Trabalho apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Ciências Farmacêuticas; O Arbutus unedo L., vulgarmente designado por medronheiro, é uma planta
endémica da região Mediterrânica amplamente utilizada na alimentação e na medicina
tradicional devido às suas actividades biológicas. Estudos fitoquímicos realizados nesta
espécie evidenciaram que os extractos da folha e fruto contêm diversos antioxidantes
naturais. As folhas apresentam na sua constituição compostos fenólicos, tais como
taninos, flavonóides e glicosídeos fenólicos e α-tocoferol, enquanto os frutos possuem
também variados compostos fenólicos (antocianinas, derivados do ácido gálhico,
taninos e flavonóides), vitamina C e E e carotenos. Assim, é de prever que a espécie
Arbutus unedo L. tenha um considerável potencial antioxidante. No entanto, as
propriedades antioxidantes desta espécie em membranas biológicas não foram até então
Por esta razão, o presente trabalho deteve-se na avaliação do efeito protector dos
extractos aquosos da folha e fruto de Arbutus unedo L. na danificação oxidativa
induzida por radicais livres, utilizando como modelo celular in vitro o eritrócito
humano. O 2...
Despite strawberry tree (Arbutus unedo L.) leaves had a long use in traditional medicine due to its antiseptic,
diuretic, astringent and depurative properties, the potential of their antioxidant activity are still
lacking. Our study goals to assess the antioxidant and free radical scavenging potential of water, ethanol,
methanol and diethyl ether extracts of A. unedo leaves. Total phenols content was achieved spectrophotometrically
using Folin–Ciocalteau reagent with gallic acid as standard. Antioxidant activity was evaluated
using three different methods: reducing power of iron (III)/ferricyanide complex assay, scavenging
effect on DPPH (2,2-diphenyl-1-picrylhydrazyl) radicals and scavenging effect on superoxide radicals by
using the PMS–NADH–nitroblue tetrazolium system. Ethanol extracts of A. unedo leaves were the highest
in reducing power (IC50 232.7 lg/mL) and DPPH scavenging effect (IC50 63.2 lg/mL) followed by water
extracts (with IC50 of 287.7 and 73.7 lg/mL, respectively); whereas diethyl ether extracts were the lowest.
In the scavenging on superoxide radical assay, methanol extracts obtained the best results (IC50
6.9 lg/mL). For all the methods tested the antioxidant activity was concentration dependent. In accordance
with antioxidant activity...
O medronheiro (Arbutus unedo L.) é uma árvore fruteira de elevado valor ornamental, ambiental, económico e medicinal devido às propriedades atribuídas às flores, frutos e folhas. No entanto, nos últimos anos não tem sido atribuída a devida importância a esta espécie na região de Trás-os-Montes, ao contrário do que se verifica no resto do país, registando-se um declínio da área ocupada por A. unedo nesta região. Com o objectivo de preservar e potenciar a produção desta espécie procedeu-se, numa primeira fase, à avaliação da diversidade de 4 populações naturais de A. unedo do interior Norte e Centro de Portugal, através da caracterização morfológica e molecular. Numa segunda fase, avaliaram-se as propriedades químicas de 19 genótipos provenientes da região de Bragança, com vista a seleccionar os de superior qualidade. Desta forma espera-se contribuir para a multiplicação e valorização de populações de maior interesse.
A diversidade de populações naturais de A. unedo foi avaliada através da análise morfológica e genética pela utilização de marcadores moleculares, num total de 46 genótipos. Para tal, foram colhidas 40 folhas aleatoriamente em cada árvore, nas quais se mediu o comprimento...
In the Mediterranean Basin, the strawberry tree (Arbutus unedo L.) has medicinal, ornamental, economical, and environmental importance. During the last decades, several occurrences have caused the decline of the strawberry tree in Portugal being replaced by other species. In this context, the knowledge of genetic variability in the wild populations is essential for a proper conservation. This work aimed to characterize genetically and morphological 38 A. unedo genotypes, 30 of which located in Trás-os-Montes e Alto Douro region, and the remaining 8 genotypes in Beira Interior region, to assess their biodiversity.
