Toxicological studies of SUNACTIVE Zn-P240 and SUNACTIVE Fe-P80 = SUNACTIVE Zn-P240과 SUNACTIVE Fe-P80의 독성학적 연구

국문 초록 (Abstract)

아연보충제는 정상적인 성장과 발달에 중요한 영양소인 아연의 결핍을 예방하고 치료하는 효과적이며 효율적인 수단이다. 기존 아연 보충제의 단점인 낮은 생체이용률과 풍미 변화 등을 해결하기 위해 나노기술이 적용된 아연보충제가 시판되고 있으나, 이에 대한 안전성 자료는 제한적으로 알려져있다. 본 연구는 Sprague–Dawley 랫드에서 새로운 유형의 아연보충제인 SUNACTIVE Zn-P240의 28일 반복 경구투여로 인해 나타나는 잠재적인 독성영향을 평가하였다. 암·수 각 20마리의 랫드를 무작위법으로 군분리하여 부형제대조군 (멸균증류수)과 SUNACTIVE Zn-P240 500, 1000 및 2000 mg/kg/day 투여군의 4개 시험군으로 구성하였으며, 추가적으로 대조군과 고용량군에서 회복군(군당 5마리)을 두고 2주간 관찰하였다. 시험결과, SUNACTIVE Zn-P240의 28일 반복경구투여는 2000 mg/kg/day 투여군에서 증체량 감소, 사료섭취량 감소, 간 중량의 증가, 그리고 위, 간 및 췌장의 조직병리학적 변화를 초래하는 것으로 확인되었다. 또한, 혈액 및 혈청생화학적 검사에서 알칼라인 포스파타제 (alkaline phosphatase; ALP) 및 알라닌 아미노전달효소 (alanine aminotransferase) 수치 증가와 헤모글로빈(hemoglobin), 헤마토크리트 (hematocrit), 평균적혈구용적 (mean corpuscular volume; MCV), 평균적혈구혈색소량 (mean corpuscular hemoglobin; MCH), 평균적혈구혈색소농도 (mean corpuscular hemoglobin concentration), 총단백질 (total protein; TP)과 알부민 (albumin)의 수치 감소를 유발하였다. 1000 mg/kg/day 투여군에서는 ALP 수치가 증가하였으며, MCV, MCH와 TP 수치가 감소하였다. SUNACTIVE Zn-P240 투여로 인해 관찰된 혈액학적 및 혈청 생화학적 변화들은 암·수 모두에서 명확한 용량의존성을 나타내었으며, 간 중량의 증가 및 조직병리학적 이상소견이 동반된 것으로 보아 시험물질의 투여에 기인된 독성소견으로 판단된다. 따라서, 본 시험조건 하에서 SUNACTIVE Zn-P240의 무독성량(no-observed-adverse-effect level)은 암·수 모두 500 mg/kg/day으로 판단되었고, 표적장기는 위, 간, 췌장, 그리고 적혈구로 확인되었다.
철분제는 철 결핍성 빈혈을 예방할 뿐만 아니라, 체내에 부족한 영양분을 효과적으로 공급해줄 수 있는 주요 수단이다. 기존 철분제의 단점인 낮은 생체이용률과 풍미 변화 등을 해결하기 위해 나노기술이 적용된 철분제가 시판되고 있으나, 이에 대한 안전성 자료는 제한적이다. 본 연구는 Sprague–Dawley 랫드에서 나노기술이 적용된 철분 보충제인 SUNACTIVE Fe-P80의 28일 반복 경구투여로 인해 나타나는 잠재적인 독성영향을 평가하였다. 암·수 각 20마리의 랫드를 무작위법으로 군분리하여 부형제대조군(멸균증류수)과 SUNACTIVE Fe-P80 500, 1000 및 2000 mg/kg/day 투여군의 4개 시험군으로 구성하였으며, 추가적으로 대조군과 고용량군에서 회복군(군당 5마리)을 두고 2주간 관찰하였다. 28일 동안 SUNACTIVE Fe-P80을 반복 경구투여한 결과, 일반증상, 체중변화, 사료섭취량, 요검사, 안과학적 검사, 부검소견, 혈액학적 검사, 혈액생화학적 검사, 장기중량 및 조직병리학적 검사에서 시험물질 투여와 관련된 유의성 있는 독성학적 변화는 관찰되지 않았다. 따라서, 본 시험조건하에서 SUNACTIVE Fe-P80의 무독성량(no-observed-adverse-effect level)은 암·수 모두 2000 mg/kg/day 이상으로 판단되었고, 표적장기는 확인되지 않았다.
본 시험은 3종의 유전독성 표준시험법을 이용하여 SUNACTIVE Fe-P80과 SUNACTIVE Zn-P80의 잠재적인 유전독성을 조사하였고, 경제개발협력기구의 독성시험 가이드라인과 식품의약품안전처의 시험지침 및 우수실험실적격기준 (Good Laboratory Practice)을 준수하여 수행하였다. 세균을 이용한 복귀돌연변이 시험은 대사활성계의 적용 및 미적용하에 전배양 방법을 이용하여 수행하였고, 생체 외 염색체이상 시험은 대사활성계의 적용 및 미적용하에서 배양된 Chinese hamster lung 세포주를 이용하여 수행하였다. 또한, 생체 내 소핵시험은 ICR 마우스를 이용하여 수행하였다. 시험결과, SUNACTIVE Fe-P80과 SUNACTIVE Zn-P80는 Salmonella typhimurium TA98, TA100, TA1535, TA1537 균주 및 Escherichia coli WP2uvrA 균주를 이용한 복귀돌연변이 시험에서 복귀돌연변이를 유발하지 않았고, 생체 외 염색체이상 및 생체 내 소핵시험에서도 염색체 이상이나 소핵유발은 관찰되지 않았다. 따라서, SUNACTIVE Fe-P80과 SUNACTIVE Zn-P80는 돌연변이원성(mutagenic) 및 염색체이상유발성(clastogenic)을 나타내지 않는 것으로 판단되었다.

