**1. Introduction**

Natural compounds with specific reference to plant-derived active ingredients are raising interest as coadjuvant treatment of human diseases. In recent years, this trend has consolidated and many natural products have been introduced into the market under the label of nutritional supplements (NS) or foods with special medical purposes (SMPF). Despite the effective and charming marketing efforts of companies in selling these products following medical routes, the real efficacy of nutraceuticals is still a challenging issue because of the inadequate technological framework for enhancing and testing their bioavailability.

Lipophilic active ingredients belonging to the families of terpens, saponins, and vegetal oils are poorly absorbed by intestine mainly because of their low water solubility [1,2]. The interaction of active compounds with intestinal epithelium depends on the dissolution of molecules in water-based intestinal fluids [3]. A viscous mucopolysaccharides rich layer strictly adhering to the enteric epithelium and synthesized by specialized Goblet cells, called unstirred water phase (UWP), (Figure 1) plays a pivotal role in regulating the absorption rate of active ingredients. Mucins, indeed, can interact with poorly-soluble molecules preventing their penetration [4,5].

**Figure 1.** Representation of UWP in the intestine. It is possible to recognize tight junctions and Goblet cells.

Only water-soluble molecules or intimately water-dispersed molecules can readily diffuse through this layer, favoring an effective intestinal absorption [6]. According to this premise, lipophilic terpens, such as boswellic acids (BA) extracted from the resin of *boswellia* genus trees, vegetal oils such as ginger oil (GO), *Serenoa repens* (Bartram) Small oil (SRO) and many others structurally similar compounds, are not effectively absorbed through the intestine even in the presence of bile salts [7,8]. Advanced technological matrices and delivery systems able of emulsifying lipophilic compounds or making them hydro-dispersible in the duodenal tract represent promising and effective strategies to improve their bioavailability.

Over the last decades, many companies and research institutes all over the world, tried to discover new performing technologies for enhancing natural compounds bioavailability, in some cases with great success as the patented technology PhytosomeTM. PhytosomeTM consists in a physical complexation of the active ingredient with a phospholipid enabling it to overcoming the enteric barrier, through an enterocyte membrane partitioning mechanism [9–11].

Many lipophilic plant-derived active principles have shown several biological relevant actions such as anti-inflammatory activity, enteric physiology restoring activity, and osteoarticular health promoting activity. However, their positive impact on human health is often limited by their poor solubility in aqueous systems, such as digestive fluids, so leading to low enteric bio-accessibility and bioavailability. Among them, SRO is a vegetal oil largely used in clinical practice to reduce symptoms of human prostatic hyperplasia (HPH). This oil that mainly works by inhibiting 5-α-reductase enzyme in prostatic cells and other androgen-sensible cells (sebaceous glands, hair follicles etc.) and by modulating expression of PSA and inflammatory cytokines [12,13] is mainly composed of free fatty acids (80–90% *w*/*w*) and several phytosterols of which β-Sitosterol is the most represented. SRO has shown to be effective in reducing lower urinary tract symptoms (LUTS) such as normalizing urine flow and reducing gland volume [14,15] at dosage of 320 mg of oil titrated at 80–90% in fatty acids per day. Despite this efficacy and the ascertained prostatic tropism [16], not many data are available about its overall bioavailability in humans [16,17] and in particular no data are available about the role of peculiar delivery systems in improving it in humans.

Several technological forms such as nanoemulsions (NE), solid lipid nanoparticles (SLN), and self-emulsifying drug delivery systems (SEDDS) have been developed over the years to deliver insoluble or poorly bio-accessible molecules. On the other side, it must be pointed out, that the most part of natural compounds in form of oils and lipids-based formulations (LF) are mainly composed of triglycerides (TG) in form of emulsion, NE and SLN and for this reason they are partially digested by gastric lipase (GL) that cleaves TG generating free fatty acids (FFA) and diacylglycerols (DAG) [18–20]. This underestimated physiological phenomenon can dramatically affect the strategy of delivery of the molecules entrapped into these technological systems [21,22], partially or totally compromising their overall bioavailability.

