This document discusses a research project focused on Jatropha curcas and mycorrhizal fungi. The project aims to: 1) Create a germplasm bank of diverse J. curcas and mycorrhizal fungi accessions for biodiesel production. 2) Develop micropropagated J. curcas plants inoculated with mycorrhizal fungi to increase oil yields and productivity. 3) Eliminate the toxicity in J. curcas seed cake waste to allow its use as animal feed. The project is led by BioVale Energia in partnership with universities and research organizations in Brazil.

2. Jatropha-curcas: R&D project

3. Business focus
3
Agronomy, Science
 Sustainable plantation, social inclusion
for Oils Supply Programme
 Collection of accessions – Germoplasm
bank
 FMA for productivity and plant health
 Jatropha detoxification process
 Fungi production
Industry
 Jatropha planting to crude oil
 Planting, maintenance and
harvesting
 Expelling, preprocessing and
logistics
 Crude oil sales
Biovale Energia

4.  To set up a mycorrhizal germplasm reservoir of genetic diversity of
mycorrhizal fungi of jatropha-curcas with high oil yielding for biodiesel
production and the elimination of the toxicity of the meal]waste for
animal feed
 Executing organization :
BIOVALE ENERGIA – an emerging biodiesel company based in the Minas
Gerais in partnership with various stakeholders in the oil production chain
 Co-executing organizations:
Federal University of Viçosa – DMB/BIOAGRO, ranked as the best university
of Minas Gerais State and 3rd in Brazil
SECTES – Minas Gerais State Science and Technology Secretariat
EPAMIG – EMPRESA DE PESQUISA DE AGROPECUÁRIA DE MINAS, national
bench-mark in jatropha development
CENTRO TECNOLÓGICO DE MINAS GERAIS - CETEC
R&D goals and stakeholders

5. Biofuel production
has become
substantially more
efficient over the last
25 years as Brazil and
the United States
have scaled up their
industries.
Such incremental
gains are likely to
continue for years to
come.
New Technologies, New Gains
However, the greatest potential for biofuels lies in the
development of new technologies that will significantly expand
the range of biomass feedstock, increase conversion efficiencies,
and lower production costs.

6. Documentation, preservation and characterization of J. curcas
and mycorrhizal fungi germplasm
Development of micropropagated plants inoculated of ROC (Root
Organ Culture) of arbuscular mycorrhizal (AM) fungi and their in
vitro mass inoculum production
Selection of jatropha genotypes with high oil yielding, envisaging
the biodiesel production
Toxicity elimination of the waste for animal feed
Utilization and reclamation of industry created wastelands
Expected results
The mycorrhizal technology offers biological means of assuring plant health in an
economically profitable and ecologically friendly manner. The only known fungal system
categorized as a biofertilizer, mycorrhizae provide plant roots with extended arms that
help them tap soil nutrients that are otherwise beyond their reach. This means greater
availability of other, enriching soil, increasing health, and decreasing dependence on
chemical fertilizers.

7. It exists a wide diversity within the AM fungi which form association
with the roots of almost 80% plant species and enhance mineral
nutrient acquisition and water uptake, as well as increase tolerance
towards different environmental stresses conditions.
To conserve and exploit their diversity, BioVale Project contemplates
to build J. curcas and mycorrhizal fungi culture depository houses
and maintain cultures from different agro-ecological zones, in
different areas of Brazil.
The expected result is to trap isolate a variety of J. curcas and
polysporal/monosporal cultures, setting up a J. curcas and
mycorrhizal fungi germplasm collection of genetic diversity of
agriculturally and industrially species for oil production .
Documentation, preservation and characterization
of J. curcas and mycorrhizal fungi germplasm

8. Jatropha has been identified as one the best plant
alternative to offer clean fuel for achieving energy
security. Jatropha seeds inoculated with in vitro-
raised mycorrhiza exhibits early fruition and
flowers from the 7th month onwards as against a
year with conventional clonal plantations and two
years from seed raised plantations.
The mycorrhized Jatropha also exhibited 20%–30%
higher yields as compared to non-mycorrhizal
plantations.
The mycorrhized Jatropha will be widely tested
covering several agro-climatic regions in Brazil to
prove their adaptation in diverse soils.
More and earlier productivity

9. The ROC (root organ culture) system is the most
attractive and advanced cultivation methodology for
AM (arbuscular mycorrhizal ) fungi: it uses root-
inducing transfer-DNA-transformed roots of a host
plant to develop the symbiosis on a specific medium
in vitro which provides pure, viable, contamination-
free inoculum using reduced space.
BioVale R&D Project aims at gaining expertise in
ROC for in vitro culture and utilize mass production
of AM fungi under in vitro.
Development of ROC of different AM fungi and their
mass inoculum production under in vitro

10. Although the facts and figures of potential role of mycorrhizal
association in enhanced nutritional and water needs of plants in
laboratories, the major bottleneck for its widespread application
to reach the end-users is its bulk production to cater the huge
requirement.
A known fact that culturing mycorrhizal fungi in laboratory
conditions like other microbes was not possible due to its strict
biotrophic nature of proliferation in the presence of suitable
host has been the major reservation of its future contribution in
agriculture.
The mass inoculum technology envisages to exploit the
genetically modified host roots using the Agrobacteriumum
rhizogenes carrying Ri T-DNA plasmid. The technology offers the
mass production of viable, healthy, genetically pure and high
quality fungal propagules, without any pathogenic contamination
under in vitro environment.
Mass inoculum technology

