Conservation, Utilization, Genetic and
Chemodiversity of Germplasm of Genipap
(Genipa Americana L.) in Brazil
Alex Souza De Jesus1, Jose Guedes Da Sena Filho2, Adenir Vieira Teodoro2, Milena Nascimento Cardoso1, Ana Letícia Sirqueira Nascimento1, Ana Da Silva Ledo2 and Ana Veruska Cruz Da Silva2*
1Universidade Federal de Sergipe, Brazil
2Embrapa Tabuleiros Costeiros, Brazil
Submission: February 21, 2019; Published: March 22, 2019
*Corresponding author: Ana Veruska Cruz da Silva, Embrapa Tabuleiros Costeiros, Brazil
How to cite this article: Alex S D Jesus, Jose G D S F, Adenir V T, Milena N C, Ana Letícia S N, et al. Conservation, Utilization, Genetic and Chemodiversity
of Germplasm of Genipap (Genipa Americana L.) in Brazil. Curr Trends Biomedical Eng & Biosci. 2019; 18(4): 555995.
Genipa americana L. or genipap (Rubiaceae) is brazilian native species. Its leaves and fruits are used in the folk medicine and are greatly consumed in natura in the form of sweets, juices, composts and wines. Moreover, it can be used for the recovery of contaminated soils, in civil and naval construction, dye production for fabric dyeing, ceramic artifacts and non-permanent tattoos. Phytochemical analysis from the leaves and fruits reported in the literature detected the presence of iridoids, monoterpenoids, fatty acids, steroids, carboxylic acids and phenolic compounds, some of which bioactive compounds.
Despite the economic and ecological importance, this species has suffered a considerable loss of genetic variability caused, mainly, by the expansion of farming areas in the Brazilian. The genetic diversity and structure of a Genipap Genebank and natural populations has been evaluated using molecular markers, generating data useful to help programs of plant breeding, prospection and conservation. Application of techniques of tissue culture overcome the difficulties imposed by the traditional genipap propagation system, enabling large-scale multiplication and the conservation of this Rubiaceae.
Genipap (Genipa americana L. - Rubiaceae) is a species widely distributed in tropical Central and South America, including the Cerrado biome . Is a native from Amazon region, with medicinal importance and is widely used in Brazil . This tree is one of the most promising for sustainable harvesting programs and restoration of Brazilian riparian forests  (Figure 1). G. americana has economic and environmental importance, being valued for food production, recuperation of degraded areas, in the composition in areas of permanent preservation and agroforestry [3,4]. Your wood is widely used in civil and naval construction, and carpentry .
Extractivism is the most common form of exploration of this species. Commercial uses, however, has been encouraged and is especially recommended in the recovery of degraded areas and contaminated soils . In addition, the commercialization as fresh fruit has promising development, being held in the free trade fairs, in wholesale markets and supermarkets . They are alternative
foods for the population who live in these regions, contributing considerably to tropical forests conservation .
Unripe Fruits (green color) produces a blue to black juice that is used to tattoo the body , whereas ripe fruits were greenish-brown and possessed a soft, thin and wrinkled exocarp. The mesocarp is fleshy, yellow, and sweet and each fruit contained 105 (±10.7) seeds on average .
The tree is heliophytic, semideciduous, and selectively hygrophytic, occurring in areas with open forests and secondary vegetation of floodplains located in temporary or permanent locations flooded . It is a late secondary, with climax characteristics, moderate growth and can be used urban afforestation.
Besides the ecological and economic importance, it is worth mentioning the medicinal various parts of the plant being used in the medicine in many regions of the country . Commercializing
and validation of its benefits is employed in a folk medicine .
