Ehime University Institutional Repository

中曽, 久雄

Japanese

猿払基準の行方

池田, 彩乃

井坂, 万由

大西, 礼夏

淡野, 寧彦

岡山県瀬戸内海沿岸地域におけるい草関連産業の存続形態

MAGEE, Glenn Amon

English

Think-Pair-Share-Compare : An organizational principle for deeper learning

梶原, 克彦

オーストリア=ハンガリーにおける敵国民間人の抑留・拘禁と解放 : 日本人抑留者の事例を中心に 二

信原, 孝司

対象喪失研究と心理療法 : 子どもとの死別に関する一考察

杉田, 浩崇

教育学における脳科学受容の社会的次元 : 批判的脳科学の射程を探ることを通して

EGAMI, Sonoko

ITO, Miyuki

Changes in Sibling Conflict and Prosocial Behavior : During the "Terrible Twos" Period

Ehime University (愛媛大学)博士(農学)thesi

Md., Tazul Islam Chowdhury

（様式５）(Style5) 
学位論文全文に代わる要約 
Extended Summary in Lieu of Dissertation 
 
 
氏名： 
Name 
 
Md. Tazul Islam Chowdhury 
学位論文題目： 
Title of Dissertation 
 
Synthesis of Rare Sugar Derivatives and Their Biological Activity on Plant 
Growth 
 
（希少糖誘導体の合成と植物生長に対する生物活性） 
 
学位論文要約： 
Dissertation Summary 
 
Agriculture in the 21st century faces multiple challenges; many of today’s food production systems are 
unsustainable. In appropriate use of synthetic chemicals causes hazardous problem to flora and fauna 
(Pell et al. 1998; Aktar et al. 2009; Dreistadt et al.1994). To avoid the detrimental effects of synthetic 
chemicals, research on novel natural products have moved from the fringe to the mainstream for the 
development of ecologically acceptable, environment friendly and relatively safe natural plant growth 
retardants. Synthetic growth retardants are becoming very popular in the farmers field but due to 
toxicological concerns, it has declined markedly in recent years, particularly for edible crops 
(Rademacher 2000). Therefore, to make environmentally friendly growth retardants, scientists are 
giving more concentration on natural products and their derivatives. In this regards, rare sugar 
derivative could replace as natural growth retardants, which is biodegradable and safe for the 
environment.  
Rare sugars are monosaccharides that exist in nature in a limited amount. Izumori et al. developed a 
great idea for the mass production of rare sugar using recombinant enzymes from bacteria (Izumori 
2002, 2006). Despite their limited presence in nature, rare sugars have various biological functions and 
vast applications in pharmaceutical, cosmetics, food, and flavor industries. The biological activities of 
rare sugars such as D-psicose (Takeshita et al. 2000), D-tagatose (Levin et al.1995) and D-allose 
(Bhuiyan et al. 1998; Sui et al. 2005; Gao et al. 2011; Hossain et al. 2000) has been extensively studied.   
Based on enzymatic synthesis, my research group already reported the synthesis of 6-O-acyl-D-allose 
from rare sugar, D-allose, via regioselective lipase catalyzed transesterification and its biological activity 
against plant growth (Afach et al. 2005, 2006). D-Allose fatty acid esters showed 2-20 times greater 
inhibitory activity than D-allose itself because the hydrophobic carbon chains of the D-allose derivatives 
（様式５）(Style5) 
could enhance the surface activity and membrane permeability. Furthermore, they described that 
α-hydroxy group at C-3 might play an important role for the inhibitory activity because the 
6-O-dodecanoate of D-mannose (C-2 epimer of D-glucose) and D-galactose (C-4 epimer of D-glucose) 
showed weaker effect on rice growth than that of D-allose (Kobayashi et al. 2010). Nowadays rare sugar 
research is important for further evaluation and application of rare sugar research. Therefore, 
considering the biodegradable plant growth retardants, the intention of the first study was to investigate 
the synthetic procedure of 6-O-decanoyl-D-altrose from D-altrose and 6-O-decanoyl-D-gulose from 
D-gulose. 
The reactions of D-gulose and D-altrose with vinyl decanoate using Candida antarctica lipase in 
different organic solvents were carried out (Scheme 1 and 2) to obtain new monosacharride derivatives, 
Gul-C10 and Alt-C10 respectively. Then, the biological activity of Gul-C10 and Alt-C10 was evaluated 
against different plant species. Both of them showed similar inhibitory activity against cress, Italian 
ryegrass and rice seedlings but exhibited weaker activity against of lettuce. 
   
