KR20020072130A - antibacterial-activity material extracted from cactus of opumtia genus - Google Patents
antibacterial-activity material extracted from cactus of opumtia genus Download PDFInfo
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- KR20020072130A KR20020072130A KR1020010012275A KR20010012275A KR20020072130A KR 20020072130 A KR20020072130 A KR 20020072130A KR 1020010012275 A KR1020010012275 A KR 1020010012275A KR 20010012275 A KR20010012275 A KR 20010012275A KR 20020072130 A KR20020072130 A KR 20020072130A
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- cactus
- extract
- antimicrobial
- mbt
- bacteria
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- 241000219357 Cactaceae Species 0.000 title claims description 31
- 230000000844 anti-bacterial effect Effects 0.000 title description 16
- 239000000463 material Substances 0.000 title description 8
- 230000000845 anti-microbial effect Effects 0.000 claims abstract description 65
- 239000000284 extract Substances 0.000 claims abstract description 27
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims abstract description 15
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 claims abstract description 15
- 240000001439 Opuntia Species 0.000 claims abstract description 10
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- 238000006460 hydrolysis reaction Methods 0.000 claims description 18
- 239000002904 solvent Substances 0.000 claims description 7
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- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 abstract description 5
- RJQXTJLFIWVMTO-TYNCELHUSA-N Methicillin Chemical compound COC1=CC=CC(OC)=C1C(=O)N[C@@H]1C(=O)N2[C@@H](C(O)=O)C(C)(C)S[C@@H]21 RJQXTJLFIWVMTO-TYNCELHUSA-N 0.000 abstract description 5
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- RHJZFALAFLMQQW-UHFFFAOYSA-N 1-methylsiline Chemical compound C[Si]1=CC=CC=C1 RHJZFALAFLMQQW-UHFFFAOYSA-N 0.000 description 1
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- 208000018522 Gastrointestinal disease Diseases 0.000 description 1
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 0.000 description 1
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- 230000009471 action Effects 0.000 description 1
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- LKCWBDHBTVXHDL-RMDFUYIESA-N amikacin Chemical compound O([C@@H]1[C@@H](N)C[C@H]([C@@H]([C@H]1O)O[C@@H]1[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O1)O)NC(=O)[C@@H](O)CCN)[C@H]1O[C@H](CN)[C@@H](O)[C@H](O)[C@H]1O LKCWBDHBTVXHDL-RMDFUYIESA-N 0.000 description 1
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- SRZNHPXWXCNNDU-RHBCBLIFSA-N cefotetan Chemical compound N([C@]1(OC)C(N2C(=C(CSC=3N(N=NN=3)C)CS[C@@H]21)C(O)=O)=O)C(=O)C1SC(=C(C(N)=O)C(O)=O)S1 SRZNHPXWXCNNDU-RHBCBLIFSA-N 0.000 description 1
- 229960004755 ceftriaxone Drugs 0.000 description 1
- VAAUVRVFOQPIGI-SPQHTLEESA-N ceftriaxone Chemical compound S([C@@H]1[C@@H](C(N1C=1C(O)=O)=O)NC(=O)\C(=N/OC)C=2N=C(N)SC=2)CC=1CSC1=NC(=O)C(=O)NN1C VAAUVRVFOQPIGI-SPQHTLEESA-N 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 229960002227 clindamycin Drugs 0.000 description 1
- KDLRVYVGXIQJDK-AWPVFWJPSA-N clindamycin Chemical compound CN1C[C@H](CCC)C[C@H]1C(=O)N[C@H]([C@H](C)Cl)[C@@H]1[C@H](O)[C@H](O)[C@@H](O)[C@@H](SC)O1 KDLRVYVGXIQJDK-AWPVFWJPSA-N 0.000 description 1
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- ZSKVGTPCRGIANV-ZXFLCMHBSA-N imipenem Chemical compound C1C(SCC\N=C\N)=C(C(O)=O)N2C(=O)[C@H]([C@H](O)C)[C@H]21 ZSKVGTPCRGIANV-ZXFLCMHBSA-N 0.000 description 1
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- UWYHMGVUTGAWSP-JKIFEVAISA-N oxacillin Chemical compound N([C@@H]1C(N2[C@H](C(C)(C)S[C@@H]21)C(O)=O)=O)C(=O)C1=C(C)ON=C1C1=CC=CC=C1 UWYHMGVUTGAWSP-JKIFEVAISA-N 0.000 description 1
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Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/33—Cactaceae (Cactus family), e.g. pricklypear or Cereus
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
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- Health & Medical Sciences (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Medical Informatics (AREA)
- Mycology (AREA)
- Engineering & Computer Science (AREA)
- Alternative & Traditional Medicine (AREA)
- Biotechnology (AREA)
- Botany (AREA)
- Analytical Chemistry (AREA)
- Medicinal Chemistry (AREA)
- Microbiology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Medicines Containing Plant Substances (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
Description
본 발명은 항생제 내성세균 및 일반세균의 생육저해를 일으키는 항균성 천연물질에 관한 것으로, 보다 상세하게는 천연선인장으로부터 유래되고 추출분획된 약제다제 내성균에 활성을 나타내는 항균 활성물질에 관한 것이다.The present invention relates to an antimicrobial natural substance causing growth inhibition of antibiotic resistant bacteria and general bacteria, and more particularly to an antimicrobial active substance derived from natural cactus and exhibiting activity against drug-drug-resistant bacteria extracted and fractionated.