The fruits, leaves and bark are indicated in cases of cough,
anemia, bruises, dislocations, as well as purgative, diuretic,
osteoporosis, stomach problems, nervousness, diabetes,
cholesterol, besides being excellent tonic in the fight against the
indisposition, fatigue and weakness . Satisfactory amounts of
iridoids, tannins, and genipacetol, with pharmacological activity
Roots showed antigonorrheal properties and the
decoction its leaves are used for diarrhea and syphilis . An
ethnopharmacological survey carried out in the rural area of Serra
da Raiz, Paraíba state, Northeastern Brazil, showed that genipap
is one of the main species that are widely used in the treatment
of various diseases, such as respiratory problems, inflammation
and blood diseases . Moreover, because of its high mannitol
content, the fruit is recommended for its antihypertensive
properties . In the industry, it is employed to production of hair,
skin products and natural skin dyes.
In North and Northeast regions of Brazil, fruits have potential
due to high nutritive value and lipid content . Genipap in natura
have considerable moisture values, e.g. total carbohydrates,
mainly dietary fiber, vitamin C and phenolic compounds and high
antioxidant activity. Therefore, the inclusion of genipap in the diet
could improve its quality, contributing to food and nutritional
security of the cerrado population. In food it can be used in natura
in preparations or in beverages e. g. jams, candies, crystallized,
liqueurs, juice, wine and brandy .
The first studies of isolation, characterization, and structural
elucidation of the bioactive compounds of genipap were the
iridoid genipin in the early 60s . In isolated genipic acid and
genipinic acid obtained from fruit extracts . Geniposidic acid
was the only glycosylated iridoid isolated from the methanolic
extract of G. americana leaves until the present moment .
Later, the isolation of the geniposid was carried out and the
tarennoside, gardenoside, and geniposidic acid were obtained by
callus formation and culture of suspended cells .
Steroids have been reported in the composition of fruits
extract, but not confirmed the occurrence of sesquiterpene,
flavonoids, or other secondary compounds (Table 1). The absence
of terpenoids, specially the sesquiterpenes may be explained by
some aspects of plant collection, such as botanic diversity or stage
of maturation of the fruit . For the Recôncavo Baiano Region
there are variability in the chemical constituents of genipap fruit
characters, such as, pH, soluble solids content and ascorbic acid.
This data suggest economic exploitation of fruits for consumption
in natura and industrialization .
The influence of the ripeness stage of genipap fruits on their
iridoid and phenolic compound composition by high-performance
liquid chromatography connected in series to a diode array
detector and mass spectrometer HPLC-DAD-MSn, was performed.
A total of 17 compounds, among them, compound phenolic and
iridoids were identified in genipap fruits, native to the Amazon
The volatile fraction of the Genipap fruits has high amounts
of carboxylic acids (butyric, 2-methylbutyric and hexanoic acids)
and 2- and 3-methylbutyric acids, which are probably related
to the strong and primary odor of genipap . Seven new
iridoid called genamesides A-D, genipacetal, genipamide and
genipaol were isolated from its fruits along with eight known
iridoid glucosides: geniposidic acid, geniposide, gardenoside and genipin-gentiobioside, genipin, gardendiol, deacetyl aperuloside
acid methyl ester and shanzhiside were reported in the fruits
[19,20]. Besides the identified of carboxylic acids, alcohols, esters
and aromatic compounds, in the volatile fraction, as well as the
monoterpenes: linalool and limonene [22,23].
In this context, the high chemodiversity of iridoids in the fruits
and the nutritional source of this specie shed a light to investigate
the secondary metabolites of the G. americana, in order to provide
data to characterize the available germplasm to serve as a basis
for the production of bioactive compounds with agroindustrial
and pharmacological potential. Nowadays, techniques are
exploring bioactivity compounds and new drugs formulations
are being made in medicinal plants, but the understanding of the
biosynthesis and the genetic variability this species need to be
The farming expansion areas in the Brazilian has been
caused considerable loss of variability genetic, although genipap
occurs natural reserves and indigenous lands. Genetic markers
can used in forest ecosystems enables advances in the genetic
understanding of the natural populations, as well as assists on
strategies to recover or restore forests .
