WO2016003148A1 - 은이온에 특이적 형광 증대 현상을 보이는 디옥시리보 핵산 기반의 은나노클러스터와 이의 제조 방법 및 이에 따른 은이온 감응 센서 - Google Patents

은이온에 특이적 형광 증대 현상을 보이는 디옥시리보 핵산 기반의 은나노클러스터와 이의 제조 방법 및 이에 따른 은이온 감응 센서 Download PDF

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WO2016003148A1
WO2016003148A1 PCT/KR2015/006672 KR2015006672W WO2016003148A1 WO 2016003148 A1 WO2016003148 A1 WO 2016003148A1 KR 2015006672 W KR2015006672 W KR 2015006672W WO 2016003148 A1 WO2016003148 A1 WO 2016003148A1
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silver
deoxyribonucleic acid
nanocluster
dna
agncs
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PCT/KR2015/006672
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English (en)
French (fr)
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김원종
이지현
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기초과학연구원
포항공과대학교 산학협력단
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Publication of WO2016003148A1 publication Critical patent/WO2016003148A1/ko

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6813—Hybridisation assays
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence

Definitions

  • the present invention relates to a deoxyribonucleic acid-based silver nanocluster that exhibits fluorescence specific phenomena specific to silver ions. More specifically, the present invention relates to a silver nanocluster formed of deoxyribonucleic acid dimers with selective ions.
  • Nanoparticles generally refer to ultra-fine particles with a particle size of 100 ran or less. When the size of a substance is reduced to the molecular or atomic level, it is possible to take advantage of the new properties that are different from the physical and chemical properties of the raw materials. It is possible to develop new materials for various purposes.
  • Representative thick metal nanomaterials are copper, silver, aluminum, zinc, cadmium and are widely used in industrial and scientific fields.
  • ICP-MS Inductively Coupled Plasma Mass Spectrometry
  • AS inductively coupled plasma mass spectrometry
  • AAS atomic absorption spectroscopy
  • the present invention is directed to silver ions.
  • It aims to provide a deoxyribonucleic acid-based silver nanocluster, a method for producing the deoxyribonucleic acid dimer, and a corresponding silver ion desensitization sensor, which exhibit specific fluorescence enhancement by forming a deoxyribonucleic acid dimer.
  • the present invention relates to a deoxyribonucleic acid-based silver nanocluster (hereinafter referred to as DNA-AgNCs) with specific fluorescence amplification of silver ions.
  • DNA-AgNCs deoxyribonucleic acid-based silver nanocluster
  • Another aspect of the present invention is directed to DNA-AgNCs having 8 to 20 cytosines in the deoxyribonucleic acid.
  • Another aspect of the present invention is the above DNA-AgNCs, which are less than 3 ran.
  • Another aspect of the present invention is a DNA-AgNCs exhibiting fluorescence enhancement specific to silver ions, which comprises the steps of stirring deoxyribonucleic acid and silver nitrate and adding a reducing agent to the complex to produce a silver nanocluster. It is about the method of manufacturing the agenda.
  • Another aspect of the present invention relates to a method for producing DNA-AgNCs having a molar ratio of 1: 4 to 1: 8 in the combination of deoxyribonucleic acid and silver nitrate.
  • Another aspect of the present invention is the above-mentioned reducing agent, which is one to two times of silver nitrate.
  • Another aspect of the present invention is that silver ion sensor using DNA-AgNCs relates to ion sensor.
  • DNA-AgNCs are mixed with deoxyribonucleic acid and silver nitrate.
  • the deoxyribonucleic acid is a cytosine (C, cytosine) of 8 to 20 chains.
  • the DNA-AgNCs form dimers by mediating silver ions.
  • DNA-AgNCs The wavelength with maximum fluorescence emission intensity represented by DNA-AgNCs is 485 at 615 nm. ran into ran, which showed a weak red fluorescence before binding to silver ions.
  • DNA-AgNCs When combined with silver ions, DNA-AgNCs show strong green fluorescence.
  • This effect can be used as a silver iontophoretic sensor.
  • the DNA-AgNCs are particularly useful if silver nanoclusters are formed.