일반적으로 세균성 질환의 치료를 위하여 많은 항균제가 개발되어 왔다. 최초의 항생제인 페니실린을 비롯하여 수많은 종류의 항균·항생제가 개발되었고 현재도 개발중에 있다. 이러한 항균제의 개발은 최초의 세균성 질환에 대한 치료를 목적으로 개발되었으며, 세균성 질환을 일으키는 균체에 들어가서 세균의 생리작용을 저해함으로써 항균작용을 나타낸다.In general, many antibacterial agents have been developed for the treatment of bacterial diseases. Numerous types of antimicrobials and antibiotics, including the first antibiotic penicillin, have been developed and are currently under development. The development of such an antimicrobial agent was developed for the purpose of treating the first bacterial disease, and enters the cells causing the bacterial disease and exhibits antimicrobial activity by inhibiting the physiological action of the bacteria.
본 발명은 미생물을 대상으로한 종래의 항균물질 생산균의 탐색방법을 선택하지 않고 식물기원에서 새로운 생리활성물질의 검색을 실시해서 얻어진 항균활성물질에 관한것이다.The present invention relates to an antimicrobial active substance obtained by searching for a new bioactive substance in plant origin without selecting a conventional method for searching for antimicrobial substance producing bacteria targeting microorganisms.
각종 항균·항생제가 세균성 질환의 치료제로 사용되어진 이 후, 이러한 항균·항생제에 대해 내성을 나타내는 새로운 내성균이 출현하게 되고, 이러한 내성균에 대한 새로운 항균·항생제의 개발이 이루어지는 과정을 거치고 있으며, 최근에는 이러한 내성균의 출현 속도에 비해 항균·항생제의 개발 속도가 미치지 못하고 있는 실정이다. 이러한 항균·항생제중 반코마이신(vancomycin)은 메틸실린 내성 황색 포도상구균(methicilin-resistantStreptococcus aureus, MRSA)과 다른 그람 양성세균에 의한 감염증을 치료할 수 있는 최후의 향균제로 여겨져 왔으나, 최근 미국, 일본 및 유럽 등지에서 이러한 반코마이신(vancomycin)에 내성을 가지는 새로운 내성세균의 출현이 보고되어 문제의 심각성을 더하고 있으며, 이에 대한 합한 항균제의 개발은 매우 어렵고 약제 내성균에 의한 파국을 방지하기 위해서는 전혀 새로운 치료제의 개발이 강력하게 요구되고 있다.Since various antibacterial and antibiotics have been used as treatments for bacterial diseases, new resistant bacteria that have become resistant to such antibacterial and antibiotics have emerged, and new antibacterial and antibiotics have been developed. Compared with the emergence rate of such resistant bacteria, the development rate of the antimicrobial and antibiotics has not been reached. Vancomycin among these antimicrobials and antibiotics has been considered as the last antifungal agent to treat infections caused by methylsilin-resistant Streptococcus aureus (MRSA) and other Gram-positive bacteria, but recently in the United States, Japan and Europe. The emergence of new resistant bacteria resistant to vancomycin has been reported in Korea, adding to the seriousness of the problem. The development of a new antimicrobial agent is very difficult and the development of a completely new therapeutic agent to prevent catastrophe caused by drug-resistant bacteria. It is strongly demanded.
상기와 같이, 서로의 생존을 위한 인간과 세균의 대결은 항생제 개발과 사용, 내성발현, 또 다른 항생제의 개발이라는 악순환을 되풀이 하고 있다. 최근에는 약제내성 메카니즘의 해명과 더불어 미생물 게놈, 세균의 병독성과 숙주세포흡착 및 침투, 기타 신규타겟을 대상으로 한 항생제 개발은 감염질환 치료에 새로운 장을 열것으로 기대하고 있다. 그러나, 현 시점에서 보면 세균의 항생제에 대한 내성 획득 속도에 비하면 새로운 약제 개발은 만족스럽지 못한 상태이고, 실제 최근에 새로이 승인 받거나 검토중인 항생제중 특이한 것은 몇가지에 불과하다.As mentioned above, the battle between humans and bacteria for each other's survival repeats a vicious cycle of antibiotic development and use, resistance expression, and development of another antibiotic. Recently, the development of antibiotics targeting the microbial genome, virulence and host cell adsorption and infiltration, and other new targets, along with the explanation of drug resistance mechanisms, is expected to open a new chapter in the treatment of infectious diseases. However, at the present time, the development of new drugs is not satisfactory compared to the bacterial acquisition rate of antibiotics, and only a few of the new antibiotics recently approved or under review are unique.