Regarding the high potential of the genipap for conservation
of its genotypes, the Genipap Genebank was created in 2009 at the
Brazilian Enterprise for Agricultural Research-Embrapa Coastal
Tablelands, in the city of Nossa Senhora das Dores, in the state of
Sergipe, Brazil (10°27’50,0” S and 37°11’39,5” W), consisting of 160
accessions from wild populations (Figure 2). Genipap Genebank is
accredited by the Genetic Heritage Management Council (CGEN),
a normative and deliberative organ of the Brazilian Ministry of
Environment (MMA), as faithful custodian of the genipap genetic
patrimony (process number 2000.002275/2013-36) .
The Genebank of Embrapa is compound by just two species:
G.americana L. and G. infundibuliformis Zappi & Semir (Table
2). The development of studies of chemical and biological
characterization accessions that compose the genotype genebank
can generate data useful to help programs of plant breeding.
The activities developed include prospection, collection, ex situ
conservation, development of in vitro conservation protocols,
morphological and molecular descriptor-based characterization,
filing and documentation.
The genetic diversity and structure of genebank was evaluated
using inter simple sequence repeat (ISSR) markers. The data
generated from 12 ISSR primers were used to determine genetic
variability via a model-based Bayesian procedure (Structure) and
molecular variance analysis . A total of 12 primers were used,
which generated 123 polymorphic fragments. Four groups were
formed from the analysis of the fragments, and the CR1-2 genotype
was isolated, being more divergent than the other genotypes .
This data can be used in the reclassification of accessions in future
breeding and conservation programs.
Recent manuscript reported the mendelian inheritance,
genetic linkage, and genotypic disequilibrium at six microsatellite
loci developed for G. americana . Results detect genetic linkage
between pairwise loci in 54.4% of the tests, but no genotypic
disequilibrium was detected between pairwise loci for adult trees
and regenerants .
The molecular characterization of accessions was performed
using 15 inter-simple sequence repeat (ISSR) primers, and
genotype clustering was performed based on genetic dissimilarity
using the Unweighted Pair-Group Method with Arithmetic Mean
(UPGMA). The NB (Núcleo Bandeirante) accession is genetically
different from the CZA (Cruz das Almas) and AS (Sabinópolis)
populations and shows superiority for most of the variables
In Sergipe state, in the lower course of São Francisco River,
the genetic diversity among 18 individuals of G. americana by
RAPD markers using 12 primers, was evaluate. Results showed
the Jaccard index, the average genetic similarity (Ags) among
individuals was 60.4%, while the highest similarity distance
obtained is between individuals G11 and G12 (83.6% ± 0.03) and
the lowest genetic similarity between individuals G4 and G18
(36.5% ± 0.02). Seven pairs were considered genetically similar
based on the Ags 78.4% . This data can be used to provide
seeds in combination with other individuals in restoration
programs for degraded areas and adequate management of forest
A construction of cDNA library generated 165 expressed
sequence tags (ESTs), in G. americana plants cultivated in nutritive
solution. The results show 81 homology to genes deposited in the
NCBI database, 67 did not show similarity to any available gene,
and 17 ESTs demonstrated homology with unknown genes .
These genes probably associated with proteins and enzymes
related to the defense mechanisms under biotic and abiotic
The genetic variability for the biochemical characteristics of
seeds was performed in two populations: one from Ilha Solteira,
SP region (ISA) and other from Mogi Guaçu, SP (MOG) show a
great genetic variation. . Narrow sense heritability estimates
of progenies were moderate (0.69 for carbohydrates in the
population ISA) to high (0.81 to 0.99 for the other traits in the two
populations), indicating that much progress can be expected with
selection strategies .
The use of biochemical and molecular markers for
characterization of genotypes useful for generating information
in future utility in specific breeding program and allows for the
identification of duplicates and germplasm exchange between
Indigenous fruits are very important as sources of food
security, balanced households nutrition . Studies the seed
bank species is important information for the conservation
strategy in situ. In addition, induce seed bank can contribute for
their regeneration, maintenance and establishment of seedlings
. G. americana seeds forms a transitory seed bank, with
maintenance of its viability until the fourth month after disposal
in natural environment. In general, the seed bank located in
the riparian forest area showed the best potential for seedling
establishment in a natural environment, which supports the
ecological behavior of this species .