The genetic diversity was assessing by RAPD (Random Amplification of Polymorphic DNA) markers. The dendrogram was obtained from the matrix of pairwise distances through the Nei and Li coefficient using the UPGMA (Unweighted Pair Group Method with Arithmetic Mean) grouping method. Morphological characterization was performed through the evaluation of several quantitative traits measured on 40 fresh leaves per plant. The quantitative traits analysed were leave length and width, leave length/width ratio, peduncle length and leave fresh and dry weigh. Distance matrix of morphologic data was assessed using Euclidean distance. A Mantel test was used to analyze correlations between genetic...
A espécie Arbutus unedo L., vulgarmente conhecida como medronheiro, é uma árvore de pequenas dimensões que surge dispersa em todo o território nacional. Vários usos tradicionais, sobretudo no âmbito medicinal e alimentar, têm sido citados para os seus frutos (medronho), folhas e raízes. Contudo, a sua potencialidade neste âmbito não tem sido devidamente explorado, o que tem contribuído para a redução da área ocupada por esta espécie nos povoamentos florestais.
O presente trabalho teve como principal objetivo complementar os conhecimentos sobre os usos tradicionais do medronheiro, pela exploração do seu potencial nutricional e nutracêutico, de maneira a utilizá-los em processos de desenvolvimento tecnológico valorizando-se, desta forma, a espécie. Para tal, foi avaliada a atividade antioxidante das folhas, frutos e, pela primeira vez, das cascas e das flores do medronheiro. Nesta análise recorreu-se a três ensaios: efeito bloqueador dos radicais livres 2,2-Difenil-1-picrihidrazilo (DPPH), poder redutor e capacidade redutora total. Nos frutos avaliou-se, adicionalmente, a sua cor e a composição nutricional e nutracêutica, bem como a influência do estado de maturação nestes parâmetros. A análise nutricional incluiu a avaliação da humidade...
Arbutus unedo has been long used in folk medicine, throughout Mediterranean countries, with the employment of
infusions and decoctions of almost all parts of this plant: leaves, fruits, barks and roots. The application of these traditional
remedies arises from several health-promoting characteristics, for treatment of gastrointestinal and urological
problems, hypertension and cardiac diseases, diabetes and as anti-inflammatory agent, among other interesting properties.
Antioxidant ability of A. unedo shrub is also known, and antimicrobial activity has also been reported. Several
compounds present in different parts of the plant may be linked to these properties. Included in those are carotenoids,
flavonoids, phenolic acids and vitamins (C and E). Other bioactive compounds may be also found in different parts
of A. unedo, like terpenoids and organic acids. This review will focus on the known composition of several parts of
A. unedo, their antioxidant ability and traditional use, and the available data sustaining the rationality of the use as part
of folk medicine.
Arbutus unedo L. is a Mediterranean shrub used in the production of jams, marmalades and beverages, as
well as in traditional medicine. In this work, the effect of ripening on the volatile compounds of the fruits
of A. unedo was evaluated, using headspace-solid phase microextraction combined with gas chromatography/
ion trap-mass spectrometry. Alcohols are the main component of the volatile fraction of A. unedo
fruits, followed by aldehydes and esters. The main volatile compounds identified were (Z)-3-hexen-1-ol,
1-hexanol, hexanal, (E)-2-hexenal and hexyl acetate, all formed through the lipoxygenase pathway (LOX).
