Synthesis of new plant growth regulator: N-(Fatty acid) O-aryloxyacetyl ethanolamine

Article abstract of DOI:10.1016/j.bmcl.2007.03.013

N-(Fatty acyl) O-aryloxyacetyl ethanolamines, prepared from N-acylethanolamine (NAE) and aryloxyacetic acid, were tested for plant growth regulating activity. Compared with N-stearoylethanolamine, most compounds exhibit improved plant growth stimulating activity. In particular, those with chlorine on aryl ring show better activity than 2,4-dichlorophenyloxyacetic acid in stimulating hypocotyls elongation of rape which indicates that chlorine on aryl ring appears significant. Moreover, these derivatives display improved solubility.

Full text of DOI:10.1016/j.bmcl.2007.03.013

Bioorganic & Medicinal Chemistry Letters 17 (2007) 3231–3234
Synthesis of new plant growth regulator: N-(Fatty acid)
O-aryloxyacetyl ethanolamine
Liang Han,a Jian-Rong Gao,a Zheng-Ming Li,b,* Yun Zhangc and Wei-Ming Guoc
aCollege of Chemical Engineering and Material Science, Zhejiang University of Technology, Hangzhou 310014, China
bNational Key Laboratory of Elemento-Organic Chemistry, Nankai University, Tianjin 300071, China
cCollege of Horticulture, Nanjing Agricultural University, Nanjing 210095, China
Received 26 October 2006; revised 13 February 2007; accepted 5 March 2007
Available online 12 March 2007
Abstract—N-(Fatty acyl) O-aryloxyacetyl ethanolamines, prepared from N-acylethanolamine (NAE) and aryloxyacetic acid, were
tested for plant growth regulating activity. Compared with N-stearoylethanolamine, most compounds exhibit improved plant
growth stimulating activity. In particular, those with chlorine on aryl ring show better activity than 2,4-dichlorophenyloxyacetic acid
in stimulating hypocotyls elongation of rape which indicates that chlorine on aryl ring appears significant. Moreover, these deriv-
atives display improved solubility.
Ó 2007 Elsevier Ltd. All rights reserved.
N-Acylethanolamines (NAEs), a family of endogenous
fatty acid amides, are known in mammals since some
decades. They are proved to be an important bioactive
substance in microgram level existing in the animal cells
and play biochemical and pharmacological role.1 How-
ever, their widespread occurrence, metabolism, and
physiological significance in plants have only recently
begun to be appreciated.2 More investigation supported
the fact that NAEs play a lipid mediator role in plants
and are implicated as transducers in plant defense sig-
naling,2d,f,3 and at elevated levels, interfered with normal
seedling root development.4 The identification and ac-
tive metabolism of NAEs in these physiological situa-
tions supports the emerging concept that NAEs are
endogenous signaling compounds in plant systems.5
nous NAEs further confirmed the recognition of their
important role in plant.2f,9
Though the above reports are exciting, NAEs have very
low solubility in water and limited solubility in most
common organic solvents, which discourages further
study and application. It urges the molecular modifica-
tion of NAEs to improve the solubility, which has not
been reported to date. On the other hand, it is well
known that substituted aryloxyacetic acids exhibit high
plant growth regulating activity.10 Yet, sometimes they
induce unfavorable responses. For example, when
substituted aryloxyacetic acids were used to improve
fruit set, some suppression of leaf growth happened.10
The report that the ester of aryloxyacetic acids can not
only improve the translocatibility but also reduce the
extent of side effects encourages us to incorporate the
aryloxyacetic acid type regulator into NAEs. A series of
N-(fatty acyl) O-aryloxyacetyl ethanolamines were syn-
thesized and their bioactivity is reported here. The prep-
aration of title compounds followed the reaction
sequence depicted in Scheme 1.
