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to the subfamily 2 are neutral or slightly
negative (Craik
et al
., 1999).
To date, cyclotides have been found in
the genera
Leonia
and
Viola
, belonging to
Violaceae. In this work we have identified
for the first time a novel cyclotide in the
genus
Hybanthus
, which belongs to the
same plant family. The presence of cyclo-
tides in
Hybanthus
could therefore be
regarded as a chemotaxonomic marker for
this genus. More than 100 cyclotides have
been identified so far, and new structures
continue to be isolated and described.
Because the number of compounds in this
family is increasing, we have proposed that
the trivial name for each compound be
formed with the acronym of the Latin name
or the scientific name of the plant from
which they are isolated for the first time,
and followed by a letter indicative of the
order of appearance. Thus, the first cyclo-
tide isolated from
H. parviflorus
has been
named hypa A (Broussalis
et al
., 2001). It is
also worth noting that hypa A is the first
cyclotide isolated from an Argentine vegetal
species.
detemined in the 50% EtOH extract by TLC
(Broussalis
et al.
, 2010).
The CH
2
Cl
2
and 50% EtOH extracts and
the fraction eluted with 25% ACN-0.1%
TFA of the tannin-free butanolic fraction of
the 50% EtOH extract were analysed by
HPLC employing a C18 column. The 50%
EtOH extract was subjected to HPLC employ-
ing a Phenomenex
TM
C18 analytical column
employing apigenin, luteolin, kaempferol,
quercitin, quercitin-methyl-ether and rutin
as reference flavonoids. The mobile phase
consisted of the following solvents: A: H
2
O-
AcOH (98:2) and B: MeOH-AcOH (98:2).
The following flavonoids were identified by
comparing their rt and UV spectra with the
rt and UV of the corresponding commercial
standards: quercetin (rt 26.59 min), luteolin
(rt 28.04 min), quercetin-3-methyl-ether (rt
28.47 min), kaempferol (rt 30.48 min) and
apigenin (rt 31.32 min) and the glycoside
rutin (rt 18.62 min). Quercetin-3-methyl-
ether and rutin were identified for the first
time in
H. parviflorus.
Because the amount of rutin present
in the 50% EtOH extract was considerably
higher than the other flavonoids, its pres-
ence was quantified. The rutin content, as
determined by HPLC, was 1.34 g/100 g
extract (1.34%) or 0.24% when referring to
dry powder (Broussalis
et al
., 2010). In
this extract the insecticide activity was
tested. The pharmacological activities of
the flavonoids apigenin, luteolin, kaemp-
ferol, quercitin and quercitin-3-methyl-
ether and the rutin glycoside have been
previously reported; however, such com-
pounds continue to be studied in the
search for novel biological activities and
therapeutic applications (Havsteen, 2002;
Menendez
et al
., 2008).
The presence of caffeic and chlorogenic
acids was assessed in the ethanolic extract
of
H. parviflorus
by HPLC, employing refer-
ence substances. The butanolic fraction of
the 50% EtOH extract eluted with 25%
CAN-0.1% TFA that had been kept for the
analysis of flavonoids was subjected to
semi-preparative RP-HPLC employing 25%
ACN-0.1% TFA as mobile phase. Upon ana-
lysing the UV spectra of this fraction, the pre-
sence of flavonoids could be demonstrated.
13.3.4
Phenolic compounds
Flavonoids and caffeoylquinic acids have
been reported to have diverse biological
activities that can be related to the insecti-
cide activity of
H. parviflorus
. For this
reason, the polyphenolic compounds (fla-
vonoids and caffeoylquinic acids) present
in the CH
2
Cl
2
and 50% EtOH extracts of
H. parviflorus
have been studied by thin-
layer chromatography (TLC) and HPLC. The
TLC analysis allowed the determination of
the presence of rutin - the principal com-
pound - and chlorogenic acid in the 50%
EtOH extract by means of the three chroma-
tographic systems employed. The analysis
of the UV spectra of the compounds present
in CH
2
Cl
2
, 50 % EtOH and its butanolic
fraction, subjected to HPLC-DAD, revealed
the presence of other flavonoids and caffe-
oylquinic acids and allowed quantification
of the principal flavonoid rutin. The pres-
ence of rutin and chlorogenic acid were
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