따라서 약제 내성균에 의한 파국을 미연에 방지하기 위해서는 기존의 미생물또는 화학적 유래가 아닌 전혀 새로운 종류의 항생제의 개발이 강력하게 요구되고 있는 실정이다.Therefore, in order to prevent catastrophe caused by drug-resistant bacteria in advance, the development of an entirely new type of antibiotic, which is not an existing microorganism or chemical origin, is strongly required.
이에 본 발명자들은 천연물에서 유래되는 항균제를 찾기 위하여 우리나라에서 재배, 자생하고 있는 식물 중, 민간요법에서 약리작용이 구전되어온 식물들을 중심으로 하여 항생제 내성세균에 대한 항균활성물질을 검색하던 중 본 발명을 완성하게 되었다.The inventors of the present invention while searching for the antimicrobial active material against antibiotic resistant bacteria centered on plants that have been pharmacologically active in folk medicine among plants grown and grown in Korea in order to find antimicrobial agents derived from natural products. It was completed.
즉, 옛날부터 민간요법과 한방에서 종기, 화상, 부종, 위장장애, 늑막염 등의 치료에 사용되어 왔으며, 멕시코에서는 당뇨병치료에 사용된다고 알려져 있는 천연 자생식물인Opuntia속 선인장이 약제 내성세균에 항균활성을 나타냄을 알게 되었다.That is, in folk medicine and oriental medicine since ancient boils, burns, swelling, gastrointestinal disorders, have been used in the treatment of pleurisy, Mexico, the antibacterial activity in the natural wild plants of Opuntia genus cactus drugs that known to be used in the treatment of diabetes-resistant bacteria It was found that the.
특히, 해안부채선인장(Opuntia littoralis cock)과 손바닥 선인장(Opuntia ficus-indicavarSaboten Makino)에서 추출된 분획물이 메틸실린내성 황색포도상구균(MRSA)과 약제내성 세균 뿐만 아니라 일반세균에서도 강력한 항균활성을 나타내는 것을 발견하고 본 발명을 완성하기에 이르렀다.Particularly, fractions extracted from Opuntia littoralis cock and Opuntia ficus-indica var Saboten Makino showed strong antimicrobial activity in general bacteria as well as methylcillin -resistant Staphylococcus aureus (MRSA) and drug-resistant bacteria. It has been found that the present invention has been completed.
따라서 본 발명은 기존의 항균제와는 다른 기원을 가지는 천연식물로부터 유래되고 약제내성 세균 뿐만 아니라 일반적인 유해세균에 항균활성을 가지는 천연추출물을 제공하는데 그 목적이 있다.Accordingly, an object of the present invention is to provide a natural extract derived from a natural plant having a different origin from an existing antimicrobial agent and having antimicrobial activity against general harmful bacteria as well as drug resistant bacteria.
도 1- 본 발명에 의한 선인장 추출물질의 일 분획물질에 의한 페이퍼-디스크에 의한 항균활성 모습을 보인다.Figure 1- shows the antimicrobial activity of the paper-disk by a fraction of the cactus extract according to the present invention.
상기와 같은 목적을 달성하기 위한 본 발명의 특징에 따르면, 선인장으로부터 추출 분획된 항균활성 추출물질을 제공한다.According to a feature of the present invention for achieving the above object, it provides an antimicrobial active extract extracted from cactus.
그리고, 바람직 하기로는Opuntia속 선인장에서 분리 추출되도록 하며, 상기 추출물질은 산 가수분해에 의한 추출물로 이루어지도록 한다. 한편, 상기 추출물질은 실리카겔 크로마토그래피(전개용매, ethyl acetate : CH2Cl2: hexane = 1 : 1 : 1)를 이용하여 Rf가 다른 7개의 각 분획물질로 이루어지도록 하고, 상기 각 분획물질은 Rf값이 0.66-0.62, 0.45-0.40, 0.40-0.35, 0.35-0.30, 0.10-0.08, 0.07-0.05 그리고, 0.00인 범위의 것으로 이루어지도록 한다.And, preferably to be separated and extracted from Opuntia genus cactus, the extract is made of an extract by acid hydrolysis. On the other hand, the extract is made of silica gel chromatography (developing solvent, ethyl acetate: CH 2 Cl 2 : hexane = 1: 1: 1) to be composed of each of the seven fractions with different Rf, each fraction is Rf values are in the range of 0.66-0.62, 0.45-0.40, 0.40-0.35, 0.35-0.30, 0.10-0.08, 0.07-0.05 and 0.00.