O
OH
OH
OH
OH
OH
O
O
Novozym 435, acetonitrile, 45 
o
C, 48 h
O
OH
O
OH
OH
OH
O
 
     D-gulose                                      6-O-decanoyl-D-gulose 
              Scheme 1 Synthesis of 6-O-decanoyl-D-gulose 
    
O
OH
OH
OH
O
O
Novozym 435, acetone, rt, 48 h
O
OH
O
OH
O
OH HO
OHOH
 
     D-altrose                                     6-O-decanoyl-D-altrose      
               Scheme 2 Synthesis of 6-O-decanoyl-D-altrose 
 
Bioassay results showed that lettuce seedlings were not significantly inhibited by the D-altrose and 
D-gulose (Fig. 1). On the other hand, D-altrose C10 and D-gulose C10 significantly inhibited lettuce 
seedlings (Fig. 1) in a higher concentration (3 mM). However, these rare sugar derivatives showed 
（様式５）(Style5) 
weaker inhibitory activity on the growth of lettuce shoot and root.   
 
Fig. 1 Biological activity of rare sugars and their derivatives on a) shoot and b) root of lettuce seedlings. 
Means ± SEM from 3 independent experiments with 10 plants for each determination are shown 
 
Next, to examine the structure-activity relationship (SAR) of the hydroxy groups at C-1 and C-2 of the 
D-allose esters, three different analogs of deoxy-D-allose ester were synthesized: 
6-O-decanoyl-2-deoxy-D-allose (2-DOAll-C10), lacking a hydroxy group at C-2; 
6-O-decanoyl-1,2-dideoxy-D-allose (1,2-DOAll-C10), which is saturated at C-1 and C-2; and 
6-O-decanoyl-1,2-didehydro-1,2-dideoxy-D-allose (1,2-DHAll-C10), having a double bond between 
（様式５）(Style5) 
C-1 and C-2. Then, their biological activities on plant growth were evaluated.  
2-DOAll-C10 (1) was synthesized as shown in Scheme 3. First, triacetyl 1,2-DHAll (4)  was added with 
AcOH (Morris and Shair 2009) to give 2-deoxy-D-allose tetraacetate (5) in a 67% yield because hydration 
with dilute H2SO4 afforded 5 in low yield (Haga and Tejima 1974).
 Then, 5 was hydrolyzed to 
2-deoxy-D-allose (6) in a 69% yield. Finally, regioselective lipase-catalyzed transesterification with 
vinyl decanoate gave 2-DOAll-C10 (1) in a 54% yield.  
6-O-Decanoyl-1,2-dideoxy-D-allose (2) and 6-O-decanoyl-1,2-didehydro-1,2-dideoxy-D-allose (3) were 
synthesized via regioselective lipase-catalyzed transesterification of 1,2-dideoxyallose (7) and 
1,2-didehydro-dideoxyallose (8) with vinyl decanoate to give 1,2-DOAll-C10 (2) and 1,2-DHAll-C10 in a 
58%  and 31% yield, respectively (Scheme 4). 
O
AcO
AcO
OAc
AcOH, Ph3PHBr, CH2Cl2
OAc
rt, 10 min, 69%
K2CO3, MeOH O
OH
HO
OH
OH
30
o
C, 21 h, 67%
Vinyl decanoate
Lipase, rt, 48 h, 54%
O
OH
HO
O
OH
(CH2)8CH3
O
O
OAc
AcO
OAc
4 5 6
1                    
               Scheme 3 Synthesis of 6-O-decanoyl-2-deoxy-D-allose (1)  
 
           
O
OH
HO
OH
Vinyl decanoate
Lipase, rt, 48 h, 58%
O
OH
HO
O (CH2)8CH3
O
O
OH
HO
OH
Vinyl decanoate
Lipase, rt, 48 h, 31%
O
OH
HO
O (CH2)8CH3
O
2
3
7
8
 
 
Scheme 4 Synthesis of 6-O-decanoyl-1,2-dideoxy-D-allose (2) and 
6-O-decanoyl-1,2-didehydro-1,2-dideoxy-D-allose 
 