Furthermore,
LPE(lysophosphatidylethanolamine),
capable of inhibiting plant PLD a (phospholipase a),
has a profound effect on the physiological symptoms
associated with postharvest senescence of flowers and
fruits6; however, NAE12:0 and NAE14:0 are at least a
100-fold more potent than LPE in inhibiting PLD a
activity, so these NAEs may found novel agrochemical
applications in the future.7,8 The applications of exoge-
Several NAE types have been identified in a variety of
plant species. These NAE types identified contain acyl
chains of 12-18C in length and up to three double
bonds, reflecting the typical acyl moieties prevalent in
higher plants.2a In terms of bioactivity, plants are partic-
ularly responsive to low concentration of medium chain
Keywords: N-Acylethanolamines; Aryloxyacetic acids; Plant growth
regulator; Synthesis.
*
Corresponding author. Tel.: +86 0 22 23503732; fax: +86 0 22
23505948; e-mail: nkzml@vip.163.com
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.03.013

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L. Han et al. / Bioorg. Med. Chem. Lett. 17 (2007) 3231–3234
+
HOCH2CH2NHCO(CH2)nCH3
indicated the reaction happened and the products had
better solubility in CHCl3 than NAEs. After the usual
workup, title compounds 1 were obtained through the
purification by column chromatography. Structures of
title compounds were confirmed by elemental analyses
NH2CH2CH2OH
CH3(CH2)nCOOH
NAE
R
R
+
OCH2COOH
SOCl2
R
OCH2COCl
1
and H NMR.13
NAE
O CH2COO NHCO(CH2)nCH3
Et3N/CHCl3
On the other hand, the solubility of title compounds and
NAEs was detected in CHCl3, EtOAc, and EtOH. The
solubility data of some title compounds with NAE12:0
and NAE18:0 are listed in Table 1. As compared with
respective NAE, title compounds show evident solubil-
ity improvement.
1
Scheme 1.
saturated NAEs; whereas animal physiology is largely
regulated by low concentration of long chain polyunsat-
urated NAEs (e.g., NAE 20:4).5 In our work, four kinds
of NAEs, namely NAE12:0, NAE14:0, NAE16:0,
NAE18:0, were synthesized to incorporate into the
structure of several aryloxyacetic acids. NAE18:0 was
chosen to couple with five kinds of aryloxyacetic acids
to study the influence of substituent group on bioactiv-
ity. And the 2,4-dichlorophenoxyacetic acid (2,4-D)
was used to react with all synthesized NAEs to investi-
gate the bioactivity variety along with chain length.
Since NAE was reported to prolong the shelf life of cut
flowers, delay deterioration and leaf drop, and extend
the overall appearance and quality of the plant cutting,9
title compounds were tested for their plant growth stim-
ulating activity in hypocotyls elongation of rape, cotyle-
don expansion of cucumber, and coleoptiles growth of
common wheat.14–16 The increase percent in these assays
over control samples with no regulator is listed in Table
2 and also are the data of NAE18:0 and the conven-
tional plant growth regulator 2,4-D for comparison.
