Study on Potential Allergenicity of TERASI, Indonesian Shrimp Seasoning, and its Reduction by Processing Technology

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Study on Potential Allergenicity of TERASI, Indonesian Shrimp Seasoning, and its Reduction by Processing
Technology

Amalia, Ulfah

北海道大学. 博士(水産科学) 甲第15252号

2023-03-23

10.14943/doctoral.k15252

http://hdl.handle.net/2115/91927

theses (doctoral)

Ulfah_Amalia.pdf

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Hokkaido University Collection of Scholarly and Academic Papers : HUSCAP

Study on Potential Allergenicity of TERASI,
Indonesian Shrimp Seasoning,
and its Reduction by Processing Technology
(インドネシア産エビ調味料テラシのアレルギー誘発性と

加工技術による低減化に関する研究)

Division of Marine Life Science
Graduate School of Fisheries Sciences
Hokkaido University
北海道大学大学院水産科学院
海洋応用生命科学専攻

Ulfah Amalia
ウルファ アマリア

2023 年

主論文の要約
博士の専攻分野の名称：博士（水産科学）

氏名: Ulfah Amalia (ウルファ アマリア)

学位論文題目

Study on Potential Allergenicity of TERASI, Indonesian Shrimp Seasoning, and
its Reduction by Processing Technology
(インドネシア産エビ調味料テラシのアレルギー誘発性と
加工技術による低減化に関する研究)

The objective of this study is to investigate the potential allergenicity of Terasi,
Indonesian fermented shrimp seasoning and discussing the processing technology for the
reduction of the IgE-binding ability of Terasi final products. The PhD thesis consists of four
chapters including one chapter overall explaining this study and three chapters presenting
results and discussion of the research.
Chapter 1. (Introduction). Terasi is a popular seasoning of Indonesian traditional
fermented shrimp paste with unique flavor and excellent nutritional value, which made by
mixing dried small shrimp with salt (10–15% w/w). The mixture is grinded and spread out on
the ground to dry with sunlight. The salted shrimp meat is solid and subjected to ferment in
anaerobic condition at ambient temperature (25–28 ℃) for at least 3 days or longer until the
unique aroma has fully established. It is reasonable to regard Terasi as a significant component
of human diet and nutrition worldwide. However, shrimp, the raw material of Terasi contains
allergenic protein, tropomyosin (TM), which is a common allergen of invertebrates, such as
crab, squid, octopus, and shellfish. Several studies have been reported that proteolysis due to
food fermentation reduced food allergenicity, but no investigation about the potential
allergenicity of Terasi.
Chapter 2. The study evaluated the safety of Terasi (Indonesian fermented shrimp paste),
the product characteristics and allergenicity of 20 types of Indonesian commercial Terasi (CT)
that meet the Indonesian National Standard were evaluated with a focus on the major shrimp
allergen TM. Marked protein hydrolysis of shrimp muscle occurred in all CT samples, and no
protein fragments or specific reaction of anti-TM IgG were observed in SDS-PAGE and
immunoblot assays. In a competitive enzyme-linked immunosorbent assay using shrimp
allergenic patient sera, it was observed a markedly diminished specific IgE reaction of CT
compared with that of shrimp muscle, whereas the IgE-binding ability remained in all CT
samples. No clear correlation was found between the degree of protein hydrolysis and IgE
reactivity. These results indicate that CT could be defined as a low–allergenic processed
seafood but has the possibility to be a causative food for shrimp allergy. Direct immunological
evaluation is required establishing the food safety of CT, because assessments of protein