The green odours are progressively replaced during ripening by floral and sweet sensations, due to the
decrease in alcohols, aldehydes and esters, resulting in the unmasking of the aromas associated with
the minor compounds (mainly monoterpenes and norisoprenoid compounds) that compose A. unedo
Arbutus unedo is a widespread shrub with economic importance, derived from the use of its berries in the
production of alcoholic beverages and in folk medicine. This work intends to evaluate for the first time the
effect of fruit ripening stage on antioxidant activity, total phenolic content, fatty acid profile and tocopherol
composition. Ripe fruits shown higher extraction yield (45.04±2.23%) when compared to other fruit ripening
stages. By contrast, total phenol contents were higher in the unripe and intermediate stage of ripeness (108
and 111 mg GAES/g dry fruit, respectively, against 60 mg/g dry fruit when ripe). Ripe and intermediate fruits
shown the lower EC50 values on the DPPH radicals (0.25±0.02 mg/mL) and reducing power assay (1.09±
0.05 mg/mL), respectively. A significant correlation was established between antioxidant activity and fruits
ripening stage. Fatty acid profiles were very similar between the ripening stages, being alfa-linolenic, linoleic
and oleic, the three major ones. Polyunsaturated fatty acids (PUFA) represent as much as 60% of the total fatty
acids, with a highly favorable omega 3/omega 6 ratio. From the analysis of the vitamin E vitamers, the most
important was γ-tocotrienol, with a clear reduction in the total free vitamin E content with ripening.
These results highlight that the fruits of intermediate ripeness can be regarded important sources of
biologically active compounds with a fatty acid profile rich in omega-3 PUFA...
Genetic analysis of plants relies on high yields of pure DNA. For the strawberry tree (Arbutus unedo) this represents a great challenge since leaves can accumulate large amounts of polysaccharides, polyphenols and secondary metabolites, which co-purify with DNA. For this specie, standard protocols do not produce efficient yields of high-quality amplifiable DNA. Here, we present for the first time an improved leaf-tissue protocol, based on the standard cetyl trimethyl ammonium bromide protocol, which yields large amounts of high-quality amplifiable DNA. Key steps in the optimized protocol are the addition of antioxidant compounds—namely polyvinyl pyrrolidone (PVP), 1,4-dithiothreitol (DTT) and 2-mercaptoethanol, in the extraction buffer; the increasing of CTAB (3%, w/v) and sodium chloride (2M) concentration; and an extraction with organic solvents (phenol and chloroform) with the incubation of samples on ice. Increasing the temperature for cell lyses to 70 °C also improved both DNA quality and yield. The yield of DNA extracted was 200.0 ± 78.0 μg/μL and the purity, evaluated by the ratio A260/A280, was 1.80 ± 0.021, indicative of minimal levels of contaminating metabolites. The quality of the DNA isolated was confirmed by random amplification polymorphism DNA and by inter-simple sequence repeat amplification...
The present study reports the antioxidant properties of Arbutus unedo L. leaf and fruit extracts using
different in vitro assays including (i) reducing power, (ii) scavenging effect on DPPH free radicals, and
(iii) inhibitory effect on AAPH-induced hemolysis and lipid peroxidation in human erythrocytes. All
assays demonstrated antioxidant efficiency for A. unedo L. aqueous extracts, being consistently higher
in the leaf. EC50 values for reducing power and DPPH radical scavenging activities were, respectively,
0.318 ± 0.007 and 0.087 ± 0.007 mg/mL for leaf, and 2.894 ± 0.049 and 0.790 ± 0.016 mg/mL for fruit
extracts. Under the oxidative action of AAPH, A. unedo leaf and fruit extracts protected the erythrocyte
membrane from hemolysis (IC50 of 0.062 ± 0.002 and 0.430 ± 0.091 mg/mL, respectively) and decreased
the levels of malondialdehyde, a breakdown product of lipid peroxidation (IC50 of 0.075 ± 0.014 and
0.732 ± 0.452 mg/mL, respectively). In accordance with antioxidant activity, phenolic content was found
to be significantly higher in leaf extract. To our knowledge, this is the first time that the antioxidant activity
of A. unedo species is evaluated using human biological membranes. Overall, our results suggest that A.
unedo leaves are a promising source of natural antioxidants with potential application in diseases mediated
by free radicals.