Just like NAE18:0 and 2,4-D, title compounds exhibit
good stimulating effect on hypocotyls elongation of
rape. In hypocotyls elongation test, improved stimulat-
ing effect was observed for all title compounds except
compound 1a when compared with NAE18:0, and com-
pounds 1e–1j with different chain length and substituent
group even have better growth stimulating activity than
2,4-D. It is interesting to note that the hypocotyls elon-
gation of rape appears strongly associated with the sub-
stituent group on the benzene ring. The compounds with
substituent chlorine are obviously superior to those with
substituent methyl and methoxy group, which suggests
that chlorine on aryl ring appears significant. For exam-
ple, compound 1g has an increase percent of 73.8% in
stimulating hypocotyls elongation of rape, while the in-
crease percent of compound 1b is only 57.0%. However,
introduction of additional chlorine does not affect the
elongation of rape hypocotyls and the activity of
4-chloro-substituent 1e is comparable to that of 2,4-
dichloro-substituent 1g. In coleoptiles growth test of
common wheat, all title compounds also exhibit better
stimulating effect than NAE18:0 except compound 1b,
NAEs were prepared by refluxing 1 mol of the fatty acid
with 1.5 mol of ethanolamine for 6 h in 60–90% yield.11
The products were purified by recrystallization from the
solution of trichloromethane and acetone instead of 95%
ethanol of Ref. 11. Aryloxyacetic acids were prepared
according to Ref. 12 and then transformed to the acid
chlorides by refluxing with SOCl2 for 5 h. NAEs, sus-
pended in CHCl3 in the presence of Et3N, were dissolved
gradually with the addition of the acid chlorides, which
Table 1. Solubility of some title compounds and parent NAEs
Compound
n
R
Solubility (g)
EtOAc
CHCl3
EtOH
NAE12:0
1j
NAE18:0
10
10
16
16
16
16
16
—
2,4-2Cl
—
1.76
15.10
0.16
0.38
11.0
0.03
2.67
8.27
1.54
3.54
9.14
10.42
0.34
1.99
3.08
1.88
1.03
1a
1b
1e
1g
m-CH3O
o-CH3
p-Cl
7.76
53.97
4.37
10.22
2,4-2Cl
Table 2. Plant growth regulating bioactivity in vitro
Compound
n
R
Increased bioactivity (%, 10 mg/L)
Hypocotyls elongation of rape Cotyledon expansion of cucumber Coleoptiles growth of common wheat
1a
1b
16 m-CH3O À1.4
À0.1
5.7
5.0
0.7
16 o-CH3
16 p-CH3
10 p-CH3
16 p-Cl
57.0
38.7
54.9
71.8
71.8
73.8
67.3
68.1
69.0
35.3
65.7
1c
1d
9.8
5.9
3.3
4.1
1e
6.3
2.9
5.0
1f
1g
10 p-Cl
16.0
7.5
16 2,4-2Cl
14 2,4-2Cl
12 2,4-2Cl
10 2,4-2Cl
0.7
3.5
1h
1i
17.0
8.9
À0.1
4.0
1j
17.9
2.1
NAE(18)
2,4-D
16
—
—
2,4-2Cl
6.2
0.8
28.3

L. Han et al. / Bioorg. Med. Chem. Lett. 17 (2007) 3231–3234
3233
Giuffrida, A.; Rodriguez de Fonseca, F.; Piomelli, D.
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unfortunately, only compound 1c shows a little im-
proved stimulating activity in cotyledon expansion test
of cucumber. The change of fatty chain length seems
to have intricate effect on the bioactivity. Compound
1f has the same bioactivity in stimulating hypocotyls
elongation of rape as 1e, though there is a difference
of six methylenes in their fatty chain structure. But when
stimulating cotyledon expansion of cucumber and cole-
optile growth of common wheat, compounds with fatty
chain of 16 carbons and 12 carbons have better activity
than compounds with fatty chain of 18 carbons and 14
carbons.
8. Sang, Y.; Zheng, S.; Li, W.; Huang, B.; Wang, X. Plant
J. 2001, 28, 1.
9. Chapman, K. D. PCT Int. Appl. WO01,301,43 A2,
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S.; Drake, T. F. J. Agri. Food Chem. 1959, 7, 118.
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In conclusion, a series of N-(fatty acyl) O-aryloxyacetyl
ethanolamines were synthesized by reaction of NAEs of
different chain length with substituted aryloxyacetic
acids. All new compounds have improved solubility.
The preliminary bioactivity data show that most of them
exhibit better plant growth stimulating effect than
NAE18:0, moreover, those with chlorine on benzene
ring have better stimulation of hypocotyls elongation
than conventional 2,4-D.