  • deoxyribonucleic acid and silver nitrate can be mixed in a molar ratio of 1: 4 to 1: 8, and more preferably, deoxyribonucleic acid and silver nitrate in a molar ratio of 1: 5 to 1: 7. It is preferred to form DNA-AgNCs best within this range.
  • DNA-AgNCs can be prepared by adding a reducing agent to the above-mentioned complex.
  • the reducing agent is capable of reducing the silver nitrate to form a silver nanocluster, and the silver nanocluster of the present invention is not particularly limited but is preferably added at a molar ratio of 1 to 2 times the silver nitrate.
  • the reducing agent is not particularly limited as long as it can reduce silver nitrate.
  • Sodium borohydride is preferably used.
  • the size of the DNA-AgNCs prepared above is not particularly limited, but is preferably less than 3 nm, more preferably 1 to 2 nm, and the maximum fluorescence emission intensity is determined according to the size of the DNA-AgNCs. If the amount of silver acid or reducing agent is higher than the composition ratio of the above manufacturing method, silver particles may be more condensed to form Ag-Nanopaticle, which is not silver nanocluster, where silver nanoparticles are higher than silver nanoclusters. A large size, which does not represent fluorescence, refers to a nano-assembly.
  • the DNA-AgNCs produced in the present invention can detect silver ions, can be detected within about 1 minute, and show stable detection even after 10 minutes. Effects of the Invention
  • DNA-AgNCs selectively deplete silver ions to deoxyribonate.
  • Nucleic acid dimers show specific fluorescence enhancement. These properties make it possible to utilize silver ion-sensitive sensors.
  • Figure 1 shows red-fluorescent DNA-AgNCs specific for silver ions.
  • FIG. 3 (A) shows a silver ion-added fluorescence spectrum of DNA-AgNCs.
  • Figure 5 (A) is a photograph taken before (-) after the silver ion addition of DNA-AgNCs
  • Figure 5 (B) is a graph showing its fluorescence spectrum.
  • Figure 6 (A) is a graph showing the fluorescence spectrum of the DNA-AgNCs according to the concentration of silver ions
  • Figure 6 (B) is a fluorescence test line graph according to Figure 6 (A)
  • y-axis is a fluorescence increase factor (I) and I at 485 nm, which means the fluorescence emission intensity of ( 10 ) and (I) before addition.
  • FIG. 8 is a graph showing fluorescence amplification factors over time and concentration, wherein 10 and I are as described above.
  • Example 1 the DNA-AgNCs according to the present invention, a method of manufacturing the same, and a silver ion desensitization sensor will be described in more detail, but the technical details of the present invention are not limited thereto.
  • Example 1 the DNA-AgNCs according to the present invention, a method of manufacturing the same, and a silver ion desensitization sensor will be described in more detail, but the technical details of the present invention are not limited thereto.
  • Deoxyribonucleic acid (C 12 -DNA) ( 12 Biioner, Korea) and silver nitrate, which were chained with 12 cytosines, were mixed and stirred in air-distilled distilled water * at a molar ratio of 1: 6, and stored at dark and room temperature for about 20 minutes.
  • the sodium borohydride was then mixed with silver nitrate in a molar ratio of 1: 1 and stored vigorously in dark and room temperature for one day. This resulted in the formation of red-fluorescent DNA-AgNCs.
  • a fluorophotometer (fluorophotometer X Shimadzu, Japan) was used.
  • a 400 ⁇ m-prepared DNA-AgNCs liquid sample was placed in a 1 cm quartz cell to confirm the emission intensity for each excitation wavelength.
  • the maximum fluorescence intensity was found to be the emission wavelength ( ⁇ ⁇ [ ⁇ ) of 615 nm at the excitation wavelength ( ⁇ « ) of about 575 nm, which indicates a weak red fluorescence. It was confirmed to indicate.
  • An absorbance photometer (UV-VIS spectrophotometer X, Shimadzu, Japan) was used to confirm the absorbance characteristics of the formed DNA-AgNCs. Ready to 10 ⁇
  • Liquid samples of DNA-AgNCs were placed in 1 cm quartz cells to measure absorption in the range of 200 ran to 800 nm. As can be seen in FIG. 4, a weak peak appeared around 575 nm related to the fluorescence of DNA-AgNCs.
  • the 0.2 ⁇ DNA-AgNCs liquid sample 200 prepared in Example 1 was mixed with 200 ⁇ silver ion solution with varying concentrations.