（様式５）(Style5) 
The biological activities of the deoxy alloses and their fatty acid esters were examined using lettuce, 
cress, Italian ryegrass, and rice seedlings. The fatty acid esters of the deoxy alloses (1–3) showed higher 
growth inhibitory activity on different test plants than the deoxy alloses. 2-DOAll-C10 (1) inhibited the 
growth in a concentration˗dependent manner, which was a similar tendency to that of All-C10. Although 
1,2-DOAll (7) showed growth-promoting activity, its ester, 1,2-DOAll-C10 (2), showed weak 
inhibitory activity at 3 mM. 1,2-DHAll-C10 (3) exhibited increased inhibitory activity to some extent 
compared with 1,2-DHAll (8). These results suggest the effectiveness of esterification of the hydroxy 
group with a medium fatty acid chain which could enhance surface activity and membrane 
permeability (Fujii 1996; Ogino 1996). 
Table 1 IC50 values of deoxy-D-allose esters and All-C10 against four plant species. 
IC50 (mM)
Lettuce Cress Rice
Shoot    Root Shoot    RootShoot    Root Secondary leaf sheath
1
2
All-C10
2.70>3.0 >3.0 >3.0 >3.0 3.0 2.56
1.02 0.98 0.37 0.40 0.51 0.48 0.19
0.84 0.69 0.32 0.36 0.48 0.51 0.18
Compound
3 2.83 2.03 0.56 0.39 1.02 0.6 0.42
Italian ryegrass
 
The concentrations required for 50% inhibition (IC50) of lettuce, cress, Italian ryegrass, and rice 
seedling hypocotyls are listed in Table 1. 2-DOAll-C10 (1) showed inhibitory activity on four plant 
species similar to that of All-C10. 1,2-DHAll-C10 (3) showed inhibitory activity on cress, Italian ryegrass, 
and rice seedlings similar to that of All-C10, but lower inhibitory activity on lettuce. On the other hand, 
saturated 1,2-DOAll-C10 (2) exhibited considerably weaker inhibitory activity than the other deoxy 
allose esters against the four plant species. These results suggest that the hydroxy group at C-2 is not 
necessary for inhibitory activity and that the hydroxy group and/or hemiacetal structure at C-1 might play 
an important role in activity. 
Then, a new deoxy derivative of All-C10 amide was synthesized and examined its inhibitory activity 
against plant growth to compare with deoxy All-C10 ester. 6-(Decanoylamino)-1,2,6-trideoxy-D-allose 
was prepared from the known compound 1,2-dideoxy-D-allose via regioselective tosylation, azidation, 
catalytic hydrogenation, and acylation with decanoyl chloride (Scheme 5).  
（様式５）(Style5) 
O
HO
OH
OR
O
HO
OH
NR
HO
HO
OH
NR
HO
OH
HO
HO
OH
OR
HO
OH
 
9: R = CO(CH2)8CH3    10: R = CO(CH2)8CH3      11: R = CO(CH2)8CH3     12: R = CO(CH2)8CH3 
   Fig. 2 Structures of All-C10 (9), dAll-C10 (10), All-C10 amide (11), and dAll-C10 amide (12) 
 
O
HO
OH
O
HO
OH
CH3CN, TsCl, Py
-5
 o
C to rt, 18 h
DMF, NaN3
90
 o
C, 48 h
dry EtOH, Pd-C, H2
rt, 48 h
O
HO
OH
O
HO
OH
H
N
O
(CH2)8CH3
54 %
93 % 95 %
CH2Cl2, decanoyl chloride
Et3N, 6 h
74 %
OH
OTs
NH2
O
HO
OH
N3
1,2-DOAll
13
14
15
dAll-C10 amide
1
2
3 4
5
6
 
   Scheme  5  Synthesis of 6-(decanoylamino)-1,2,6-trideoxy-D-allose (12) 
 
The biological activity of 6-(decanoylamino)-1,2,6-trideoxy-D-allose was evaluated using four test 
species. dAll-C10 amide inhibited the growth of the lettuce seedlings in a concentration-dependent 
manner ranging from 0.03 to 1 mM, and interestingly, slight growth promotion (119%) was observed 
of the roots at 0.1 mM (Fig. 3a). The growth of the cress and Italian ryegrass seedlings was 
significantly inhibited by dAll-C10 amide in a concentration-dependent manner (Fig. 3b and 3c). The 
growth inhibitory activity of dAll-C10 amide was also evaluated using rice seedlings. The inhibitory 
activity increased in a concentration-dependent manner from 0.1 to 1 mM, and the growth of the rice 
seedlings was completely inhibited by dAll-C10 amide at a concentration greater than 1 mM (Fig. 3d). 
 
