Caracterização do cDNA da NAD-MDH citosólica e da H+ ATPase Vacuolar no amadurecimento da anona (Annona cherimola Mill)

A Annona cherimola é um fruto exótico, com um sabor agradável. Este fruto tem um elevado potencial comercial, mas apresenta um tempo médio de vida curto devido ao seu rápido amadurecimento. Por esta razão é necessário conhecer melhor o processo de amadurecimento deste fruto. Na Região Autónoma da Madeira a cultura da anoneira é muito importante em termos comerciais. O processo de amadurecimento leva a diversas modificações bioquímicas e fisiológicas. Existem várias enzimas e substâncias que integram este processo. Neste trabalho iremos estudar os genes das enzimas malato desidrogenase e H+ ATPase vacuolar que estão envolvidos no processo de amadurecimento dos frutos. Utilizando as técnicas de RACE e sequenciação foi possível determinar a sequência nucleotídica do cDNA destes genes. O cDNA da malato desidrogenase é composto por 1364 nucleótidos, contendo uma zona 5’ UTR com 84 nucleótidos, uma zona 3’ UTR com 284 nucleótidos e um sinal de poliadenilação com a sequência AATAAA. A ORF apresenta 996 nucleótidos, codificando uma proteína com 332 aminoácidos. Para a H+ ATPase vacuolar foi amplificado o cDNA da subunidade C do domínio V1. Esta apresenta 799 nucleótidos, dos quais 36 são da 5’ UTR, 266 da 3’ UTR e 498 da ORF. A ORF codifica uma proteína com 166 aminoácidos.

Comparative analysis of the volatile fraction from Annona cherimola Mill. cultivars by solid-phase microextraction and gas chromatography–quadrupole mass spectrometry detection

The analysis of volatile compounds in Funchal, Madeira, Mateus and Perry Vidal cultivars of Annona cherimola Mill. (cherimoya) was carried out by headspace solid-phase microextraction (HS-SPME) combined with gas chromatography–quadrupole mass spectrometry detection (GC–qMSD). HS-SPME technique was optimized in terms of fibre selection, extraction time, extraction temperature and sample amount to reach the best extraction efficiency. The best result was obtained with 2 g of sample, using a divinylbenzene/carboxen/polydimethylsiloxane (DVB/CAR/PDMS) fibre for 30 min at 30 °C under constant magnetic stirring (800 rpm). After optimization of the extraction methodology, all the cherimoya samples were analysed with the best conditions that allowed to identify about 60 volatile compounds. The major compounds identified in the four cherimoya cultivars were methyl butanoate, butyl butanoate, 3-methylbutyl butanoate, 3-methylbutyl 3-methylbutanoate and 5-hydroxymethyl-2-furfural. These compounds represent 69.08 ± 5.22%, 56.56 ± 15.36%, 56.69 ± 9.28% and 71.82 ± 1.29% of the total volatiles for Funchal, Madeira, Mateus and Perry Vidal cultivars, respectively. This study showed that each cherimoya cultivars have 40 common compounds, corresponding to different chemical families, namely terpenes, esters, alcohols, fatty acids and carbonyl compounds and using PCA, the volatile composition in terms of average peak areas, provided a suitable tool to differentiate among the cherimoya cultivars.

Integrated valorization of Anona Cherimola Mill. seeds

Agricultural and agro-industrial residues are often considered both an environmental and an economical problem. Therefore, a paradigm shift is needed, assuming residues as biorefinery feedstocks. In this work cherimoya (Annona cherimola Mill.) seeds, which are lipid-rich (ca. 30%) and have a significant lignocellulosic fraction, were used as an example of a residue without any current valorization. Firstly, the lipid fraction was obtained by solvent extraction. Extraction yield varied from 13% to 28%, according to the extraction method and time, and solvent purity. This oil was converted into biodiesel (by base-catalyzed transesterification), yielding 76 g FAME/100 g oil. The obtained biodiesel is likely to be incorporated in the commercial chain, according to the EN14214 standard. The remaining lignocellulosic fraction was subjected to two alternative fractionation processes for the selective recovery of hemicellulose, aiming different products. Empirical mathematical models were developed for both processes, aiming future scale-up. Autohydrolysis rendered essentially oligosaccharides (10 gL-1) with properties indicating potential food/feed/pharmacological applications. The remaining solid was enzymatically saccharified, reaching a saccharification yield of 83%. The hydrolyzate obtained by dilute acid hydrolysis contained mostly monosaccharides, mainly xylose (26 gL-1), glucose (10 gL-1) and arabinose (3 gL-1), and had low content of microbial growth inhibitors. This hydrolyzate has proven to be appropriate to be used as culture media for exopolisaccharide production, using bacteria or microbial consortia. The maximum conversion of monosaccharides into xanthan gum was 0.87 g/g and kefiran maximum productivity was 0.07 g.(Lh)-1. This work shows the technical feasibility of using cherimoya seeds, and materials as such, as potential feedstocks, opening new perspectives for upgrading them in the biorefinery framework.