  • the red fluorescent DNA-AgNCs with the maximum emission intensity at 615 nm were maximum at 485 nra.
  • the emission intensity was transformed into super-fluorescent DNA-AgNCs, which allowed us to measure the emission intensity at 485 nm emission wavelength at 270 nm excitation wavelength and obtain a black line for the amount of silver ions. It was possible to qualitatively and quantitatively identify unknown samples containing no silver ions, and to determine whether other metals affected the 485 nm emission fluorescence of DNA-AgNCs.
  • each DNA-AgNCs was prepared to have a silver concentration of 50, 100, 250, 350, 500, 1000, 2000, and 3000 nM at 0.1 ⁇ M.
  • Each prepared sample was prepared to emit 485 nm using 270 nm excitation wavelength. The fluorescence emission intensity at was measured and represented by a black line as shown in FIG. 6.
  • Samples containing DNA-AgNCs and 1 ⁇ of silver ions were prepared.
  • the prepared solution was subjected to fluorescence emission intensity at the emission wavelength of 485 nm using 270 nm excitation wavelength.
  • the response time for detecting silver ions in DNA-AgNCs was determined. 0.1 ⁇ M of DNA-AgNCs were prepared for samples of 50, 100, 250, 500, and 1000 ⁇ , respectively. 1, 2, 3, 4, 5, 7.5, 10 minutes), the fluorescence emission intensity at 485 nm was measured. As shown in FIG. 8, the silver ions were detected by DNA-AgNCs within about 1 minute and showed stable detection ability even after 10 minutes.

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Abstract

본 발명은 은이온에 특이적 형광 증대 현상을 보이는 디옥시리보 핵산 기반의 은나노클러스터에 관한 것으로, 보다 자세하게는 사이토신 8 ~ 20개를 사슬로 가지는 디옥시리보 핵산을 이용한 은나노클러스터의 제조 방법을 제공한다. 또한 은이온에 특이적 형광 증대 현상을 보이는 성질을 이용한 은이온 감응 센서를 제공함으로써, 환경 및 의학 분야에 유용하게 활용될 것이다.

Description

명세서
발명의명칭:은이온에특이적형광증대현상을보이는 디옥시리보핵산기반의은나노클러스터와이의제조방법및이에 따른은이온감웅센서
기술분야
[1] 본발명은은이온에특이적형광증대현상을보이는디옥시리보핵산기반의 은나노클러스터에관한것으로，보다자세하게는은이온과선택적으로 감웅하여디옥시리보핵산이합체를이루는은나노클러스터에관한것이다.
[2] 또한，특정한서열을가지는디옥시리보핵산과질산은을이용하여
은나노클러스터를제조하는방법과은이은에특이적형광증대현상을보이는 성질을이용한은이온감웅센서에관한것이다.
배경기술
[3] 일반적으로나노입자는입경 100 ran이하의초미세입자 (Ultrafme particle)를 말한다.어떤물질의크기가분자또는원자수준으로작아지면원재료의 물리화학적성질과는전혀다른새로운성질이나타나는데이를이용하여 다양한용도의신소재를개발할수있게되었다.
[4] 그러나나노물질의새롭고다양한특성은본질적으로지금까지알려지지않은 새로운생체반웅을유발할수도있다는것을의미하며따라서나노물질의 인체독성과환경에미치는잠재적위해성이새로운이슈로등장하게되었다.
[5] 대표적인증금속나노물질은구리 (Copper),은 (Silver),알루미늄 (Aluminum), 아연 (Zinc),카드뮴 (Cadmium)둥으로써산업분야나과학분야에서광범위하게 사용되고있다.
[6] 최근은나노입자에대한산업적웅용이증가하고있는데은은예로부터사진， 보석，배터리，전기제품등에사용되어왔으며,의류，정수기,세탁기，치약등 인체에직접적인영향을미칠수있는분야에많이사용되어왔다.특히，이미 오래전부터항균력이인정되어의약품으로사용되어왔으며현재에도 화상치료제성분으로광범위하게사용되고있다.
