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Digital archive of theses discussed at the University of Pisa


Thesis etd-03022014-182211

Thesis type
Tesi di dottorato di ricerca
Thesis title
Biotechnological approach in Lamiaceae species for the production of antioxidant and antibacterial compounds
Academic discipline
Course of study
tutor Dott. Pistelli, Laura
commissario Ruffoni, Barbara
commissario Natali, Lucia
  • carnosic acid
  • HPPR
  • in vitro cultures
  • rosmarinic acid
  • Rosmarinus officinalis
  • Salvia officinalis
Graduation session start date
The interest in natural-derived molecules rosmarinic acid (RA) and carnosic
acid (CA) has raised up during the last years due to their well documented
antioxidant properties: they are in fact used in foodstuff as preserving agent
as well as in pharmacology due to their antibacterial and medicinal properties.
In vitro cultures of medicinal plants are considered an alternative way to
conventional methods for the production of valuable secondary metabolites;
undifferentiated cells (callus and cell suspensions) are also suitable for both
plant biomass production and scaling-up and for the study of metabolites
In this work Salvia officinalis L. (common sage) cell suspension cultures were
established, and a specific cell line was selected for the high antioxidant
capacity of its methanolic extract, which was characterized by a high content
of RA. Scavenger activity (DPPH test) and total RA content were evaluated
during cell growth. Gene coding for hydroxyphenylpyruvate reductase, the key-
enzyme responsible for the RA metabolic biosynthesis, was cloned from common
sage (SoHPPR). Its transcript expression level was monitored during cell
suspension cultures, and showed a relationship with scavenger activity and RA
yield. Our results suggest the potential use of this gene as a marker and
target for the modulation of RA production in controlled conditions. In
addition, the optimum conditions for Agrobacterium-mediated transformation were
determined by monitoring the transient expression of β–Glucuronidase (GUS)
marker gene in callus culture.
We also set up a protocol for the in vitro culture of Rosmarinus officinalis,
one the few species able to synthesize the phenolic diterpene CA. The target of
this study was to evaluate the capacity of different tissues of synthesize CA.
Our results suggest that all the tissues can accumulate varying amounts of CA,
depending on the differentiation grade of the cell culture type: among the
different culture types, neo-formed shoots coming from irradiated calli
accumulated the highest content of CA. Finally we evaluated whether CA extract
may be used as a natural substitute of antibiotics commonly used in plant
tissue cultures: our data suggest a bacteriostatic activity of CA against
common pathogens without affecting plant growth and physiology.