +45 • C), which allows it to be cultivated in many parts of the globe [7]. It is one of the fastest-growing deciduous trees in the world-it can reach up to 16 m in height and 35 cm in trunk diameter in only six years. After cutting, the trunk can regrow four to five times. Thanks these regenerative abilities, the wood can be harvested more than once. In addition, its root system can reach a depth of 9 m [25]. The tree can also rejuvenate contaminated soil and improve groundwater retention [112]. It absorbs large amounts of CO 2 , i.e., up to 111 tons/ha/year; in comparison, oak can assimilate only 9.1 tons/ha/year [25]. Another advantage of Oxytree is that it only produces infertile seeds, thus reducing the risk of it becoming an invasive species [6].
As wood harvested from Paulownia Clone in vitro 112 is lightweight and durable [6], it is sometimes referred to as 'aluminum wood'; 1 m 3 weighs approximately 310 kg and is rated as class I on the Janka scale, indicating a very soft wood [36]. The wood of Oxytree is lighter than the wood of other species of deciduous trees. Stochmal et al. [25] estimated that it is approximately 50% lighter, while Bikfalvi [112] reported that it is lighter by 30%. Moreover, Biotree wood is a good thermal insulator, has fine texture, and is resistant to deformation [112].

Chemical Content of Paulownia Clone In Vitro 112 and Its Biological Activity
Currently, there are few publications describing the chemical constituents of Paulownia Clone in vitro 112. As the chemical content varies between different species and cultivars, more research is needed to ascertain the content of secondary metabolites and their properties [4,113]. Adach et al. showed that the majority of active constituents of Oxytree leaves were phenolic compounds, the most predominant being verbascoside and its derivatives (methoxyverbascoside, hydroxyverbascoside). Other phenolics included apigenin-HexA-HexA, luteolin-HexA-Hex, and caffeic acid-Hex-dHexA. The total phenolic content was 205.5 mg·g −1 ± 6.41 [24,114]. Dżugan et al. reported that the total phenolic content of leaf tissue is 248.51 mg GAE/g (mg of gallic acid equivalents per gram of dry mass), and total flavonoid content is 147.71 mg QE/g (mg of quercetin equivalent per gram of dry mass) [4].
In addition, the leaf extract contained the iridoids catalpol and aucubin or 7-hydroxy tomentoside, with the total iridoid content of 15.16 mg·g −1 ± 0.274. Out of these, aucubin and its isomer 7-hydroxytomentoside were present in the highest concentrations. The extract also contained a small number of triterpenoids, including C 30 [66]. The substances presented in Table 4 were isolated by Adach et al. in 2020 and 2021.
Catalpol can penetrate the blood brain barrier and has anti-inflammatory, antioxidant, anti-apoptosis, antitumor, and neuroprotective properties [75,76]. It is used to treat agerelated macular degeneration, which is a disease manifested by visual distortions, dark spots, and impaired central vision [118].
Oxytree leaf extract had antioxidant activity. Adach et al. tested the effect of the extract and four fractions (A, B, C, and D) on human plasma treated with H 2 O 2 /Fe. Fractions A-C contained mostly verbascoside and its derivatives, as well as apigenin diglucuronide and luteolin diglucuronide. Fraction A and B contained iridoids. Fraction D contained phenolics, mainly acetylverbascoside and dimethylverbascoside, as well as apigenin and luteolin. Both the extract and all of the fractions significantly inhibited lipid peroxidation and oxidation of plasma protein thiol groups at the two highest concentrations (10 and 50 µg/mL). Moreover, fractions C and D were able to inhibit carbonylation of plasma proteins at all tested concentrations (1, 5, 10, and 50 µg/mL) [114].
Dżugan et al. have reported that the extracts from Oxytree leaves had lower antioxidant and antibacterial activity than extracts from other tested clones, with P. tomentosa x P. fortunei clones demonstrating the strongest effects. Additionally, the leaf blade extract showed four to nine times greater biological activity than the petiole extract. Higher activity correlated with a higher polyphenol concentration and a greater share of flavonoids in the polyphenol fraction (which ranged from 60 to 86% in the majority of cases). P. elongata x P. fortunei had the lowest antioxidant activity and polyphenol and flavonoid contents out of all tested clones [4].
The extract and four fractions (A-D) from the leaves of Clone in vitro 112 also showed anti-platelet activity, successfully inhibiting ADP-induced platelet aggregation at the highest concentration (50 µg/mL). They also lowered the adhesion of thrombin-activated platelets to fibrinogen and collagen. Lipid peroxidation was reduced in thrombin-activated platelets at all tested concentrations (1, 5, 10, and 50 µg/mL), although these results were not always statistically significant. The strongest effect was observed with fraction D; at the concentration of 50 µg/mL, the peroxidation was reduced by 60%. All the preparations increased the concentration of O 2 − . in resting and activated platelets. Overall, the extract had stronger antiplatelet activity than the fractions [24]. Moreover, fraction D showed the strongest anticoagulant activity in whole blood, which was determined with the Total Thrombus-Formation Analysis System (T-TAS) [113].

Conclusions
The reviewed literature demonstrated that the trees of Paulownia genus produce many promising chemical compounds (e.g., verbascoside, diplacone, mimulone, apigenin, catalpol, aucubin, and maslinic acid) with various biological activities (e.g., antioxidant, anti-inflammatory, antiproliferative, antibacterial, antiviral, neuroprotective, and hepatoprotective activities). Extracts and fractions from various Paulownia organs show beneficial properties as well. For example, the leaves of P. Clone in vitro 112 had anti-radical and anticoagulant effects, making them potential candidates for natural preparations (Figure 1). However, there is a need for more studies that would clarify the exact mechanisms of action and determine which active compounds are responsible for these effects. Moreover, animal studies and clinical trials should be performed to determine the in vivo efficiency of preparations and compounds from Paulownia and check their long-term effects and safety.