[7] 의약품으로서의질산은 (AgN03)에대한독성은이미오래전부터연구된바 있으며피부도포제로서의은제제는상대적으로독성이높지않은것으로 평가되어왔다.그러나최근입자형태의은나노물질의산업적용도가
증가하면서은나노물질의독성연구도매우심도있게연구되고있다.
[8] 은이은은박테리아，바이러스，곰광이등에고도로독성이며，인간에게도
은이온의과량축적은은피증 (argyria)의원인이된다.또한환경에도악영향을 미치는것으로보고되고있다.이러한이유로은이온에대한높은감도，높은 선택성및빠른속도의검출성능이필요하다.
[9] 기존은이온의검출방법으로유도결합플라즈마질량분석법 (ICP-MS, inductively coupled plasma mass spectrometry)및원자톱수분광법 (AAS, atomic absorption spectroscopy)을이용한방법이개발되었지만，이들은고도로정교한 장비와복잡한제조공정을필요로한다.
[10] 또한다양한유기화합물，양자점 (quantum dots),나노입자，고분자및 DNA가 화학센서로사용되어왔다ᅳ이러한방법은은이온의검출에있어고감도를 발휘하지만，시약합성의어려움，복잡한검출시스템및제한된지속시간등의 단점이있다.
발명의상세한설명
기술적과제
[11] 상기와같은문제점을해결하기위하여본발명은은이온에선택적으로
감웅하여디옥시리보핵산이합체를이룸으로써특이적형광증대현상을 보이는，디옥시리보핵산기반의은나노클러스터와이의제조방법및이에따른 은이온감웅센서를제공하는것을목적으로한다.
과제해결수단
[12] 본발명은은이온에특이적형광증대현상을보이는디옥시리보핵산기반의 은나노클러스터 (이하， DNA-AgNCs로표기)에관한것이다.
[13] 또한본발명의또다른양태는상기디옥시리보핵산에있어,사이토신 8 ~ 20개를가지는 DNA-AgNCs에관한것이다.
[14] 또한본발명의또다른양태는상기 DNA-AgNCs에있어, 3 ran이하인
DNA-AgNCs에관한것이다.
[15] 또한본발명의또다른양태는디옥시리보핵산과질산은을교반하는단계및 상기흔합물에환원제를첨가하여은나노클러스터를제조하는단계를포함하는 은이온에특이적형광증대현상을보이는 DNA-AgNCs의제조방법에관한 것이다.
[16] 또한본발명의또다른양태는상기디옥시리보핵산과질산은의흔합물에 있어, 1:4ᅳ 1:8의몰비율을가지는 DNA-AgNCs의제조방법에관한것이다.
[17] 또한본발명의또다른양태는상기환원제에있어，질산은의 1 ~ 2배의
몰비율로첨가되는 DNA-AgNCs의제조방법에관한것이다.
[18] 또한본발명의또다른양태는 DNA-AgNCs를이용한은이온감웅센서은이온 감웅센서에관한것이다ᅳ
[19]
[20] 이하，본발명에따른각성분에대해설명한다.
[21] 본발명에따른 DNA-AgNCs는디옥시리보핵산과질산은올섞은후이를
환원시켜형성된것으로，상기디옥시리보핵산은사이토신 (C, cytosine) 8 ~ 20개의사슬로된것이다.
[22] 상기 DNA-AgNCs는은이온을매개로하여이합체를형성한다.이로인해
DNA-AgNCs가나타내는최대형광방출세기를가지는파장이 615 nm에서 485 ran로변화하게되는데，은이온과결합전엔붉은색의약한형광을나타내던
DNA-AgNCs가은이온과결합후엔초록색의강한형광을나타내게된다.
이러한효과를이용하여은이온선 적인감웅센서로이용할수있다.
[23] 본발명에서상기 DNA-AgNCs은은나노클러스터가형성된다면특별히
제한되진않으나예를들면，디옥시리보핵산과질산은을 1:4 ~ 1:8의몰비율로 섞어흔합물을제조할수있으며，더욱좋게는디옥시리보핵산과질산은을 1:5 ~ 1:7의몰비율로섞는것이바람직하다.이범위내에서 DNA-AgNCs를가장잘 형성한다.
[24] 본발명에서상기흔합물에환원제를첨가하여 DNA-AgNCs를제조할수있다. 상기환원제는질산은을환원시켜은나노클러스터를형성할수있도록하는 것으로써，본발명의은나노클러스터가생성된다면특별히제한되진않으나 예를들면，질산은의 1 ~ 2배의몰비율로첨가하는것이바람직하다.