이하 상기와 같은 특징을 가지는 본 발명을 좀 더 상세히 설명하기로 한다. 먼저, 실험을 위한 재료로서 항균활성물질의 추출을 위하여 손바닥 선인장을 항균물질 추출 원료로 사용하였다. 이는 원료의 공급면을 고려한 것으로서, 상기 손바닥선인장은 제주도 등에 자생하고 있는 다년생 식물일 뿐만 아니라 대량으로 재배되고 있기 때문에 원료수급면에서 최대의 장점을 가지고 있다. 그러나, 본 발명의 특징은 상기 손바닥 선인장에 한정되지 않으며,Opuntia속 선인장이면 어느 것이나 가능하다는 것이다. 한편, 본 발명은 약제 내성균의 감염치료를 위한 새로운 의료용 약재소재 개발뿐만 아니라 기능성화장품 개발 및 어류의 세균성질병 치료제등 여러 가지 세균성 질환에 대한 항균성을 가지는 생리활성물질을 제공하는데 가장 큰 특징이 있는 것이다.Hereinafter, the present invention having the above characteristics will be described in more detail. First, palm cactus was used as an antimicrobial material extraction material for the extraction of antimicrobial active material as a material for the experiment. This is considering the supply side of the raw material, the palm cactus is not only a perennial plant native to Jeju Island, but also has the greatest advantage in terms of supply and demand because it is grown in large quantities. However, the feature of the present invention is not limited to the above palm cactus, and any cactus of the genus Opuntia can be used. On the other hand, the present invention is the biggest feature to provide a biologically active substance having antimicrobial activity against various bacterial diseases such as development of functional cosmetics and treatment of bacterial diseases of fish as well as the development of new medical medicinal materials for the treatment of infection of drug-resistant bacteria. .
시험균주Test strain
이러한 다양한 세균에 대한 항균성을 알아보기 위하여 생리활성물질 검색단계에서부터 항균력 검사를 위한 시험균주를 광범위하게 선택하였다. 사용한 시험균주는 표준균주들 이외에 한국인 환자로부터 분리된 병원분리 약제내성균, 어병 병원체와 혐기성균인 여드름 유발원인세균과 충치유발 원인균 등을 시험균주로 사용하였으며, 본 발명의 실험을 위하여 사용된 균주는 표 1과 같다.In order to examine the antimicrobial activity against these various bacteria, a test strain for screening for antimicrobial activity was selected from the screen of bioactive substances. In addition to the standard strains, the isolates were used as test strains such as pathogen-resistant drug isolates from Korean patients, fish pathogens and anaerobes such as acne-causing bacteria and caries-causing bacteria. Same as 1.
시험방법Test Methods
Opuntia속 선인장의 항균활성물질에 대한 항균력 검사는 항생제 내성균 등의 시험균주가 접종된 한천 평판배양기에 선인장의 추출물과 분획물 등을 흡수시킨 페이퍼-디스크(paper-disc)를 놓고, 세균의 생육저해 활성을 관찰하는 페이퍼-디스크(paper-disc)방법을 이용하여 항균력을 표시하였다.The antimicrobial activity test on the antimicrobial activity of Opuntia genus cactus was carried out by placing a paper-disc that absorbed the extracts and fractions of the cactus in an agar plate incubator inoculated with the antibiotic-resistant bacteria. The antimicrobial activity was expressed using a paper-disc method.
시험균주의 배지 및 배양방법Test strain medium and culture method
Staphylococcus속과Pseudomonas속은 Tryptic soy broth,Lactococcus,Edwardsiella와Vibrio은 brain heart infusion,Streptococcus은 MRS broth배지를 각각 사용하여 생육적온에서 배양한 다음 한천 평판 배지에 접종하였다. 그러나 혐기성균인Propionibacterium은 GAM broth배지를 사용하여 다른 균주와는 달리 배양 및 항균시험에서 혐기상태를 유지해 주기 위해서 gas generating kit를 사용하였다. Staphylococcus genus and Pseudomonas genus were incubated at the growth temperature using Tryptic soy broth, Lactococcus , Edwardsiell a and Vibrio at brain heart infusion, and Streptococcus at MRS broth medium, respectively. However, unlike the other strains, propionibacterium, an anaerobic bacterium, used a gas generating kit to maintain anaerobic conditions in culture and antimicrobial tests.
