Natural deep eutectic solvents (NADESs) can serve as solvents for enzymes, are biodegradable, and have low toxicities. Eight NADESs with different hydrogen bond acceptors and donors were tested to improve the stability and activity of a lipoxygenase from Basidiomycete Pleurotus sapidus (LOXPSA). Betaine:sorbitol:water (1:1:3, BSorbW) and betaine:ethylene glycol (1:3, BEtGly) had the best impact on the peroxidation of linoleic acid and the side reaction of piperine to the vanilla-like scented compound piperonal. The yield of piperonal in NADESs increased by 43% in BSorbW and 40% in BEtGly compared to the control. The addition of BSorbW also enhanced the enzyme\'s stability at various temperatures and increased its activity during incubation at 60 °C. The demonstrated improvement in lipoxygenase activity and stability indicates versatile applications in industry, expanding the potential uses of the enzyme.

Substituted benzaldehydes are the most commonly used natural-occurring flavours in the world. The consumer\'s preference for \'natural or organic\' aromas has increased the request for flavours possessing the \'natural\' status. The resulting shortage of aromatic aldehydes of extractive origin, such as vanillin, veratraldehyde and piperonal, can be offset by developing a new biotechnological synthesis method. Here, we report a study on the microbiological reduction of five natural benzoic acid derivatives, namely p-anisic, vanillic, veratric, piperonylic and eudesmic acids, to produce the corresponding fragrant aldehydes. We found that different Basidiomycota strains can efficiently perform this transformation, with good chemical selectivity and tolerance to the toxicity of substrates and products. Besides confirming the carboxylic acid reductase activity of the already studied fungi Pycnoporus cinnabarinus, we discovered that other species such as Pleurotus eryngii, Pleurotus sapidus and Laetiporus sulphureus as well as the non-ligninolytic fungi Lepista nuda are valuable microorganisms for the synthesis of anisaldehyde, vanillin, veratraldehyde, piperonal and 3,4,5-trimethoxybenzaldehyde from the corresponding acids. According to our findings, we propose a reliable process for the preparation of the above-mentioned aldehydes, in natural form. KEY POINTS: • Fragrant benzaldehydes were obtained by biotransformation. • Basidiomycota strains reduced substituted benzoic acid to the corresponding aldehydes. • Anisaldehyde, vanillin, veratraldehyde, piperonal and 3,4,5-trimethoxybenzaldehyde were prepared in natural form.

Piperonal is a compound of key industrial importance due to its attractive olfactory and biological properties. It has been shown that among the fifty-six various fungal strains tested, the ability to cleave the toxic isosafrole into piperonal through alkene cleavage is mainly found in strains of the genus Trametes. Further studies involving strains isolated directly from different environments (decaying wood, fungal fruiting bodies, and healthy plant tissues) allowed the selection of two Trametes strains, T. hirsuta Th2_2 and T. hirsuta d28, as the most effective biocatalysts for the oxidation of isosafrole. The preparative scale of biotransformation with these strains provided 124 mg (conv. 82%, isolated yield 62%) and 101 mg (conv. 69%, isolated yield 50.5%) of piperonal, respectively. Due to the toxic impact of isosafrole on cells, preparative scale processes with Trametes strains have not yet been successfully performed and described in the literature.

Candida infections have been always a serious healcare related problem. The present study reports the preparation of hydrogels and formulations based on piperonyl-imino-chitosan derivatives and Amphotericin B drug for the treatment of Candida infections. The hydrogels were obtained by the imination reaction of chitosan with piperonal monoaldehyde, followed by the self-assembling of the resulted imines, while the formulations were obtained by an in situ hydrogelation method of chitosan with piperonal in the presence of Amphotericin B antifungal drug. The structural characterization of both hydrogels and formulations by Fourier transform infrared spectroscopy and Nuclear magnetic resonance spectroscopy revealed the formation of imine units between the reagents, while their supramolecular characterization using polarized optical microscopy and wide angle X-ray diffraction demonstrated that hydrophilic/hydrophobic segregation is the process which governed the formation of gel like systems. The systems were further investigated from the point of view of their further applications revealing that they were biodegradable, presented high swelling ability and were able to release the antifungal drug in a sustained manner, presenting promising antifungal activity against five Candida strains.

Piperonal is a simple aromatic aldehyde compound with a characteristic cherry-like aroma and has been widely used in the flavor and fragrance industries. Despite piperonal being an important aroma in black pepper (Piper nigrum), its biosynthesis remains unknown. In this study, the bioinformatic analysis of the P. nigrum transcriptome identified a novel hydratase-lyase, displaying 72% amino acid identity with vanillin synthase, a member of the cysteine proteinase family. In in vivo substrate-feeding and in vitro enzyme assays, the hydratase-lyase catalyzed a side-chain cleavage of 3,4-methylenedioxycinnamic acid (3,4-MDCA) to produce 3,4-methylenedioxybenzaldehyde (piperonal) and thus was named piperonal synthase (PnPNS). The optimal pH for PnPNS activity was 7.0, and showed a K m of 317.2 μM and a k cat of 2.7 s-1. The enzyme was most highly expressed in the leaves, followed by the fruit. This characterization allows for the implementation of PnPNS in various microbial platforms for the biological production of piperonal.
UNASSIGNED: The online version contains supplementary material available at 10.1186/s13765-022-00691-0.