（様式５）(Style5) 
 
Fig. 3  Biological activity of dAll-C10 amide in a) lettuce, b) cress, c) Italian ryegrass, and d) rice 
seedlings. Values are mean ± SEM from three independent experiments 
 
Comparing with those of other D-allose derivatives, the activity of dAll-C10 amide was slightly weaker 
than that of All-C10 but significantly higher than that of dAll-C10. These results indicate that 
replacement of the ester group with an amide group has opposite effects on D-allose derivatives and 
deoxy-D-allose derivatives. The concentrations required for 50% inhibition (defined as IC50) of Alt-C10, 
Gul-C10, 1,2-DHAll-C10, 1,2-DOAll-C10, 2-DOAll-C10 and dAll-C10 amide (Fig. 4) against lettuce, 
cress, Itlian ryegrass, and rice seedling hypocotyls were displayed in Fig. 5.  All-C10, Alt-C10, 
Gul-C10, 1,2-DHAll-C10, 2-DOAll-C10 and dAll-C10 amide showed almost similar inhibitory 
activity on rice, cress and Italian ryegrass seedlings. But incase of lettuce, Alt-C10, Gul-C10 and 
1,2-DHAll-C10 exhibited lower inhibitory activity than  All-C10, 2-DOAll-C10 and dAll-C10 amide, 
while 1,2-DOAll-C10 displayed very weak activity against all the test species. Among the four 
different test plants, rice was most sensitive to show the inhibitory activity.  
（様式５）(Style5) 
 
  
Fig. 5 IC50 values of the rare sugar derivatives on a) shoot and b) root of different plant species 
 
These results suggested that the hydroxy group at C-1 and C-3 position is important to exhibit 
inhibitory activity on plant growth for rare sugar derivatives of 2-DOAll-C10 and All-C10. Moreover, 
the amide group of deoxy allose at C-6 is also important to increase the retarding activity on plant 
growth. 
To disclose the mechanism of growth-inhibitory activity of rare sugar derivatives on plants, the effect 
of active gibberellin with rare sugar derivatives on rice seedlings were examined because it has also 
been reported that some growth inhibitors retarded the rice shoot elongation and this growth was able 
to be recovered by the treating with GA3 (Rademacher et al. 1991; Graebe 1987). In these studies, the 
（様式５）(Style5) 
recoveries of rice growth were also observed by the co-addition of GA3. 
O
HO
OH
H
N
O
(CH2)8CH3
O
OH
HO
O (CH2)8CH3
O
OH
HO
O
OH
HO
O (CH2)8CH3
O
O
O (CH2)8CH3
O
OH
H
1
2
3
O
OH
HO
O (CH2)8CH3
O
1
2
3
O
OH
HO
O (CH2)8CH3
O
2
3
Alt-C10 Gul-C10 1,2-DOAll-C10
1,2-DHAll-C10 2-DOAll-C10
OH
1 12
3 4
5
6
1
2
3
4
12
3
HO
OH
dAll-C10 amide
                        Fig. 4 Rare sugar derivatives  
 
Moreover, daminozide was used as a positive control for growth recovery experiment. Rice growth was 
recovered in the similar tendency to rare sugar esters when GA3 was co-added with a solution of 
daminozide (Fig. 6).   
 
Fig. 6 Effect of dAll-C10 amide, daminozide (Dam), and GA3 on rice seedlings. Relative lengths (% of 
control) of shoot and second leaf sheath of rice at 7 days after treatment. Values are mean ± SEM (n = 
14), and bars with different letters indicate significant difference as determined by Tukey’s honestly 
significant difference comparison (p<0.05) 
 
（様式５）(Style5) 
 
Fig. 7 Effect of uniconazole (Uni), GA3, and dAll-C10 amide on rice seedlings. Relative lengths (% of 
control) of shoot and second leaf sheath of rice at 7 days after treatment. Values are mean ± SEM (n = 
14), and bars with different letters indicate significant difference as determined by Tukey’s honestly 
significant difference comparison (p<0.05) 
 