To design a platform for increasing enteric absorption of lipophilic molecules, some requirements should be considered pivotal to achieve a good pharmaceutical delivery tool: (i) delivery systems able of retaining active ingredients in the stomach and releasing them in the duodenum; (ii) systems able of enforcing bile salts effect in forming fine emulsions or micellar dispersions of lipophilic molecules in the first duodenum; (iii) systems stable to enzymes and peculiar environment of the gastro-enteric tract (iii) systems able of entrapping lipophilic compounds, even in oily form, in a hydro-dispersible solid matrix powder to be easily transformed in capsules or tablets. Taking into consideration the aforementioned requirements, a new technological matrix has been designed and developed to enhance lipophilic compounds enteric absorption. This technological platform, named Lipomatrix (Labomar Research, Istrana, TV, Italy), contains a molten fats core aimed to entrapping lipophilic compounds into a gastric-refractory environment and to emulsify them once exposed to duodenal fluids. This technology is based on a peculiar association of low melting point fats such as mono and diglycerides of fatty acids (MDGFA) commonly used in food industry (E471), and ascorbyl palmitate (ASP), an ester of L-ascorbic acid (AA) and palmitic acid (PA) widely employed in food, pharmaceutical and cosmetic industry as antioxidant agent (E304) [23,24]. The precise chemical features of ASP and its peculiar mechanism of action as emulsifying agent and drugs absorption enhancer are fully described in the Supplementary file section.

In this work, we present for the first time, the technology Lipomatrix and its efficacy in favoring apparent absorption and improving biological activities of SRO through human cell lines models. Beside this main goal, the apparent enteric permeability rate of an extract of Cranberry (FlowensTM) and Lycopene vehiculated in the mentioned technological matrix with SRO was assessed.

#### **2. Results and Discussion**

#### *2.1. Differential Scanning Calorimetry (DSC) Analysis*

DSC analysis was performed on MDGFA, mannitol, ASP, SRO, and Lipomatrix as described in Section 3.2.1.2, in order to evaluate possible changes in the crystallinity structure of the single raw material once embedded into the matrix and thereby speculate on its prevalent amorphous or crystalline state. In Supplementary file DSC thermograms for the single substances are reported.

From the analysis of the Lipomatrix thermogram achieved in the range temperature of 25–125 ◦C (Supplementary file) it is possible to recognize a melting pick between 47 and 55 ◦C coherent with the melting point of MDGFA even though there is a significant decrease in the melting temperature pick of about 10 ◦C in respect to MDGFA alone. This could mean that some interaction phenomena of MDGFA with the other matrix components occurs during Lipomatrix preparation, with an overall increase of the amorphous structure respect to the pure raw material [25,26]. Interestingly, comparing the overlapping thermograms of single components and Lipomatrix one in the range 30–300 ◦C (Figure 2, Table 1), it is possible to ascertain that both endothermic and exothermic picks of ASP completely disappear, while the only distinctive endothermic pick seems to coincide to Mannitol one. This fact could be explained considering the effective dissolution of ASP into the warmed lipidic matrix, according to the method of production, taking place to a prevalent amorphous or disordered crystalline phase molecular dispersion in the lipid matrix after cooling [27]. Finally, the appearing of

a well-shaped exothermic pick in the Lipomatrix sample around 220 ◦C, not recognized in none of the single components thermograms, could means that a sort of crystallization phenomenon occurs during the warming of the mix of lipophilic components and its further rapid cooling onto the Mannitol to get the final matrix. This phenomenon could be a further proof of an occurring structural change.

**Figure 2.** Overlapping thermograms of LIPOMATRIX containing SRO and single components. Black line represents ASP. Red line represents MDGFA. Green line represents Lipomatrix with SRO. Blue line represents Mannitol. Pink line represents SRO.

**Table 1.** Cristallinity index, entropy, and extrapolation pick time of Lipomatrix and any of its constituents.