The valuable aroma compound piperonal with its vanilla-like olfactory properties is of high interest for the fragrance and flavor industry. A lipoxygenase (LOXPsa 1) of the basidiomycete Pleurotus sapidus was identified to convert piperine, the abundant pungent principle of black pepper (Piper nigrum), to piperonal and a second volatile product, 3,4-methylenedioxycinnamaldehyde, with a vanilla-like odor through an alkene cleavage. The reaction principle was co-oxidation, as proven by its dependence on the presence of linoleic or α-linolenic acid, common substrates of lipoxygenases. Optimization of the reaction conditions (substrate concentrations, reaction temperature and time) led to a 24-fold and 15-fold increase of the piperonal and 3,4-methylenedioxycinnamaldehyde concentration using the recombinant enzyme. Monokaryotic strains showed different concentrations of and ratios between the two reaction products.

The fungal secretome comprises various oxidative enzymes participating in the degradation of lignocellulosic biomass as a central step in carbon recycling. Among the secreted enzymes, aryl-alcohol oxidases (AAOs) are of interest for biotechnological applications including production of bio-based precursors for plastics, bioactive compounds, and flavors and fragrances. Aryl-alcohol oxidase 2 (PeAAO2) from the fungus Pleurotus eryngii was heterologously expressed and secreted at one of the highest yields reported so far of 315 mg/l using the methylotrophic yeast Pichia pastoris (recently reclassified as Komagataella phaffii). The glycosylated PeAAO2 exhibited a high stability in a broad pH range between pH 3.0 and 9.0 and high thermal stability up to 55 °C. Substrate screening with 41 compounds revealed that PeAAO2 oxidized typical AAO substrates like p-anisyl alcohol, veratryl alcohol, and trans,trans-2,4-hexadienol with up to 8-fold higher activity than benzyl alcohol. Several compounds not yet reported as substrates for AAOs were oxidized by PeAAO2 as well. Among them, cumic alcohol and piperonyl alcohol were oxidized to cuminaldehyde and piperonal with high catalytic efficiencies of 84.1 and 600.2 mM-1 s-1, respectively. While the fragrance and flavor compound piperonal also serves as starting material for agrochemical and pharmaceutical building blocks, various positive health effects have been attributed to cuminaldehyde including anticancer, antidiabetic, and neuroprotective effects. PeAAO2 is thus a promising biocatalyst for biotechnological applications. KEY POINTS: • Aryl-alcohol oxidase PeAAO2 from P. eryngii was produced in P. pastoris at 315 mg/l. • Purified enzyme exhibited stability over a broad pH and temperature range. • Oxidation products cuminaldehyde and piperonal are of biotechnological interest. Graphical abstract.

Piperonal, an important aromatic aldehyde containing the benzene ring fused to 1, 3- dioxolane moiety, widely occurred in nature and known for its fragrance similar to vanillin and cherry. Alterations in its structural features led to the access of a wide range of piperonal derivatives viz. pyrazolines, chalcones, benzothiazepines, benzoxazoles, triazoles etc. having remarkable pharmacological properties like antimicrobial, anticancer, antidiabetic, antitubercular activities, etc. Therefore, the present study has been designed to highlight the progress made in the area of piperonal and its derivatives till date with respect to their synthesis and pharmacological activities. This may facilitate the synthesis of more novel derivatives with improved biological activities. Various search engines like google, Pubmed, Scopus etc. were used to gather the related information using different keywords.

Six quinoline-piperonal hybrids were synthesized and evaluated as potential drugs against Alzheimer\'s disease (AD). Theoretical analysis of the pharmacokinetic and toxicological properties of the compounds suggest that they present good oral bio-availability and are also capable of penetrating the blood-brain barrier, qualifying as leads for new drugs against AD. Evaluation of their inhibitory capacity against acetyl- and butyrilcholinesterases (AChE and BChE) through Ellmann\'s test showed that three compounds present promising results with one of them being capable of inhibiting both enzymes. Further docking studies of the six compounds synthesized helped to elucidate the main interactions that may be responsible for the inhibitory activities observed.

Black pepper or Piper nigrum is a well-known spice, rich in a variety of bioactive compounds, and widely used in many cuisines across the world. In the Indian traditional systems of medicine, it is used to treat gastric and respiratory ailments. The purpose of this investigation is to study the antihyperlipidemic and antiobesity effects of piperonal in high-fat diet (HFD)-induced obese rats.
Piperonal, an active constituent of Piper nigrum seeds, was isolated and confirmed by HPLC, 1H and 13C NMR spectroscopy. Male SD rats were fed on HFD for 22 weeks; Piperonal was supplemented from the 16th week as mentioned in the experimental design. Changes in body weight and body composition were measured by TOBEC, bone mineral composition and density were measured by DXA, and adipose tissue distribution was measured by 7 T-MRI. Plasma levels of glucose, insulin, insulin resistance and lipid profiles of plasma, liver and kidney, adipocyte hormones and liver antioxidants were evaluated using standard kit methods. Expression levels of adipogenic and lipogenic genes, such as PPAR-γ, FAS, Fab-4, UCP-2, SREBP-1c, ACC, HMG-COA and TNF-α were measured by RT-PCR. Histopathological examination of adipose and liver tissues was also carried out in experimental rats.
HFD substantially induced body weight, fat%, adipocyte size, circulatory and tissue lipid profiles. It elevated the plasma levels of insulin, insulin resistance and leptin but decreased the levels of adiponectin, BMC and BMD. Increased expression of PPAR-γ, FAS, Fab-4, UCP-2, SREBP-1c, ACC, and TNF-α was noticed in HFD-fed rats. However, supplementation of piperonal (20, 30 and 40 mg/kg b.wt) for 42 days considerably and dose-dependently attenuated the HFD-induced alterations, with the maximum therapeutic activity being noticed at 40 mg/kg b.wt.
Piperonal significantly attenuated HFD-induced body weight and biochemical changes through modulation of key lipid metabolizing and obesogenic genes. Our findings demonstrate the efficacy of piperonal as a potent antiobesity agent, provide scientific evidence for its traditional use and suggest the possible mechanism of action.