Next, the effect of dAll-C10 amide on the GA signaling pathway in rice was examined using another 
known growth retardant, uniconazole, which inhibits the conversion of ent-karuene to ent-karuonic 
acid in an earlier stage of gibberellin biosynthesis (Rademacher 2000). Co-addition of GA3 and 
dAll-C10 amide on rice seedlings pre-treated with uniconazole resulted in similar recovery (Fig. 7), 
suggesting that dAll-C10 amide does not inhibit GA signaling in the way D-allose does. This is the 
main difference with D-allose beacause it has been shown that D-allose inhibits the gibberellins signal 
transduction pathway; thus growth seedlings and elongation of the second leaf sheath of rice were 
inhibited (Fukumoto et al. 2011, 2013).  
Therefore, it may conclude that rare sugar derivative might mainly block the formation of active 
gibbrellins in the same way to the known plant growth retardants daminozide and 16,17-dihydro-GA5. 
These results suggest that rare sugar derivative could be a new class of lead compounds for plant 
growth regulators similar to known growth retardants and may contribute to establish a biodegradable 
and environmental friendly plant growth regulator.   
 
 
 
 
（様式５）(Style5) 
                                References 
 
Afach G, Kawanami Y, Izumori K (2005) Synthesis of D-allose fatty acid esters via lipase catalyzed 
regioselective transesterification. Biosci Biotechnol Biochem 69:833–835 
Afach G, Kawanami Y, Kato-Noguchi H, Izumori K (2006) Practical production of 
6-O-octanoyl-D-allose and its biological activity on plant growth. Biosci Biotechnol Biochem 
70:2010–2012 
Aktar MW, Sengupta D, Chowdhury A (2009) Impact of pesticides use in agriculture: their benefits 
and hazards. Interdisc Toxicol 2(1):1–12 
Bhuiyan SH, Itami Y, Rokui Y, Katayama T, Izumori K (1998) D-allose production from D-psicose 
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Izumori K, Ohara T, Shigematsu Y, Tanaka K, Ishida Y, Nishizawa Y, Tada Y, Ichimura K, Gomi 
K, Akimitsu K (2011) Rare sugar D-allose suppresses gibberellin signaling through 
hexokinase-dependent pathway in Oryza sativa L. Planta 246(6):1083–1095 
Fukumoto T,  Kano A,  Ohtani K, Inoue M, Yoshihara A,  Izumori K, Tajima S, Shigematsu Y, Tanaka K,  
Ohkouchi T,  Ishida Y,  Nishizawa Y, Tada Y, Ichimura K, Gomi K,  Yoo S, Sheen J, Akimitsu K (2013) 
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inhibition in Oryza sativa L. Planta 237:1379–1391 
Gao D, Kawai N and Tamiya T (2011) The anti-inflammatory effects of D-allose contribute to 
attenuation of cerebral ischemia-reperfusion injury. Med Hypotheses 76:911-13 
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2,3,4,6-tetra-O-acetyl-1,5-anhydro-D-ribo-hex-1-enitol. Carbohydrate Research 34:214–218 
Hossain MA, Wakabayashi H, Goda F, Kobayashi S, Maeba T, Maeta H (2000) Effect of the 
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Transplant Proc 32:2021–2023 
Izumori K (2002) Bioproduction strategies for rare hexose sugars. Naturwissenschaften 89:120–124 
Izumori K (2006) Izumoring: a strategy for bioproduction of all hexoses. J Biotechnol 124:717–722 
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against plant growth. Biosci Biotechnol Biochem 74:216–217 
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characteristics, and potential health benefits. Am J Clin Nutr 62:1161S-1168S 
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of pesticide effects in soil. Ambio 27:24–28 
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296–310 
Rademacher W (2000) Growth retardants effects on gibberellins biosynthesis and other metabolic 
pathways. Annu Rev Plant Physiol Plant Mol Biol 51:501–531  
Sui L, Dong Y, Watanabe Y, Yamaguchi F, Hatano N, Izumori K (2005) Growth Inhibitory Effect of D-allose on 
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Takeshita K, Suga A, Takada G, Izumori K (2000) Mass production of D-psicose from D-fructose by a 
continuous bioreactor system using immobilized D-tagtose 3-epimerase. J Biosci Bioeng 
90:453-455  
 
 
 
 
 
 
 
 
 
(注) 要約の文量は，学位論文の文量の約１０分の１として下さい。図表や写真を含めても構いません。
(Note)  The Summary should be about 10％ of the entire dissertation and may include illustrations 