13. General method for preparing title compounds. A mixture of
NAE (1 mmol) and Et3N (1.2 mmol) in dry CHCl3 (5 ml)
was stirred at room temperature and then the solution of
aryloxyacetic acid chloride (1.2 mmol) in dry CHCl3 (2 ml)
was added drop by drop. During the addition, NAE was
dissolved gradually and the solution turned yellow. After
stirred for 12 h, the solution was washed with water and
then dried with MgSO4. The solvent was removed and
column chromatography of the residue with PE/EtOAc
(1.5:1, v/v) gave the product as a white solid. Compound
1b, yield 75%: mp 63–64 °C; 1H NMR (CDCl3, 300 MHz):
d 7.20–7.12 (dd, 2H, ArH), 6.94–6.90 (t, 1H, ArH), 6.72–
6.69 (d, 1H, ArH), 5.45 (br s, 1H, NHCO), 4.70 (s, 2H,
ArOCH2), 4.28–4.24 (t, 2H, OCH2CH2N), 3.51–3.46 (dd,
2H, OCH2CH2N), 2.30 (s, 3H, ArCH3), 2.08–2.02 (t, 2H,
CH2CONH), 1.58–1.53 (t, 2H, CH2CH2CO), 1.25 (br s,
28H, (CH2)14), 0.90–0.86 (t, 3H, CH3(CH2)14) ppm; Anal.
calcd for C29H49NO4: C 73.22, H 10.38, N 2.94; found: C
73.24, H 10.40, N 2.85.
Acknowledgment
We are grateful for the financial support from National
Natural Science Foundation NNSFC#20432010.
Supplementary data
Supplementary data associated with this article can be
found, in the online version, at doi:10.1016/
j.bmcl.2007.03.013.
References and notes
14. Hypocotyls elongation test of rape. After soaking, the seeds
with similar magnitude were chosen to use. Tested
compounds were dissolved in DMF and later dropped
evenly on 6-cm-diameter fitter paper. After air-volatiliza-
tion of solvent, the filter paper was placed in 6-cm-
diameter glass utensil with distilled water to give 10 mg/L
compound solutions, and then ten seeds were added. The
seeds were cultured at 25 °C in dark. The length of
hypocotyls was measured after 3 days and compared with
those treated with distilled water to estimate the activity.
Two replicates were included in the evaluation.
15. Cotyledon expansion test of cucumber. After soaking,
the seeds were germinated in covered enamelware
containing 0.7% agar and cultured for 3 days in dark
at 26 °C, and then the cotyledons of similar magnitude
were chosen to use. Tested compounds were dissolved
in DMF and later dropped evenly on 6-cm-diameter
fitter paper. After air-volatilization of solvent, the filter
paper was placed in 6-cm-diameter glass utensil with
distilled water to give 10 mg/L compound solutions and
ten pieces of cotyledons were added. The cotyledons
were cultured in light (300Lux, 26 °C). After 3 days,
the total weight of cotyledon was measured and
compared with those treated with distilled water to
estimate the activity. Two replicates were included in
the evaluation.
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L. Han et al. / Bioorg. Med. Chem. Lett. 17 (2007) 3231–3234
16. Coleoptiles growth test of common wheat. After soaking,
the seeds were germinated in covered enamelware con-
taining 0.7% agar and cultured for 3 days in dark at 25 °C.
When the seedling grew to 2.5–3.0 cm tall, the first 3 mm
of coleoptile top was rejected. Coleoptile (5 mm) was
truncated and dunked in distilled water for 1 h to remove
endogenous hormone, then it was chopped into 10
segments. Tested compounds were dissolved in DMF
and later dropped evenly on 6-cm-diameter fitter paper.
After air-volatilization of solvent, the filter paper was
placed in 10 ml beaker with 0.01 mol/L phosphoric acid-
citric acid buffer solution (pH 5) containing 2% sucrose to
give 10 mg/L compound solutions and ten coleoptile
segments were added afterward. The coleoptiles were
cultured at 25 °C in dark for 18–20 h and then the length
of coleoptiles was measured and compared with those
treated without tested compounds to estimate the activity.
Two replicates were included in the evaluation.