재료의 선택Choice of materials
Opuntia속 선인장의 항균활성물질 존재 여부를 확인하는 예비실험은 먼저 해안부채 선인장과 손바닥선인장의 동결건조 분말시료를 50% MeOH으로 추출한 농축액 등을 사용하였다. 선인장 추출액은 표 1에 표시된Staphylococcus aureusKCTC 1621,Pseudomonas aeruginosaKCTC 1636, 병원분리균인 MRSA와 약제내성Pseudomonas aeruginosa, 혐기성균인Propionibacterium acnesATCC 3320을 시험균주로 선택해서 paper-disc법에 의하여 항균활성을 측정한 결과 충분한 항균활성을 나타냄을 알 수 있었으며, 두 선인장의 항균력 비교시험에서 두 선인장의 활성이 거의 동일하게 나타났다. 따라서 본 발명자들은 시료의 대량확보가 수월한 제주도 손바닥선인장을 시험균주에 의한 항균활성을 위한 주 시료로 선택하여 실험하였다.Preliminary experiments were conducted to determine the presence of antimicrobial activity in the cactus of Opuntia genus, using a concentrated extract of 50% MeOH from lyophilized powder samples of coastal lichen cactus and palm cactus. Cactus extracts were selected as Staphylococcus aureus KCTC 1621, Pseudomonas aeruginosa KCTC 1636, pathogen isolate MRSA and drug-resistant Pseudomonas aeruginosa , and anaerobic Propionibacterium acnes ATCC 3320 as test strains, and measured by the paper-disc method. As a result, it was found that it showed sufficient antimicrobial activity, and the activity of the two cacti was almost identical in the antibacterial activity test of the two cacti. Therefore, the present inventors experimented by selecting the Jeju cactus cactus easy to secure a large amount of the sample as the main sample for the antibacterial activity by the test strain.
이하에서는 손바닥선인장으로부터 항균활성물질의 추출과정 및 실험과정 그리고 그 결과를 설명하기로 한다.Hereinafter will be described the extraction process, the experimental process and the results of the antimicrobial active material from palm cactus.
추출예 1Extraction Example 1
산에 의한 가수분해Hydrolysis by acid
손바닥선인장의 동결건조 분말을 300g 준비하고, 이를 2N H2SO4(aq) 700ml용매 하에서 110℃를 유지하면서 약 10시간 동안 가수분해시켰다.300 g of lyophilized powder of palm cactus was prepared and hydrolyzed for about 10 hours while maintaining 110 ° C. under 700 ml of 2N H 2 SO 4 (aq) solvent.
반응이 완결된 후 실온에서 냉각시켜 2N NaOH(aq)로 중화시키고 중화된 수용액을 메틸렌클로라이드(CH2Cl2) 혹은 클로로포름(CHCl3)을 이용하여 추출한 후 Na2SO4로 건조시키고, 남아 있는 용매를 감압증류에 의해 제거시켜, 산 가수분해에 의한 추출물 약 3.0g을 얻었다.After completion of the reaction, the mixture was cooled to room temperature, neutralized with 2N NaOH (aq), and the neutralized aqueous solution was extracted with methylene chloride (CH 2 Cl 2 ) or chloroform (CHCl 3 ), dried over Na 2 SO 4 , and remaining. The solvent was removed by distillation under reduced pressure to obtain about 3.0 g of the extract by acid hydrolysis.
추출예 2Extraction Example 2
상기 추출예 1에서 얻어진 산 가수분해 생성물을 실리카겔 크로마토그래피(전개용매, ethyl acetate : CH2Cl2: hexane = 1 : 1 : 1)를 이용하여 Rf가 다른 7개의 분획물질을 얻었다. 본 실험에서는 산 가수분해 생성물 총 분획과 실리카겔 크로마토그래피를 이용하여 얻은 7개의 분획물질에 대한 항균활성을 조사하였다.The acid hydrolysis product obtained in Extraction Example 1 was subjected to silica gel chromatography (developing solvent, ethyl acetate: CH 2 Cl 2 : hexane = 1: 1: 1) to obtain 7 fractions having different Rf. In this experiment, the antimicrobial activity of the total fraction of acid hydrolysis products and the seven fractions obtained by silica gel chromatography were investigated.
추출예 3Extraction Example 3
염기에 의한 가수분해Hydrolysis by base
추출예 1과 유사한 방법으로 선인장 분말 300g을 2N NaOH(aq) 700ml용매 하에서 가수분해시켜 염기 가수분해 생성물 약 2.5g을 얻었다.In a similar manner to Extraction Example 1, 300 g of cactus powder was hydrolyzed under 700 ml of 2N NaOH (aq) solvent to obtain about 2.5 g of base hydrolysis product.
추출예 4Extraction Example 4
추출예 3에서 얻어진 염기 가수분해 생성물을 실리카겔 크로마토그래피(전개용매, ethyl acetate : CH2Cl2: hexane = 1 : 1 : 1)를 이용하여 산 가수분해물로 부터 얻은 7개의 분획물과 동일한 7개의 분획물질을 얻었다. 산 가수분해물과 유사하게 염기 가수분해 생성물과 실리카겔 크로마토그래피로 분리한 7개의 분획물들도 항균활성을 조사하였다.Seven fractions identical to the seven fractions obtained from the acid hydrolysates by silica gel chromatography (developing solvent, ethyl acetate: CH 2 Cl 2 : hexane = 1: 1: 1) obtained in the extraction example 3 Material was obtained. Similar to the acid hydrolyzate, the seven fractions separated by the base hydrolyzate and silica gel chromatography were also examined for antimicrobial activity.
