초록

1. 면역증강 사료첨가제 생강 분말/추출물 첨가 BFActiva의 제조공장 등록 및 상품화

2. 신기능성 특허균주 Lactobacillus plantarum PSCPL13 균주를 활용한 BFProbi의 제조공장 등록 및 상품화

(출처 : 보고서 요약서 3p)

Abstract ▼

III. RESULTS OF THE STUDY
For mass production process of BFP-1, we used Lactobacillus plant arum PSCPL13, patented strain, as an industrial strain. This strain showed salt tolerance and acid resistance. To make industrial media for mass culture of the strain, we used CSL to replace expensive yeas

III. RESULTS OF THE STUDY
For mass production process of BFP-1, we used Lactobacillus plant arum PSCPL13, patented strain, as an industrial strain. This strain showed salt tolerance and acid resistance. To make industrial media for mass culture of the strain, we used CSL to replace expensive yeast extract as the primary nitrogen source. As a result, a complex medium of CSL 50 g/L and yeast extract 3 g / L showed the same production when incubated with 15 g/L yeast extract. However the culture time must be extended to 6 hours. The complex media can reduce the manufacturing cost.

The optimum extraction conditions of yacon and ginger showed the extraction temperature and time were 80℃ and 3 hours in terms of Brix. The study was carried out to establish indicator ingredients of ginger and yacon (BFA - 1). The indicator materials of ginger and yacon were determined by zingiberene and chlorogenic acid, respectively.

Toxic substance residual analysis of prototype BFA —1 was carried out to evaluate by using ICP analysis device. The reason is yacon and ginger planted in the soil due to possibility of residual heavy metals. Antibacterial activity of BFA —1 was examined by the killing rate curve to confirm bactericidal or bacteriostatic effects. As the results, there are no heavy metals in yacon and ginger. In antibacterial activity, both yacon and ginger extract inhibited the lag phase of fish pathogenic activity as well as exhibited the antioxidant activity.

BFA — 1 and BFP — 1 were carried out for the safety assessment of a single oral toxicity study in the rat at GLP institution. LD₅₀ of BFA —1 and BFP — 1 was confirmed more than 2000 mg/kg bw.

BFA — 1 and BFP — 1 were carried out to obtain fish toxicity because they are products intended for fishery. Carp was used for fish toxicity for the 96 hour exposure at limited concentration of BFA — 1 10 mg/L and 100 mg/ L, respectively. The half lethal concentration (LC₅₀) of BFA — 1 is more than 10.0 mg/L since we could not observed swimming inhibition for 96 hours. The half lethal concentration (LC₅₀) of BFP — 1 is more than 100.0 mg/L since we could not observed swimming inhibition for 96 hours.

After oral administration of the lactic acid bacterium, the adherence at intestine, the distribution, the pharmacokinetic experiment was carried out for suggesting optimal dosage. PSC PL13 was administered orally with two doses of 10⁶ and 10⁹ CFU/mL corresponding with 10⁹ and 10¹² CFU/ L . The result confirmed that PSCPL13 was adhered to intestine at 12 hours after administration. In probiotic pharmacokinetic, the biological half—life of high dose (10¹² CFU/ L) and low dose (10⁹ CFU/ L) showed 9.13 hours and 6.82 hours.

In order to confirm the safety and pathogenic for the Lactobacillus plantarum strains, it was performed PSCPL13 single oral toxicity tests. As the results, the culture broth of L. plantarum PSC PL13 and BFP — 1 did not show the pathogenicity and toxicity. Also, Lacrobacillus is one of GRAS.

For the quality control of raw materials and products of the BFA —1 was subjected to biological activity and physicochemical evaluation along with assessment. All samples tested had an concentration of 1,024 ㎍ / ml as a result could not recognize the antimicrobial activity. Three fish pathogenic strains compared to the untreated group at 10³ CFU / mL inhibit the growth of bacteria were effective.