11. One of the main challenges that mankind will be facing during this century will
be the necessity to drastically increase animal production levels and
productivity in order to fulfill the feeding demands of a growing world
population.
J. curcas is a multipurpose tree of significant economic importance because
of its several industrial and medical uses. Jatropha grows throughout most of
the tropics. It survives on poor stony soils and is resistant to drought,
reaches a height from 3 m to 8 m and can be used to reclaim land. The seed
weighs approximately from 0.53 -0.86 g and its kernel contains 22 -27 %
protein and 57 -63 % lipid indicating good nutritional value.
The oil from these seeds can serve as fuel for diese! engines, indicating its
potential as a renewable energy source . The potential impact is immense for
countries with no indigenous fossil fuel or for regions remote from a source of
supply. Furthermore, use of biodiescl is friendly to the environment. The
seeds can be transported without deterioration and at low cost due to its high
specific mass.
However, the seed and oil have been found to be toxic to. Hence, its use as a
food or feed source is presently limited.

12. Biological detoxification is another environmental sound approach
for making J. curcas suitable for animal feeding. White-rot fungal
basidiomycetes are well known for their ability to degrade a wide
range of xenobiotics, such as polycyclic aromatic hydrocarbons,
polychlorinated biphenyls and synthetic dyes due to their enzymatic
systems [7, 8].
Enzymatic complexes produced by white-rot fungi have also an
enormous potential to be used in the treatment of fibrous
feedstuffs improving its nutritive value through the sequential
degradation of certain refractory cell wall components
Incresing aggregated value of the residue J. curcas
The oil expell

13. Treatment of J. curcas seed cake by solid-state incubation with white-rot fungi is
able to promote the decrease of phorbol esters content to non-toxic levels,
showing that this might be an alternative treatment with future potential. In fact,
most of the physical and chemical treatments being proposed so far have not
been able to definitively decrease the levels of these toxic compounds. The
development of a biological approach to treat J. curcas seed cake is an
environmentally sound and a more economical process, which might be
advantageous since it can effectively decrease the level of toxic compounds.
End result

14. de germoplasma pinhão
manso:
-Banco de germoplasma:ex
vitro (no campo) e in vitro
- Caracterização molecular
Montou-se o banco de germoplasma de pinhão manso no campus de
Florestal e em Viçosa, num total de 49 acessos.
Foi ajustado protocolo para desinfestação, estabelecimento e proliferação
de pinhão manso in vitro. Este protocolo permitiu montar banco de
germoplasma in vitro.
Uso dos reguladores BAP e AIA e dos antioxidantes ácido cítrico e ácido
ascórbico no meio de crescimento é eficaz na indução de brotos e
multiplicação permitindo formar banco de germoplasma in vitro do
pinhão manso
Caracterização molecular de 46 acessos, somente três foram divergentes.
Por possuírem alto valor em distância genética, podem ser utilizados
como genitores de um híbrido no programa de melhoramento

15. O substrato PlantMax isolado ou misturado com areia constitui-se um
substrato adequado para o desenvolvimento vegetativo das sementes e
estacas;
O banco de germoplasma do pinhão-manso foi desenvolvido com sucesso
tanto em Viçosa –MG quanto em Florestal-MG;
Pinhão-manso apresenta potencial para ser utilizado na micropropagação
tanto de sementes quanto de estacas.
Produção de mudas de
pinhão manso

16. Diversidade de Fungos
micorrízicos
arbusculares (FMAs) em
plantios de pinhão manso
Foram identificadas 19 espécies de FMAs na rizosfera de plantas de
pinhão manso.
O gênero Glomus foi o mais abundante em número de espécies entre os
FMAs na rizosfera de pinhão manso nas áreas estudadas.
Plantas de pinhão-manso ocorrendo em regiões distantes, porém, em
condições edafoclimáticas semelhantes, possuem populações de FMAs
semelhantes próximas às raízes.
A utilização em conjunto das metodologias morfológica e molecular
fornece informações mais completas a respeito da diversidade de FMAs
presentes na rizosfera das plantas em campo.
Plantas de pinhão manso apresentam elevada percentagem de colonização
micorrízica e relativa abundância no número de esporos na rizosfera.

17. Pleurotus ostreatus tem alta capacidade de degrada éster de forbol, ácido
fítico e compostos ligninocelulósicos;
P. ostreatus apresenta alta produtividade e eficiência biológica de
produção de cogumelos em substratos à base da torta de pinhão manso;
Os cogumelos produzidos na torta de pinhão manso apresentam teor de
éster de forbol em níveis considerados não tóxicos, podendo ser indicado
para o consumo, mas testes biológicos devem ser efetuados para
comprovar a inexistência dos compostos tóxicos ou anti nutricionais.

18. Características
nutricionais de resíduo
de pinhão manso tratado
com fungo da podridão
branca
O fungo da podridão branca aumentou os teores de nutrientes na torta de
pinhão manso. Apesar da torta de pinhão manso ser tóxica, a partir
dessas
características nutricionais a mesma serve para serem adicionados como
fonte de proteína, carboidrato e energia.
Enzymatic complexes produced by white-rot fungi have also an enormous
potential to be used in the treatment of fibrous feedstuffs improving its
nutritive value through the sequential degradation of certain refractory cell
wall components

19. Avaliação do
desempenho dos animais
alimentados com ração à
base de torta de pinhão
manso tratada -
digestibilidade in vivo
A torta destoxificada de pinhão manso pode ser utilizada com segurança
em até 20% da MS na dieta de caprinos em crescimento como alimento
Alternativo
In vitro digestible organic matter (DOM), metabolizable energy (ME) and rumen
degradable nitrogen