실시예Example
상기 추출예 1 내지 추출예 4에서 얻어진 각 추출물들은 동일 조건을 유지하도록 동일한 농도(0.1g/150㎕)로 제조하고, 표 1의 각종 시험균주가 배양된 한천평판배양기상에 동일농도의 추출물들을 흡수시킨 페이퍼-디스크(paper-disc)를 놓고 배양한 다음, 생육저지환의 크기에 따른 항균활성 능력을 조사하였다. 그리고 항균활성 능력을 나타내는 생육저지환의 크기를 비교하기 위하여 대조군으로 클로로포롬을 사용하였다.Each extract obtained in Extraction Example 1 to Extractive Example 4 was prepared in the same concentration (0.1g / 150μl) to maintain the same conditions, and extracts of the same concentration on the agar plate incubator cultured various test strains of Table 1 After incubation with the absorbed paper-disc, the antimicrobial activity was investigated according to the size of growth-lowering ring. In addition, chloroform was used as a control to compare the size of growth-lowering ring showing antimicrobial activity.
실험결과Experiment result
(1)산 가수분해 생성물과 염기 가수분해 생성물의 항균력(1) Antibacterial activity of acid hydrolysis products and base hydrolysis products
표 2에 나타난 바와 같이, 선인장분말의 산 가수분해물이 물질 중량비로 비교할 때 염기 가수분해물보다 항균활성물질의 항균력이 보다 양호함을 알 수 있었다. 그리고 선인장 분말 300g으로부터 산 가수분해 생성물은 약 3.0g을 얻을 수 있었으며, 염기 가수분해 생성물은 약 2.5g을 얻을 수 있었으나, 산 가수분해방법이 항균활성물질을 추출하는 방법으로 더 좋은 것으로 나타났다.As shown in Table 2, it can be seen that the acid hydrolyzate of the cactus powder has a better antibacterial activity than the base hydrolyzate, compared to the base hydrolyzate. The acid hydrolysis product was about 3.0 g from the cactus powder 300 g and the base hydrolysis product was about 2.5 g, but the acid hydrolysis method was better as an antimicrobial active material extraction method.
(2) 산·염기 가수분해 생성물의 실리카겔 크로마토그래피(2) Silica Gel Chromatography of Acid-Base Hydrolysis Products
산·염기에 의한 가수분해 생성물의 실리카겔 크로마토그래피를 이용한 항균활성물질 분리에서 Rf값이 0.66-0.62인 MBT-01101, 0.45-0.40인 MBT-01102, 0.40-0.35인 MBT-01103, 0.35-0.30인 MBT-01104, 0.10-0.08인 MBT-01105, 0.07-0.05인 MBT-01106, 0.00인 MBT-01107이라는 7개의 분획물질을 얻었다.Separation of antimicrobial actives by hydrogel acid-base hydrolysis product using silica gel chromatography MBT-01101 with Rf value of 0.66-0.62, MBT-01102 with 0.45-0.40, MBT-01103 with 0.40-0.35, 0.35-0.30 Seven fractions were obtained: MBT-01104, MBT-01105 with 0.10-0.08, MBT-01106 with 0.07-0.05, and MBT-01107 with 0.00.
이러한 7개의 분획물질들에 대한 항균범위와 항균력을 표 3에 나타내었다. 상기 표에 나타난 바와 같이, 7개의 분획물질 중에서 항균범위가 넓고 항균력이 뛰어난 것은 MBT-01105와 MBT-01106임을 알 수 있었다. 그러나 모든 분획물질들이 특정 균주에 대한 활성은 나타내므로 각 분획물질들에 따른 사용 가능성을 충분히 알 수 있었다. 그리고, 이러한 분획물질들중 하나의 실시예로서 MBT-01105에 의한 페이퍼-디스크법에 의한 항균활성 모습을 도 1의 사진에 나타내었다. 상기 사진에서 보는 바와 같이 대조군인 클로로포롬과 비교되는 각 시험균주에 대한 항균활성을 나타내는 생육저지환의 크기를 뚜렷이 확인할 수 있었다. 그리고 상기 분획물질중 가장 다양한 균주에 항균활성을 나타내는 분획물인 MBT-01105의 분획수율은 산과 염기의 가수분해 생성물로부터 각각 300mg과 150mg을 얻을 수 있었으며, 또한 MBT-01106은 각각 50mg과 30mg을 얻을 수 있었다.Table 3 shows the antimicrobial range and antimicrobial activity of these seven fractions. As shown in the table, it was found that the broad antimicrobial range and excellent antimicrobial activity among the seven fractions are MBT-01105 and MBT-01106. However, all the fractions showed activity against a specific strain, so the possibility of use according to each fraction was sufficiently understood. In addition, as an example of one of these fractions, the antimicrobial activity of the paper-disk method by MBT-01105 is shown in the photograph of FIG. 1. As shown in the picture, the size of the growth hypolipidemia showing antimicrobial activity against each test strain compared to the control group chloroform was clearly identified. In addition, the fraction yield of MBT-01105, the fraction showing the antimicrobial activity against the most various strains of the fractions, was obtained from 300 mg and 150 mg, respectively, from the hydrolysis products of acid and base, and MBT-01106 was able to obtain 50 mg and 30 mg, respectively. there was.