[25] 상기환원제는질산은을환원시킬수있는것이라면특별히제한되지는
않으나，예를들어수소화붕소나트륨 (sodium borohydride),하이드라진 (hydrazine) 또는하이드로퀴논 (hydroquinone)등을사용할수있으며,이중
수소화붕소나트륨을사용하는것이바 직하다.
[26] 상기제조된 DNA-AgNCs의크기는특별히제한되진않으나， 3 nm이하，더욱 좋게는 1 ~ 2 nm가바람직하며, DNA-AgNCs의크기에따라최대형광방출 세기가결정된다.상기제조방법에서디옥시리보핵산의비율에비해질산은 또는환원제의양이상기제조방법의조성비율보다높으면，은입자가더욱잘 웅집되어은나노클러스터가아닌은나노파티클 (Ag-Nanopaticle)이형성될수 있다.여기서은나노파티클은은나노클러스터보다크기가큰，형광을나타내지 않는은나노집합체를말한다.
[27] 본발명에서제조된 DNA-AgNCs는은이온을검출하는데있어，약 1분이내에 검출이완료될수있으며， 10분전후까지도안정한검출능력을보여줄수있다. 발명의효과
[28] 본발명에따른 DNA-AgNCs는은이온에선택적으로감웅하여디옥시리보
핵산이합체를이룸으로써특이적형광증대현상을보인다.이러한특성을 이용하여은이온감응센서로활용이가능하다.
도면의간단한설명
[29] 도 1은붉은형광을내는 DNA-AgNCs가은이온과특이적감웅하여초록
형광을내는것을도식화한것이다.
[30] 도 2는 DNA-AgNCs의형태를확인하기위한전자현미경이미지이다.
[31] 도 3의 (A)는 DNA-AgNCs의은이온첨가전형광스펙트럼을나타낸
그래프이고，도 3의 (B)는은이온첨가후형광스펙트럼을나타낸그래프이다.
[32] 도 4는 DNA-AgNCs의흡광스펙트럼을나타낸그래프이다.
[33] 도 5의 (A)는 DNA-AgNCs의은이온첨가전 (-)후 (+)를찍은사진이고，도 5의 (B)는이의형광스펙트럼을나타낸그래프이다.
도 6의 (A)는은이온의농도에따른 DNA-AgNCs의형광스펙트럼을나타낸 그래프이고，도 6의 (B)는도 6의 (A)에따른형광검정선그래프로, y축은 형광증대인자 (fluoroscene enhancement factor)이며， I。및 I는 485 nm에서은이은 첨가전 (10)및후 (I)의형광방출세기를의미한다.
도 7은은이온및기타금속이온에대한선택성을나타낸그래프로， 10및1는 상기와같다.
도 8은시간및농도에따른형광증대인자를나타낸그래프로， 10및 I는상기와 같다.
발명의실시를위한형태
이하,본발명에따른 DNA-AgNCs와이의제조방법및은이온감웅센서에 대해더욱상세하게설명하겠으나，본발명의기술적사항이이에한정되는것은 아니다. [실시예 1]
사이토신 12개를사슬로가지는디옥시리보핵산 (C12-DNA)(한국， Bioneer사)과 질산은을 1:6의몰비율로공기가포화된증류수 *에섞어교반하여약 20분간 암실및실온에보관하였다.이후수소화붕소나트륨을질산은과 1:1의몰비율로 섞어격렬히교반한후암실및실온에서하루동안보관하였다.이과정을 통하여은이은이붉은형광을가지는 DNA-AgNCs를형성하였다.