In recent years the strawberry tree (Arbutus unedo L.) is being gradually replaced by other species with
higher economic value. With the ultimate goal of selecting superior genotypes, the present work was
initiated to study the antioxidant and antimicrobial activities, and total phenolic content in 19 different
genotypes of A. unedo leaves from the Trás-os-Montes region of Portugal.
The genotype Braganc¸ a 1 contains higher total phenolic content (215.0 mg GAE/gextract) whereas the
Vila Boa 4 genotype shows lower total phenolic content (148.0 mg GAE/gextract). In both methods tested
to evaluate the antioxidant activity, Vila Verde and Donai displayed the highest antioxidant capacity
(EC50 values of 0.088 and 0.090 mg/mL, respectively, for DPPH; EC50 values of 0.233 and 0.245 mg/mL,
respectively, for reducing power assay) while Vila Boa 2 reported the lowest antioxidant potential (EC50
values of 0.142 and 0.378 mg/mL, respectively, in DPPH and reducing power methods). Linear negative
correlations were established between the total phenol contents and the EC50 values for both of the
antioxidant activity methods tested. Preliminary assays for antimicrobial potential showed that extracts
from A. unedo leaves display antibacterial activity...
The genetic diversity within and between four natural Arbutus unedo L. populations in Portugal was
investigated using morphological characters, inter-simple sequence repeats (ISSR) and random amplified
polymorphic (RAPD) markers, for a proper conservation of this resource. The results showed a high
morphological variation between and within-populations. The dry-weight, length, width and peduncle
length of the leaves were the most discriminating characters, being responsible for 52.6% of the total variation.
The molecular analysis, based on both RAPD and ISSR, indicated moderate genetic diversity within
populations, with values of Nei’s gene diversity ranging from 0.16 to 0.39 and the Shannon’s diversity
from 0.24 to 0.57. The highest genetic diversity occurred within Braganc¸ a population. At the species level,
the genetic variation was mainly within populations (75%), suggesting a low-moderate genetic differentiation
among populations. The high level of gene flow estimated among populations (1.499) showed to
be the main reason for their low differentiation. The cluster analysis showed that only one population
formed a distinctive cluster. The remaining formed a second cluster due to their geographical approach,
which has been showed to affect significantly the genetic distances among populations. No correlation
between RAPD and ISSR markers...
Arbutoid mycorrhizae were synthesized between adult selected
clones of Arbutus unedo L. and Pisolithus arhizus. Two micropropagated
clones were tested: AL1, in vitro and C1 (acclimatized plants)
in nursery and later in a field trial. In vitro, rooted shoots were transferred
to test tubes containing the substrate previously inoculated with
mycelium cultured on agar. In the nursery, two inoculation treatments
were tested (vegetative inocula or dry sporocarps) and compared to control
plants. In the field trial, plants from nursery inoculation treatments
were compared and an additional control treatment using seedlings was
implemented. Plant height was evaluated 4 months later in the nursery
and 20 months later in the field trial. Roots were examined by morphological
and histological studies: a) in vitro plantlets one month after
inoculation and nine months after acclimatization; and b) 20 months after
the field trial was established. Arbutoid mycorrhizae were observed in
vitro one month after inoculation, indicating compatibility between A.
unedo and P. arhizus. These showed the presence of a mantle, Hartig net,
and intracellular hyphal complexes confined to the epidermal root cells.
Arbutoid mycorrhizae were also observed nine months after acclimatiza-
Fund project: This work was supported by a PhD fellowship
(SFRH/BD/37170/2007) from the Portuguese Foundation for Science
and Technology (FCT)
The online version is available at http://link.springer.com
Filomena Gomes ( ) • Esteban San Martin
Filomena Gomes. CERNAS...