profiles and hydrolysis are not useful for determining the safety of Terasi. Moreover, among
CTs, raw materials and protocol used are less informed; therefore, the following chapters
discuss their potential.
Chapter 3. This study examined the effect of the Terasi manufacturing process on the
loss of the allergen TM and its IgG/IgE-binding ability. Terasi was produced from three
shrimps, Akiami (Acetes japonicus), Okiami (Euphausia pacifica), and Isazaami (Neomysis
awatchensis). Protein degradation and TM IgE-binding activity were examined by
immunoblotting using anti-TM rabbit IgG and competitive enzyme-linked immunosorbent
assays using shrimp-allergic patients’ sera. TM in the materials was degraded during the
manufacturing process, and the IgG-specific response in Akiami meat disappeared at the
second fermentation step but remained in both Okiami and Isazaami Terasi. In contrast, TM
IgE-binding ability of TM in all shrimp meats decreased gradually with the progress of the
manufacturing process and nearly disappeared in Akiami Terasi (AT), indicating Terasi can be
recognized as a low allergenic seafood. However, in order to ensure food safety of Terasi, an
effective mean for decreasing IgE-binding ability of the final product should be introduced to
the Terasi manufacture, because progress of the IgE-binding loss varied depending on raw
materials.
Chapter 4. As countermeasures against the technical issues raised in Chapter 3, the
backslopping method was applied to the Terasi manufacture, and its contribution to reducing
the potential allergenicity of Terasi was examined. That is, three kinds of starters, the low
allergenic commercial Terasi as CT, AT (produced in Chapter 3) and HAT (heat treated AT)
were added to manufacturing process of the Isazaami Terasi which highly remained IgEbinding ability. This chapter demonstrated that backslopping method using Terasi products is
an effective manner to produce low allergenic Terasi by inducing reduction of IgE-binding
ability of TM. Addition of the starter accelerated the fermentation of the raw material,
effectively promoting the degradation of the shrimp protein and the reduction of the IgEbinding ability of TM. However, the backslopping effect was dependent on the type of Terasi
used as a starter, and the commercial Terasi used in this study did not contribute to the
allergenicity reduction of the final product. Interestingly, Terasi added as a starter would have
acted primarily as a nutrient source to promote microbial fermentation rather than as a source
of fermenting microorganisms and endogenous proteases.
Conclusively, Terasi manufacture is an effective manufacturing process to reduce the
IgE-binding ability of TM and to ensure the low allergenicity potential. Terasi can be
recognized as a low allergenic seafood when produced under an appropriate manufacturing
condition. It is probable that the measurement of the whole protein hydrolysis is effective for
allergenicity evaluation of Terasi under certain conditions, and it could be used as a screening
index. However, analysing IgE-binding activity of the final products is the most important
manner to estimate Terasi as a low allergenic seafood. It should be noted that the backslopping
method using Terasi final products could contribute to improving food safety of Terasi by
reducing the IgE-binding ability of the final products. I believe that these research results will
definitely help ensuring food safety of Terasi.

ACKNOWLEDGEMENT
The journey that one has to embark on, to attain a Doctoral degree, is an arduous one. The
foremost challenge in this journey is that of the uncertainty of things that may occur in the
foreseeable future and the risk that one is willing to take to overcome them. But then,
innovation by its very nature involves risk, and innovation is the very essence of the research
leading to a higher degree. The success achieved at the end of this path is because of a collective
effort and hard work of the many; and I take this opportunity to express my gratitude to
everyone who supported me throughout my journey.
At this moment of accomplishment, first and foremost, I would like to express my sincere
gratitude to my advisor, Professor Hiroki Saeki. He has been a source of constant motivation
and support not only in my academic life but also in my personal life. His distinctive outlook
towards life, work, and family has been an inspiration for me to be a better person. His positive
attitude and strategic thinking towards research have been a unique example in shaping my
professional conduct. I thank SAEKI Sensei for helping me develop my research career; this
PhD thesis would not have been possible without his support and advice.
I would like to thank my co-advisor Dr. Yutaka Shimizu for his willingness to spend time
out of his busy schedule to help me with the practical aspects of my work and valuable advice
and suggestion during my Ph.D. progress. A special thanks and acknowledge to Assistant
Professor Ga-Hyun Joe for being a great research student monitor, advising me with valuable
comments on my progress and PhD completion, and for her support with conference and
workshop participations.
I would like to express my sincere gratitude to Professor Koji Yamazaki for reviewing this
thesis and providing insightful comments. Thank to him, I was able to improve the quality of
my doctoral thesis. I am also indebted to Professor Hideyuki Kurihara for the valuable
comments and discussions to improve my doctoral thesis.
I am very grateful to Dr. Kazuhiko Watanabe (Watanabe Kazuhiko Pediatric Clinic,
Sapporo, Japan) for providing access to the sera of shrimp-allergic patients, to Dr. Atsushi
Yamaguchi (Graduate School of Fisheries Science, Hokkaido University) for identifying the
species of Okiami shrimp (Eupashia pacifica) used in my study. I would also like to thank
Associate Professor Fumiaki Beppu for very kind discussion and help on setting the Kjeldahl
apparatus for my experiment.
I am very fortunate and proud to be part of an amazing team of the Allergy Group, and I
would like to express my gratitude to the members, past and present; with special thanks to Mr.
Hirano, Mr. Minamiyama, Mr. Makino, Ms. Dong Su Chi, Ms. Watanabe, Mr. Shibazaki,
and Mr. Kikuchi for not only supporting me with the experimental work but also to make my
life fun outside of the lab. I would also like to extend my gratitude to members of laboratory,
to Mr. Yang Boxue (Ph.D. cand.), Mr. Li Wenzhao (Ph.D. cand.), Mr. Nezaki, Mr. Fujimoto,
Ms. Sakai, Mr. Shimada, Mr. Sato, Ms. Shimada, Mr. ...

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