In Korean rockfish (Sebastes schlegelii) infected artificially with Streptococcus iniae infections, Feed administration of yacon and ginger extract showed the inhibition of cumulative mortality as compared with control group (P < 0.05). The result indicate that they can be used antibacterials at the beginning of bacterial infection. Similarly, the groups with addition of feed yacon and ginger extract were compared to the control group in terms of the cumulative mortality. Both yacon and ginger administration was observed to decrease the cumulative mortality when compared to the control. In flounder (Paralichthys olivaceus) infected artificially with Edwardsiella tarda, the groups of yacon extract (Yacon extract, YCE) extracts added to feed showed the inhibition of cumulative mortality as compared with that of control group, indicating the potential one of feed additives. Also, in Vibrio anguillarum infection experiment, they showed the same results.

In the single oral toxicity test of the prototype BFA —1 with BFP — 1 in Flounder, the experiment period was for 14 days. During the test, we could not observe abnormal clinical symptom such as swimming and coloring blackening of the fish. In conclusion, the prototype BFA —1 and BFP — 1 was determined LC₅₀ was more than 2,000 mg/kg bw.

After inoculation with fish pathogenic bacteria, immune activity and mortality test of the prototype BFA —1 with BFP — 1 were tested. After the inoculation attack (S. iniae), the mortality rate of BFA—1 과 BFP — 1 were a 47.5% and 55%, respectively. The untreated group showed a mortality of 72%. On the other hand after the attack inoculated with E. tarda, the cumulative mortality in BFA —1 and BFP — 1 showed 65% and 68% and non—treated group showed a 100% mortality rate.

For clinical trials of phase 3, the prototype BFP — 1 administration was performed in the outdoor test fishery located in Jeju Island and Gangwon. To prevent lactic acid bacteria spread into water, lactic acid is adsorbed into the vermiculite according to company’s patented manufacturing process. A s the results, the total number of bacteria in the intestinal bacteria showed a similar level in BFP — 1 test group and the control group before the experiment. After the administration of BFP — 1, the BFP — 1 group showed the increased lactic acid bacteria grown on MRS agar. In metagenome analysis, the BFP — 1 group showed 16 phylum and 96 genus and the control group showed 15 phylum and 52 genus. In comparison of intestinal bacterial communities in groups, BFP — 1 test and control group exhibited the 13.2% and below 1% of lactic acid bacteria, respectively.
Interestingly, BFP — 1 test and control group also exhibited the 13.2% and below 1% of Vibrio, respectively. From the result, the function of BFP — 1 in flounder was confirmed that it tends to promote the growth of lactic acid bacteria and suppresses the growth of harmful bacteria.

(출처 : SUMMARY 14p)