The strawberry tree (A. unedo L.) is native from the Mediterranean region and it is implanted all over the Portuguese territory. In Portugal, A. unedo fruits have been used in the production of spirit, liqueurs and jam. Considering the wide distribution through Portuguese territory and the limited number of applications, it becomes crucial to value A. unedo fruits. For this purpose it is necessary to understand the chemical composition of A. unedo fruits, particularly during maturation and storage. Therefore, some quality parameters of A. unedo fruits were studied throughout maturation, particularly ºBrix, pH, titratable acidity, total phenolic content and antioxidant activity. In addition, the lipophilic extractives profile was assessed in ripen fruits. To promote the consumption of A. unedo fresh fruits it was studied the effect of cold storage (≈ 5 ºC) using fruits from two different ripeness degrees (19 and 21 ºBrix). The final phase of this thesis was focused on innovation and development of new food products based on A. unedo fruits.
This study allowed us to conclude that throughout maturation, the ºBrix content and pH increased, whereas titratable acidity, total phenolic content and the antioxidant activity exhibited no remarkable tendency. Moreover...
Mestrado em Engenharia do Ambiente - Tecnologias Ambientais - Instituto Superior de Agronomia; Panasqueira mine is one of the last three mines actually in labour. Tungsten, copper
and tin are explored for over a century. The visual and chemical impact of the exploration is
due to active and old huge tailings and due to draining from mine, water plant treatment and
seepage water. The objective of this study was to evaluate the biogeochemical impact of the
mine activity in soil, water and strawberry tree (Arbutus unedo L.).
Soil is contaminated with copper, zinc, arsenic, tungsten and lead and the
consequent impact was observed in microbial soil enzymes. Selective chemical extraction of
trace elements allows to assess elements in soils and their absorption in plant roots and
translocation to branches, leaves and fruits. Arbutus unedo is a manganese and zinc
tolerant, and is not accumulator of trace elements. However, this species presented high
concentration of cadmium in the aerial parts. The analysis of sediments and water from
Ribeira of Bodelhão indicate that trace elements are dispersed from the primary
contaminated area, to Zêzere River, where the dilution drastically eliminates the
The strawberry tree fruits can be used to produce alcohol as they are not
contaminated. A. unedo can be used in phytostabilisation programs of contaminated soils
Mestrado em Engenharia Agronómica - Instituto Superior de Agronomia; The strawberry tree (Arbutus unedo L.) is native to the Mediterranean region. The
use of molecular markers in this species has been limited to the use of RAPDs,
ISSRs as well to the cross-amplification of SSRs from other Ericaceae. In this work,
we developed a protocol for extracting nuclear DNA from the strawberry tree and
performed partial next-generation sequencing of the Arbutus unedo L. genome using
the "Ion Torrent" (Life Technologies) platform. The next-generation sequencing
resulted in 198,856 sequences ("raw data") with an average size of 123 bp, which
were uploaded to the NCBI database "Sequence Read Archive" (SRA) with the
accession number: SRX341237. Data analysis led to the identification of 1085
microsatellite-containing sequences, which were also uploaded with accession
numbers: from KF023636 to KF024720 to the NCBI databases. Primers were
designed for 18 microsatellite loci of which only three have proved to be polymorphic
in a panel of 16 samples. Based on identified 25 SNPs one CAPS marker was
developed, which despite being heterozygous revealed to be monomorphic among
the 16 analyzed samples.
Mestrado em Engenharia Agronómica - Instituto Superior de Agronomia; This study had two objectives that aim to contribute to a greater knowledge of Arbutus unedo. The first objective was to characterize morphologically different clones installed in a commercial orchard in Oleiros, Castelo Branco, in order to detect the morphological differences between them, as well as to monitor the vegetative cycle. The second objective was to study the physiological behavior of the fruit in the post-harvest period, in order to better assess the evolution of the maturation.
The clones showed significant differences in the cross sectional area of the trunk and in the canopy height. Leaves showed significant differences in the length (L), width (W), L/W ratio, petiole length and leaf thickness.
The percentage of trees with fruits, the average number of inflorescences and the number of fruits and shoots per inflorescence were significantly different. Fruits at different stages of maturity showed significant differences in firmness, L, hue and saturation. Except for the green fruits, all states of maturation showed an increase in the soluble solids content and pH while titratable acidity decreased. It was not found a marked evolution of vitamin C and total phenolic contents in the postharvest period.