Synthesis of Rare Sugar Derivatives and Their Biological Activity on Plant Growth

Khairul, Hafiz bin Mohd Yusoff

（第３号様式）(Form No. 3) 
学  位  論  文  要  旨  
Dissertation Abstract 
 
 
氏名：  
Name 
Khairul Hafiz bin Mohd Yusoff  
 
学位論文題目：  
Title of Dissertation 
Influences of Agricultural Landuse on Soil Morphological and 
Physicochemical Properties on Sandy Beach Ridges Interspersed 
with Swales in East Coast of Peninsular Malaysia 
(半島マレーシア東海岸における浜堤砂質土壌の断面形態と理
化学性への農業活動による影響 ) 
 
 
 
 
 
 
 
 
 
 
 
 
 
学位論文要旨：  
Dissertation Abstract 
 
Sandy Beach ridges interspersed with swales (BRIS) distribute along the east coast of 
Peninsular Malaysia. Although being regarded as a problema tic lowland soil because of 
its very sandy texture and concomitant infertility, BRIS soils have been used for 
farming practices with recently expending and elevating impacts. The purpose of this 
study was to evaluate the influences of agricultural activiti es on BRIS soils with special 
reference to the contrasting farming practices; conventional fruit gardens with a 
relatively intensive managements and local people’ homegardens. The study area were 
located in Cherating District, Pahang state, the east coast of peninsular Malaysia. Typic 
Quartzipsamments distributed close to the shoreline while Typic Haplorthods and Typic 
Haplohumods in the inland-ward location. 
 
A) Influences of fruit garden with intensive management on the BRIS soils  
Three one-kilometer transects from the shoreline running inland were 
established; two transects passed through the experimental field of Malaysian 
Agriculture Research Institute (MARDI) while one transect passed through an adjacent 
remnant forest. In the field, a high input of chemical fertilizers and organic amendments 
were applied upon planting, followed by monthly application of NPK compounds and 
urea. Major findings were; 1) The A horizons of soils in the MARDI field had lower in 
T-C and T-N compared with those in the forest, suggesting loss of soil organic matter in 
the former due enhanced decomposition and insufficient input of organic materials to 
（第３号様式）(Form No. 3) 
sustain the level of soil organic matter. 2) While the levels of pH, and exchangeable Ca 
and Mg of the soils in the MARDI were higher than those in the forest, that of 
exchangeable K was occasionally under the detection limit despite of fertilizer 
application. This phenomenon might be ascribable to the fact that variable negative 
charges on soil organic matter surfaces with preferen tial adsorption of divalent over 
monovalent cations. 3) The Spodosols in the inland -ward location in the MARDI showed 
the highest peak of available P in the Bhs horizon despite its low level in the overlying 
A and E horizons suggesting the downward translocation of mineral P from fertilizer and 
its retention in the Bhs horizon with higher contents of Fe and Al oxides.  
 
B) Influences of homegarden by local people on the BRIS soils  
The survey was conducted in the Sungai Ular village. Homegardens in the villa ge 
were divided into the inland-ward area (HG-I) and the shoreline area (HG-S) in terms of 
soil types as well as residents’ types. Ash and charcoal produced from burning garbage 
and plant litter, with a few amount of chemical fertilizers if any, were appli ed to both 
types of the homegardens. The difference of the managements in the homegarden types 
was found in the application of homemade-manure in HG-I and the application of 
seafood waste from fish processing in HG-S. Major findings were; 1) In comparison 
with the MARDI, HG-I soils showed higher levels of soil organic matter and nutrients. 
It is suggested that the cumulative effects of the combined application of charcoal and 
manure in the HG-I contributed to the augmentation of the soil organic matter pool . 2) 
In HG-I, the levels of T-C, T-N and CEC at the 0-10 cm depth were tended to be high 
with increasing ages of homegarden. 3) The soils in HG -I were also significantly higher 
in T-C and T-N and CEC values, compared with HG-S. In contrast, the level of available 
P was higher in HG-S than HG-I, which might be ascribable to seafood waste 
application in the shoreline area. 4) However, investigation of soil profile reveals that 
the levels of exchangeable Ca and K and available P tended to be high down to deepe r 
soil layers, suggesting that significant amounts of nutrients leached down beyond plant 
rooting depth. In this sense, unlike tropical homegardens in the other environment, the 
BRIS homegarden could not be regarded as closed, sustainable systems.  
 