그리고 부언하면, 표 3의 결과에서 보는 것처럼 MBT-01105는 MRSA와 약제내성Pseudomonas aeruginosa를 비롯하여 사용된 모든 균주에 대해 강한 항균활성을 나타내었다. 그리고 MBT-01106은 혐기성균인Propionibacterium acnes균주를 제외한 모든 시험균에 항균력을 나타내었으며 특히 병원분리 약제내성균에 대한 항균활성은 매우 강력하였으며 MBT-01105 보다 더 강하게 나타났다.In addition, as shown in the results of Table 3, MBT-01105 showed strong antimicrobial activity against all strains used, including MRSA and drug-resistant Pseudomonas aeruginosa . In addition, MBT-01106 showed antimicrobial activity against all test bacteria except Anaerobic Propionibacterium acnes strain, especially antimicrobial activity against pathogenic drug-resistant bacteria was stronger than MBT-01105.
* 각 분획물질의 농도는 0.1g/150㎕로 동일하며, paper-disc에 의한* The concentration of each fraction is the same as 0.1g / 150µl, by paper-disc
생육저지환의 크기는 mm로 나타내었다.The size of growth-lowering ring is expressed in mm.
그외의 분획물질인 MBT-01103과 MBT-01104도 어병 병원체에 대한 항균력을 나타내었으며 이들 물질도 어병 병원체에 대한 항균물질로 개발할 가치가 충분한 분획물질로 평가된다.The other fractions, MBT-01103 and MBT-01104, also showed antimicrobial activity against fish pathogens, and these substances are also considered to be valuable fractions to be developed as antimicrobials against fish pathogens.
이상에서 살펴 본 바와 같이 손바닥 선인장으로 부터 추출된 산·염기 가수분해 생성물과 가수분해 생성물로부터 분획된 물질인 MBT-01101, MBT-01102, MBT-01103, MBT-01104, MBT-01105, MBT-01106과 MBT-01107은 항균성 시험균주에 대해 특이적인 항균활성을 나타낸다는 것을 알 수 있다. 특히 MBT-01105와 MBT-01106은 병원성 분리약제 내성균에 대해 뛰어난 항균력을 나타내기 때문에 신규 항균활성물질의 약제개발에 그 유용성이 매우 클 것으로 기대 된다. 또한 MBT-01102, MBT-01103과 MBT-01104도 MRSA에 대해 항균활성을 가지고 있기 때문에 새로운 작용메커니즘을 가진 항균성 약제개발에 기여할 수 있는 물질로 평가될 수 있다.As described above, the acid-base hydrolysis products extracted from the palm cactus and the substances fractionated from the hydrolysis products MBT-01101, MBT-01102, MBT-01103, MBT-01104, MBT-01105, MBT-01106 It can be seen that MBT-01107 shows specific antimicrobial activity against the antimicrobial test strain. In particular, MBT-01105 and MBT-01106 are expected to be very useful for the development of new antimicrobial active agent drugs because they show excellent antimicrobial activity against pathogenic isolates resistant to bacteria. In addition, since MBT-01102, MBT-01103 and MBT-01104 have antimicrobial activity against MRSA, it can be evaluated as a material that can contribute to the development of antimicrobial drugs with a new mechanism of action.
이상에서 설명한 바와 같이, 본 발명에 따르면, 선인장에서 유래된 추출물이 약제내성세균 뿐만 아니라 일반세균에서도 강한 항균생리활성을 나타내므로 천연물로부터 유래된 새로운 항균제를 제공할 수 있을 뿐만 아니라 그 기원이 천연물이므로 인체에 전혀 무해하고 안전하게 사용할 수 있는 새로운 개념의 항균제를 제공하는 효과가 있다.As described above, according to the present invention, since the extract derived from the cactus exhibits strong antimicrobial physiological activity not only in drug-resistant bacteria but also in general bacteria, it is possible to provide a new antibacterial agent derived from natural products, since its origin is natural It is effective in providing a new concept of antimicrobial that can be used safely and harmless to the human body.
그리고 특히, 추출 분획물질 중 두가지 분획물질(MBT-01105 , MBT-01106)은 기존의 항생제에 내성을 나타내는 메티실린 내성 황색포도상구균(MRSA)과 약제내성녹농균(R-Pseudomonas aeruginosa)에 탁월한 항균작용을 나타내므로 현재 별다른 항균제가 없는 이러한 내성세균에 대한 새로운 대안이 될 수 있는 효과가 있다.In particular, two fractions (MBT-01105, MBT-01106) of the extract fractions have excellent antimicrobial activity against methicillin-resistant Staphylococcus aureus (MRSA) and drug-resistant Pseudomonas aeruginosa (R- Pseudomonas aeruginosa ), which are resistant to conventional antibiotics. Since there is no current antimicrobial agent there is an effect that can be a new alternative to these resistant bacteria.