목차 Contents

• 표지 ... 1
• 제 출 문 ... 2
• 보고서 요약서 ... 3
• 요 약 문 ... 4
• SUMMARY ... 14
• Contents ... 18
• 목차 ... 21
• 제 1 장 연구개발과제의 개요 ... 24
• 제1절 연구개발의 개요 ... 24
• 1. 연구개발의 목적 ... 24
• 2. 연구개발의 필요성 및 범위 ... 29
• 제 2 장 국내외 기술개발 현황 ... 37
• 1. 사료첨가제 시장 동향 ... 37
• 2. 국내 사료시장 ... 40
• 3. 국외 주요 사료회사 ... 41
• 4. 국내외 기술 수준 ... 42
• 제 3 장 연구개발수행 내용 및 결과 ... 45
• 제1절 BFP-1의 대량생산 공정 확립 ... 45
• 1. 시험목적 ... 45
• 2. 재료 및 방법 ... 45
• 3. 결과 및 고찰 ... 49
• 제2절 시제품 BFA-1 대량생산 공정 확립 ... 56
• 1. 시험목적 ... 56
• 2. 재료 및 방법 ... 56
• 3. 결과 및 고찰 ... 59
• 제3절 BFA-1 의 유해물질 잔류와 항균 활성 평가 ... 65
• 1. 시험목적 ... 65
• 2. 재료 및 방법 ... 65
• 3. 결과 및 고찰 ... 68
• 제4절 Rat에서 시제품 BFA-1 와 BFP-1의 안전성 평가 ... 71
• 1. 시험목적 ... 71
• 2. 재료 및 방법 ... 71
• 3. 결과 및 고찰 ... 73
• 제5절 시제품 BFA와 BFP의 어류독성평가 ... 80
• 1. 시험목적 ... 80
• 2. 재료 및 방법 ... 80
• 3. 결과 및 고찰 ... 82
• 제6절 BFP-1 (Lactobacillus plantarum PSCPL13)의 경구 투여후 넙치의 장내 미생물 분리 동정 ... 87
• 1. 시험목적 ... 87
• 2. 재료 및 방법 ... 87
• 3. 결과 및 고찰 ... 89
• 제7절 BFP (Lactobacillus plantarum PSCPL13)의 단회경구독성시험 ... 93
• 1. 시험목적 ... 93
• 2. 재료 및 방법 ... 93
• 3. 결과 및 고찰 ... 94
• 제8절 BFA (야콘, 블루베리, 생강) 생물 활성 평가 ... 101
• 1. 시험목적 ... 101
• 2. 재료 및 방법 ... 101
• 3. 결과 및 고찰 ... 105
• 제9절 조피볼락용 병원성 세균 억제 및 면역증강 양어 기능성 사료첨가제 개발 ... 109
• 1. 실험목적 ... 109
• 2. 재료 및 방법 ... 109
• 3. 결과 및 고찰 ... 122
• 제10절 설치류에서 시제품 BFP-1의 단회경구독성시험 ... 163
• 1. 시험목적 ... 163
• 2. 재료 및 방법 ... 163
• 3. 결과 및 고찰 ... 167
• 제11절 설치류에서 BFA-1의 단회경구독성시험 ... 173
• 1. 시험목적 ... 173
• 2. 재료 및 방법 ... 173
• 3. 결과 및 고찰 ... 177
• 제12절 넙치에서 BFA-1의 안전성 시험 ... 181
• 1. 시험목적 ... 181
• 2. 재료 및 방법 ... 181
• 3. 결과 및 고찰 ... 183
• 제13절 넙치에서 시제품 BFP-1의 안전성 시험 ... 188
• 1. 시험목적 ... 188
• 2. 재료 및 방법 ... 188
• 3. 결과 및 고찰 ... 190
• 제14절 넙치에서 신규 Lactobacillus plantarum PSCFL13의 경구투여 후 약동학 ... 195
• 1. 시험목적 ... 195
• 2. 재료 및 방법 ... 196
• 3. 결과 및 고찰 ... 197
• 제15절 넙치에서 어류병원성세균 접종 후 시제품 BFP-1과 BFA-1의 면역활성 평가 ... 199
• 1. 시험목적 ... 199
• 2. 재료 및 방법 ... 199
• 3. 결과 및 고찰 ... 202
• 제16절 넙치 (Paralichthys olivaceus)에서 시제품 BFP-1 투여에 의한 효능 시험(동일수산, 강원도) ... 208
• 1. 시험목적 ... 208
• 2. 재료 및 방법 ... 208
• 3. 결과 및 고찰 ... 214
• 제17절 넙치에서 시제품 BFP-1의 효능평가 및 식품품질분석(제주도, 용천수산) ... 227
• 1. 시험목적 ... 227
• 2. 재료 및 방법 ... 227
• 3. 결과 및 고찰 ... 229
• 제 4 장 목표달성도 및 관련분야에의 기여도 ... 247
• 제1절 목표 및 연구결과 ... 247
• 제2절 관련 분야 기술 발전 기여도 ... 252
• 제 5 장 연구개발결과의 활용계획 ... 254
• 제1절 추가연구의 필요성 ... 254
• 제2절 타 연구에의 이용 ... 255
• 제3절 기업화 추진방안 ... 256
• 제4절 성과 활용 및 사업화 프로세스 ... 257
• 제 6 장 연구개발과정에서 수집한 해외과학기술정보 ... 261
• 제1절 특허 정보 기술 ... 261
• 제2절 논문 정보 기술 ... 263
• 제3절 해외 제품 정보 기술 ... 265
• 제 7 장 참고문헌 ... 267
• 끝페이지 ... 273