다국어 초록 (Multilingual Abstract)

This study was conducted to investigate the potential toxicity of repeated oral dose of SUNACTIVE Zn-P240, a new type of zinc supplement, in Sprague-Dawley rats. SUNACTIVE Zn-P240 was administered once daily by gavage at doses of 0, 500, 1000, and 2000 mg/kg/day for each group over a 28-day period. At 2000 mg/kg/day, there were increases in serum alkaline phosphatase (ALP) and alanine aminotransferase, liver weight, histopathological changes in stomach, liver, and pancreas and decreases in body weight, food consumption, hemoglobin, hematocrit, mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration, total protein (TP), and albumin. At 1000 mg/kg/day, there was an increase in the serum ALP level and there were decreases in the MCV, MCH, and TP. There were no treatment-related adverse effects in the 500 mg/kg/day group. Under the present experimental conditions, the target organs in rats were determined to be the stomach, pancreas, liver, and erythrocyte and the no-observed-adverse-effect level (NOAEL) in rats was considered to be 500 mg/kg/day.
This study aimed to investigate the potential toxicity of a 28-day repeated oral dose of SUNACTIVE Fe-P80, a new type of iron supplement, in Sprague-Dawley rats. SUNACTIVE Fe-P80 was administered once daily by gavage to male and female rats for 28 days at doses of 0, 500, 1000, and 2000 mg/kg/day (n = 5, for male and female rats for each dose). Additional recovery groups from the control and high-dose groups were observed for a 14-day recovery period. At the end of the treatment and recovery periods, the animals were sacrificed, their organs weighed, and blood samples collected. There were no treatment-related adverse effects in the context of clinical signs, body weight, food consumption, ophthalmoscopy, urinalysis, necropsy findings, organ weights, and hematologic, serum biochemical and histopathological parameters at any dose tested. Under the present experimental conditions, the no-observed-adverse-effect level (NOAEL) of SUNACTIVE Fe-P80 was ≥ 2000 mg/kg/day in both the sexes, and no target organs were identified.
This study evaluated the potential genotoxic effects of the nutrient supplements SUNACTIVE Fe-P80 and SUNACTIVE Zn-P240 in vitro and in vivo. Genotoxicity tests were performed following the guidelines of the Organization for Economic Cooperation and Development, applying Good Laboratory Practice. A bacterial reverse mutation test was performed using the pre-incubation method, while the in vitro chromosome aberration test was performed using a cultured Chinese hamster lung cell line in the presence or absence of the S9 mixture. The in vivo micronucleus test was performed using ICR mice. The bacterial reverse mutation test revealed that SUNACTIVE Fe-P80 and SUNACTIVE Zn-P240 did not induce genetic mutations at the tested doses in Salmonella typhimurium (TA98, TA100, TA1535, and TA1537) and Escherichia coli (WP2uvrA) strains. Meanwhile, the results of the in vitro chromosomal aberration and in vivo micronucleus tests revealed that SUNACTIVE Fe-P80 and SUNACTIVE Zn-P240 did not induce chromosomal aberrations. These results suggest that SUNACTIVE Fe-P80 and SUNACTIVE Zn-P240 did not exhibit mutagenic or clastogenic properties in vitro and in vivo.

목차 (Table of Contents)

• LIST OF ABBREVIATIONS 1
• LIST OF FIGURES 3
• LIST OF TABLES 4
• GENERLA ABSTRACT 8
• GENERAL INTRODUCTION 11
• REFERENCES 17
• CHAPTER I Evaluation of 28-day repeated oral dose toxicity of SUNACTIVE Zn-P240 in rats 22
• ABSTRACT 23
• INTRODUCTION 24
• MATERIALS AND METHODS 28
• RESULTS 37
• DISCUSSION 57
• REFERENCES 63
• CHAPTER II Evaluation of 28-day repeated oral dose toxicity of SUNACTIVE Fe-P80 in rats 69
• ABSTRACT 70
• INTRODUCTION 71
• MATERIALS AND METHODS 74
• RESULTS 83
• DISCUSSION 98
• REFERENCES 102
• CHAPTER III Evaluation of genotoxicity of SUNACTIVE Fe-P80 and SUNACTIVE Zn-P240 in vitro and in vivo 109
• ABSTRACT 110
• INTRODUCTION 111
• MATERIALS AND METHODS 114
• RESULTS 121
• DISCUSSION 134
• REFERENCES 138
• ABSTRACT IN KOREA 144

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