또한 이외에 MBT-01102 , MBT-01103과 MBT-01104도 MRSA에 항균력을 가지고 있으며, 기타 각 분획물질들이 특이적으로 항균활성을 나타내는 균주가 존재함을 알 수 있다. 따라서 이러한 분획 추출물들을 이용하면 새로운 항균활성물질은 원내감염 다제 내성균의 치료제 개발에 획기적인 전환점을 마련할 것으로 본다.In addition, MBT-01102, MBT-01103 and MBT-01104 also have antimicrobial activity in MRSA, and it can be seen that there are strains that each of the other fractions exhibit antimicrobial activity. Therefore, using these fraction extracts, new antimicrobial actives are expected to provide a significant turning point in the development of therapeutic agents for multi-drug resistant bacteria in hospitals.
대부분의 병원 획득감염을 유발하는 황색포도상구균(S.aureus)은 국소피부감염에서부터 심내막염·골수염·패혈증·폐렴·수술부위 감염까지 다양한 감염질환을 일으키는 병원성 세균이다. 특히 최근 임상에서 포도상구균과 다른 그람양성세균의 최후 항생제로 여겨져 왔던 반코마이신(Vancomycin)내성균주가 발견되면서 약제내성균의 문제는 더욱더 심각해지고 있다. 또한Pseudomonas aeruginosa(녹농균)도 항생제 내성과 임상적 중요성에서 그람음성균중 현재 가장 문제가 되고 있는 균종이다. 녹농균 imipenem내성주의 증가는 치료상의 문제점을 증폭시키고 있다. 이와 같은 상황에서 민간요법에서 사용되어온Opuntia속 선인장으로부터 병원분리 약제 내성균에 강한 활성을 나타내는 새로운 생리활성물질을 찾았다는 실험결과는 인체에 안전하고 새로운 항균메카니즘을 가진 약제개발에 새로운 장을 열 것으로 기대된다.Most Staphylococcus aureus (S.aureus) that cause hospital acquired infections from a pathogenic bacteria that cause local skin infections, osteomyelitis, endocarditis, septicemia, pneumonia, various infections to surgical site infections. In particular, as vancomycin-resistant strains, which have been considered as the last antibiotics of staphylococci and other Gram-positive bacteria, have recently been discovered, the problem of drug-resistant bacteria becomes more serious. Pseudomonas aeruginosa (Pseudomonas aeruginosa) is also the most problematic species of Gram-negative bacteria in terms of antibiotic resistance and clinical significance. The increase in P. aeruginosa imipenem resistance is amplifying therapeutic problems. In this situation, a new bioactive substance showing strong activity against pathogen-resistant drug resistant bacteria from Opuntia cactus, which has been used in folk medicine, is expected to open a new chapter in the development of drugs that are safe for humans and have a new antibacterial mechanism. do.
그리고 손바닥 분획물질들의 항균범위와 항균력에서 볼 수 있는 바와 같이, 각 분획물질들에 대한 항균활성을 고려해서 약제를 개발한다면, 어류의 세균성 질병치료와 여드름 치료제 및 기능성 화장품개발 등 여러 가지 다른 용도로 사용할수 있는 또 다른 효과도 있다.And as can be seen in the antimicrobial range and antimicrobial activity of palm fractions, if the drug is developed in consideration of the antimicrobial activity of each fraction, it can be used for various other purposes such as the treatment of bacterial diseases of fish, treatment of acne and functional cosmetics. There is another effect you can use.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100906625B1 (en) * | 2008-03-18 | 2009-07-10 | (주) 메이지바이오 | Growth inhibitor of red or green algae |
| KR101011465B1 (en) * | 2008-06-13 | 2011-01-31 | (주)도우기술협력회 | Piping connection structure with backwash function |
| US11806378B1 (en) * | 2023-03-22 | 2023-11-07 | King Faisal University | Therapeutic composition including phenolic compounds derived from Opuntia littoralis |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100906625B1 (en) * | 2008-03-18 | 2009-07-10 | (주) 메이지바이오 | Growth inhibitor of red or green algae |
| KR101011465B1 (en) * | 2008-06-13 | 2011-01-31 | (주)도우기술협력회 | Piping connection structure with backwash function |
| US11806378B1 (en) * | 2023-03-22 | 2023-11-07 | King Faisal University | Therapeutic composition including phenolic compounds derived from Opuntia littoralis |
| US11975038B1 (en) * | 2023-03-22 | 2024-05-07 | King Faisal University | Therapeutic composition including phenolic compounds derived from opuntia littoralis |
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