C) Conclusion  
In comparison with conventional cropping systems which rely on heavy input of 
chemical fertilizer, management practices employed in the homegarden can be 
considered more appropriate on the BRIS to maintain soil fertility for sustainable 
agriculture. 


Influences of Agricultural Landuse on Soil Morphological and Physicochemical Properties on Sandy Beach Ridges Interspersed with Swales in East Coast of Peninsular Malaysia

Ehime University (愛媛大学)博士(学術)thesi

Winnie, Naa Adjorkor Sowah

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学位論文全文に代わる要約 
Extended Summary in Lieu of Dissertation 
 
 
氏名： 
Name 
Winnie Naa Adjorkor Sowah 
 
学位論文題目：    
Title of Dissertation 
A study on the genetic structure and diversities of tilapia from Ghana and Japan 
（ガーナと日本におけるティラピアの遺伝的集団構造と遺伝的多様性） 
 
 
学位論文要約： 
Dissertation Summary 
 
Tilapia is native to Africa. However, it has been introduced to many tropical and subtropical 
regions in the world. They belong to the family Cichlidae which has at least 1650 species. Three 
genera are commonly referred to as tilapia; Oreochromis, Sarotherodon and Tilapia. Oreochromis spp 
and Sarotherodon spp comprise the mouthbrooders and Tilapia spp are substrate brooder. However, 
the distinguishing feature between Oreochromis spp and Sarotherodon spp is for the former, the 
females incubate the eggs whiles in the latter both male and female incubate the eggs. 
Tilapias are the third most farmed fish resource after carps and salmonids. In 1978, Nile tilapia 
(Oreochromis niloticus) was introduced to China which currently leads in world production of tilapia 
and has consistently produced more than half the global production every year from 1992 to 2008. 
Tilapia introduced to Japan in 1962 was distributed from Hokkaido to Kyushu and Okinawa. Nile 
tilapia from Cote D’Ivoire was introduced to Brazil in 1971 and from Brazil it was sent to the USA in 
1974. Due to certain characteristics, they are easily bred which makes them an excellent choice for 
aquaculture. These include, ease of reproduction, adaptability to intensive culture, acceptability of low 
input sustainable feeds, resistance to impaired water quality, resistance to diseases, fast growth, 
efficient use of natural aquatic food, tendency to consume a variety of supplementary feeds, 
herbivorous nature and widespread consumer acceptance (Balarin et al., 1979). 
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From the 1980s, tilapia was introduced as aquaculture species that are often farmed in cages in 
open bodies of water. The fish can escape if the cage becomes damaged due to environmental forces, 
such as hurricanes, storms, or human actions. Throughout the world, documented cases of tilapia 
introductions are frequently due to both release and escape (ISSG, 2006). Blue tilapia (Oreochrmomis 
aureus) were introduced to Gulf states for weed control, in other cases it was for weed and insect 
control. They also have been released from aquariums and fish farms (GSMFC, 2003). Tilapia that 
have escaped from aquaculture facilities may interbreed and form hybrids (Costa-Pierce, 2003). Some 
species such as the Mozambique tilapia (Oreochromis mossambicus) have outcompeted native fish 
species and preyed on native larval fish in areas where it was introduced. Some species of tilapia can 
displace other species an example is, in Singapore, the problem with tilapia as an invasive species may 
be on its way of solving itself. During World War II, Oreochromis mossambicus was introduced to 
Singapore by the Japanese (this species has been named among the world’s 100 most invasive species 
(Lowe et al., 2000)) soon, it became a common fish in both fresh and brackish water, as well as in the 
sea off the northern coast. Since the late 1980s, these feral population have been declining dramatically 
and this is believed to be caused by the introduction of other tilapias, probably the hybrid between 
Oreochromis mossambicus and Oreochromis niloticus and possibly also between O. mossambicus and 
O. urolepis hornorum or O. aureus. When Oreochromis mossambicus mates with certain species and 
hybrids, the resulting batches tend to have a much skewed sex ratio where almost all fishes are male. It 
is also common for hybrids to produce a lower number of fry per spawning. In addition to this, it is not 
unreasonable to assume that offspring containing genetic material from Oreochromis niloticus risk 
losing the high salinity tolerance of Oreochromis mossambicus and instead get the low salinity 
tolerance of Oreochromis niloticus. 
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The characteristics that make it suitable for aquaculture also enable it to wreak damage in 
places where their growth is not monitored especially where they are introduced as a form of pest 