G. americana showed easy propagation by seeds, presenting
between 83 and 92% of emergency. However, the process is slow,
asynchronous and with low uniformity. These characteristics
are sources of great heterogeneity in the development of the
plants, which may hinder the conduction of cultural treatments in
nurseries producing seedlings . Another factor that affected
to vigor and germination is the water restriction. Understanding
of plant tolerance to drought is important especially for recovery
of areas where there are problems with water stress for reasons
climatic conditions or soil type conditions . Moreover, this
information is important because genipap is found riparian forest
regions, is subject to flooding and dried, that is, its fruits are
exposed to variations conditions that affect watercourses, which
may compromise the maintenance of the species throughout time
Genipap is classified as intermediate, because do not support
low temperatures for extended periods. In function of this, the best strategy is used the filed collections, not being indicated
conservation way seed bank [4, 26]. On the other hand, the
application of techniques of tissue culture is important for
species such as genipap to overcome the difficulties imposed
by the traditional propagation system, enabling large-scale
multiplication and the conservation of this species with holds
good prospect for the future (Figure 3) [35-37]. Domestication
of these trees can be successful only if successful propagation
techniques are applied . Therefore, the optimal conditions for
in vitro germination of G. americana seeds requires pre-soaking
in distilled water for 48hours and inoculation into culture media
consisting of ½ MS + 15 g L-1 sucrose, with stratification in the
dark for 16days, followed by the transfer to growth chambers
with lighting provided by white fluorescent lamps .
The temperature for germination for genipap is 25 °C, 30 °C
and 35 °C and the substrate is vermiculite and soil substrates were
the best conditions for seed germination . The substrates
consisting of washed sand + dried coconut shell powder, at a 1:1
ratio by volume, and Topstrato HT® may be recommended for
genipapo acclimatization Sabinópolis (AS), Cruz das Almas (CZA)
and Núcleo Bandeirante (NB) accessions .
Drying seed genipap can be done in a greenhouse (temperature
of 33 °C with relative humidity of 70%) for 24 hours or laboratory
environment (temperature of 28 °C with relative humidity of 75%)
for 48 hours without compromising the physiological quality .
For the immersed seeds in PEG solutions, the potential -0.3MPa
allowed the osmotic conditioning, while the -0.4 MPa interfered
in vigor. In imbibed seeds in paper there was a germination
reduction, because the time, until potential events were not
observed from -0.3 MPa .
The of plant tolerance to drought and how to exploit them,
especially with regard to problems of a physiological or ecological
order, extremely important, especially for the recovery of areas
where there are problems with water stress for reasons climatic
conditions or soil type conditions.
Culture media supplementation with 1.0 mgL-1 BAP favors
the development of adventitious shoots and promotes a higher
callogenesis percentage, leaf number and feasibility at 90 days of
in vitro culture . The immersion of the basal portion of shoots
for 20 seconds in all IBA solutions induces the rhizogenesis in
the genipap accessions at 90 days of in vitro culture . In MS
medium supplemented different concentration NAA in presence
of BAP is possible in vitro regeneration of genipap from the
conversion of zygotic embryos .
The cryoprotection and dehydration does not modify
the viability of the shoot apices of genipap encapsulated or
unencapsulated. Immersion for 24h in cryoprotecting solution
and the dehydration in a laminar flow cab by 2h have potential
for use in future studies of cryopreservation by encapsulationdehydration
. Due to potential use of genipap as food, technical
strategies is need to use of tissue culture, to be used in mass
propagation in large quantity.
Due to the recognized popular use of G. americana and to
be a source of molecules with biological potential, research on
conservation, management, sustainable utilization, marketing,
ethnobotany, ethnopharmacology, food chemistry, pharmacology
and genomics it is necessary more research for generate products
of sustainable manner, low cost and environment friendly. Besides
in understanding the chemodiversity of this particular species
with a huge variety compounds.