Considering the above...
Dissertação mest., Agricultura sustentável, Universidade do Algarve, 2007; O presente trabalho teve como principal objectivo caracterizar quimicamente dois frutos
silvestres, o fruto de Arbutus unedo L (medronho) e o fruto de Rubus fruticosus Agg.
(amora), que crescem espontaneamente na Serra do Caldeirão.
Os parâmetros físico-químicos e bioquímicos estudados foram o pH, o ºBrix, a
humidade, a cinza total, os iões cálcio, magnésio, sódio e potássio, o azoto total, a
proteína total, os açúcares (frutose, glucose e sacarose) e as antocianinas (delfinidina 3-
glucosido, delfinidina 3,5-diglucosido, cianidina 3-glucosido, cianidina 3,5-diglucosido,
pelargonidina 3-glucosido e pelargonidina 3,5- diglucosido). Foram ainda determinados
os parâmetros de cor no espaço CIE (L*, a* e b*). Observaram-se diferenças
significativas nas várias determinações físico-químicas realizadas dependendo do ano e
do local de colheita das amostras.
Os frutos de Arbutus unedo L apresentaram na sua constituição teores das antocianinas
delfinidina 3-glucosido e cianidina 3-glucosido de 0,44 mg e 0,13 mg por 100 g de fruto
fresco, respectivamente. Os frutos de Rubus fruticosus Agg. apresentaram na sua
composição concentrações das antocianinas cianidina 3-glucosido...
Dissertação mest., Qualidade em análises,
Universidade do Algarve, 2006; O trabalho apresentado nesta dissertação teve como objectivo a validação de um método de análise por espectrofotómetria de absorção atómica/emissão de chama para determinação de minerais na fermentação de medronho (Arbutus unedo L.).
O método estudado, foi desenvolvido e implementado no laboratório da FERN da Universidade do Algarve, sendo utilizado em rotina para quantificação da composição mineral de alguns produtos vegetais.
No contexto da validação do método, apresentam-se os resultados da avaliação de alguns dos seus parâmetros característicos e da sua adequação à finalidade pretendida.
A validação do método foi efectuada para 10 elementos minerais, tendo os elementos Ca, Cd, Cu, Fe, Mg, Mn, Pb e Zn sido validados por espectrofotómetria de absorção atómica e os elementos Na e K validados por emissão chama.
Posteriormente procedeu-se a quantificação destes elementos em amostras de medronho fermentado, proveniente de uma fermentação anaeróbia de frutos de medronho. O processo de fermentação foi acompanhado pela recolha de 10 amostras aos tempos t= 0; 10; 12; 17; 18; 19; 24; 32; 48 e 65 dias.
Verificou-se ser possível quantificar os elementos Ca...
Plant-water relationships, leaf morphology and concentrations of pigments, proline, soluble sugars, anthocyans and nutrients of after wildfire resprout leaves of Arbutus unedo L. were com-pared with those of same age sprout leaves in unbumed plants. In spring samples, several significant differences appear: after-fire resprouts showed some juvenile characteristics, such as high tissue elasticity, small cells, low specific leaf weight, high Chla/Chlb ratio and a high anthocyan content; on the other hand, unbnrned plants showed drought tolerance characteristics by osmotic adjustment and lower tissue elasticity, larger cells, higher specific leaf weight and lower Chla/Chlb ratio. In fall samples, both kind of shoots showed drought tolerance by osmotic adjustment, low tissue elasticity, large cells and high specific leaf weight. Results show that after-fire resprouts have, at the beginning of their development, higher morphological and physiological plasticity that the new sprouts of unburned plants.; Es comparen les relacions hídriques, la morfologia foliar i les concentracions de pigments, prolina, sucres solubles, antocians i nutrients de les fulles de rebrots d'Arbutus unedo L. després d'un incendi, amb iguals característiques de les fulles de brots de plantes no cremades de la mateixa edat. A les mostres de primavera apareixen diverses diferències significatives: els rebrots...