control. They have become invasive species in many areas notably Australia, where their culture is 
strongly prohibited. In other places, they have managed to render other species within the same trophic 
level extinct or reduced their populations drastically due to completion and their ability to survive on 
many kinds of feed. 
Historically, identification of tilapias has been based mainly on morphological and behavioural 
characteristics. In recent times, molecular markers are employed to give more conclusive results and to 
identify subpopulations within a species. The most commonly used markers are allozymes, 
mitochondrial DNA (mtDNA), Restriction Fragment Length Polymorphisms (RFLPs), Random 
Amplified Polymorphic DNA (RAPD)s, Amplified Fragment Length Polymorphisms (AFLP)s, 
microsatellites (SSR), Single Nucleotide Polymorphisms (SNPs), and Expressed Sequence Tags 
(ESTs). 
In this study, mtDNA control region was used to identify and determine nucleotide and 
haplotype diversities within populations and between populations. It has been reported to better 
differentiate subpopulations due to the high variation observed.  
Samples were collected from six locations in Ghana, the Avu Lagoon in the Volta region, 
Weija Lake, Densu Estuary and Kpeshie Lagoon in the Greater Accra region, the Fosu Lagoon in the 
Central region and the Barekese dam in the Ashanti region. Genetic material was extracted by the 
standard phenol chloroform method. Polymerase chain reaction (PCR) was done with the use of 
primers L19 and 98H. This was followed by sequencing. Sequencing data analysis resulted in the 
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identification of six species; Oreochromis niloticus, Sarotherodon melanotheron, S. galilaeus, Tilapia 
zillii, T. rendalli and Hemichromis elongates (Sowah et al., 2016a).  
Sequence data from Ghana was compared to sequence data from Japan and Thailand to trace 
the roots of some unknown species that were observed in Kochi, Japan. This showed no similarities, 
therefore, more locations need to be investigated to get a definite identification for Kochi’s population. 
Many tilapia species have been introduced to Japan over the years. According to the Food and 
Agricultural Organization (FAO) of the United Nations (UN), the introduction of tilapia to Japan 
began in the 1950’s with the initial introduction in 1954. The first species introduced to Japanese 
freshwaters was the Mozambique tilapia (Oreochromis mossambicus) from Thailand. In all, nine 
species were introduced from various locations around the world, but two of those species Tilapia zillii 
and Oreochromis niloticus directly introduced from Africa (Egypt) in 1962 (De Silva et al., 2004). 
However, they were not monitored over the years so there is little or no information on their genetic 
structure and the impact of their introduction to the local population of freshwater fish. It is uncertain 
whether the tilapia in Japan were intentionally released or accidentally escaped from culture farms. 
Their prolific reproduction led to overpopulation which usually resulted in competition with 
co-occurring species (Noto Mihardio, 1986). Tilapias are considered invasive species in Australia 
where their sale or breeding, even on a subsistence basis is considered a crime 
(http://www.dpi.nsw.gov.au/fishing/pests-diseases/freshwater-pests/species/tilapia.). 
On this premise, samples were collected from Nagoya, Yamaguchi, Kochi, Oita and 
Kagoshima (2 locations) and these were transported to the laboratory for further analysis. Extraction of 
DNA was done with the phenol chloroform method. This was followed by PCR with the ORMT-F 
and ORMT-R primers (Wu et al., 2012). Sequencing was done for these samples. 
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Samples collected were identified as; O. niloticus, O. mossambicus and T. zillii. Oita was 
recorded to have only O. mossambicus, while T. zillii was recorded in one location in Kagoshima 
(Kagoshima-T). The other locations had both O. mossambicus and O. niloticus	(Sowah	et	al.,	2016b)	 	
However, our study was unable to distinguish hybrids from pure strains, as mtDNA is 
maternally inherited. Therefore, the use of other molecular markers such as microsatellite DNA is 
required in near future works to determine the level of hybridization. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
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(注)	 要約の文量は，学位論文の文量の約１０分の１として下さい。図表や写真を含めても構いません。
(Note)  The Summary should be about 10％ of the entire dissertation and may include illustrations 


A study on the genetic structure and diversities of tilapia from Ghana and Japan

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