*공기가포화된증류수의제조:신선한증류수약 10 ^를교반자와함께 20 유리병에넣고교반기를이용해하루동안교반하고，이를 20 皿주사기 필터 (미국 , Millipore사)로여과하였다. 형성된 DNA-AgNCs의형태를투과전자현미경 (transmission electron microscope)(일본， JOEL사)으로분석하였다. 5 μΜ로준비된 DNA-AgNCs액체 시료 5 ^를 400메쉬 (mesh)카본그리드 (미국， Ted Pella사)에떨어뜨리고말려 측정에사용하였다. [도 2]에서확인할수있듯이，약 l ~ 2 nm의크기를가지는 DNA-AgNCs가형성되었음을확인하였다. 형성된 DNA-AgNCs의형광특성을확인하기위하여
형광광도계 (fluorophotometerX일본， Shimadzu사)를사용하였다. 10 μΜ로준비된 DNA-AgNCs액체시료 400 를 1 cm석영셀 (quartz cell)에넣어각들뜸파장에 대한방출세기를확인하였다. [도 3의 (A)]에서확인할수있듯이，최대형광 세기는약 575 nm의들뜸파장 (λ«)에서 615 nm의방출파장 (λε[η)으로확인되었고， 이는붉은색의약한형광을나타내는것으로확인되었다. [47] 형성된 DNA-AgNCs의흡광특성을확인하기위하여흡광광도계 (UV- VIS spectrophotometerX일본， Shimadzu사)를사용하였다. 10 μΜ로준비된
DNA-AgNCs의액체시료 200 ^를 1 cm석영셀에넣어 200 ran ~ 800 nm 범위에서의흡광을측정하였다. [도 4]에서확인할수있듯이， DNA-AgNCs의 형광과관련된 575 nm부근에서약한피크 (peak)가나타남을확인할수있었다.
[48]
[49] [실시예 2]
[50] (가) DNA-AgNCs을이용하정량적이고서택적？ᅵ은이온의검출능^ 확이
[51] 실시예 1에서준비한 0.2 μΜ DNA-AgNCs액체시료 200 를다양한농도를 가지는은이온수용액 200 ^와섞었다.이때 615 nm에서최대방출세기를 가지던붉은형광의 DNA-AgNCs는 485 nra에서최대방출세기를가지는초톡 형광의 DNA-AgNCs로변하였다.이를이용하여 270 nm들뜸파장일때 485 nm 방출파장에서의방출세기를측정할수있고，은이온의양에대한검정선을 얻을수있었다.얻은검정선을이용하여은이온을포함한흑은포함하지않은 미지의시료를정성적，정량적으로구별할수있었다.또한다른금속이은들의 경우 DNA-AgNCs의 485 nm방출형광에영향을미치는지를확인하였다.
[52]
[53] (a)은이온의첨가에따른형광변화확인
[54] 은이온을 DNA-AgNCs에첨가하면최대들뜸 /방출파장이 575 nm/615 nm에서 270 nra/485 ntn로변함을확인하였다.이는 [도 3의 (A)]에서나타난것처럼 붉은색의약한형광을나타내던기존 DNA-AgNCs에서， [도 3의 (B)]에서나타난 것처럼초록색의강한형광을나타내는 DNA-AgNCs로변함을알수있었다.
[55] (b)은이온에대한 DNA-AgNCs의검정선구하기
[56] 0.2 μΜ의 DNA-AgNCs 200 ^와다양한농도의은이온용액을섞어서
최종적으로각 DNA-AgNCs 0.1 μΜ에 50， 100, 250, 350, 500, 1000, 2000, 3000 nM의은이은농도를가지도록준비하였다.준비된각시료를 270 nm들뜸파장을 이용하여 485 nm의방출파장에서의형광방출세기를측정하고 [도 6]과같이 검정선으로표시하였다.
[57] (c)기타금속이온에대한 DNA-AgNCs의선택성확인
[58] 은이온에대한선텍적인 DNA-AgNCs의형광증대특성을확인하고자기타 금속이온에대해서마찬가지실험을진행하였다.최종적으로각 0.1 μΜ의 DNA-AgNCs와 ΙΟ μΜ의알루미늄이온，카드뮴이은，크롬이온，구리이은,철이온, 칼륨이은，리튬이온，마그네슘이은，소듐이은，니켈이온，아연이은그리고 수은이은을포함한시료를각각준비하였다.마찬가지로 0.1 μΜ의
DNA-AgNCs와 1 μΜ의은이온을포함하는시료를준비하였다.준비한용액을 270 nm들뜸파장을이용해 485 nm의방출파장에서의형광방출세기를
측정하였다. [도기에서나타난것처럼,기타금속이온의농도에비해 1/10의 농도를가지는은이온의경우，선텍적으로증대됨을확인할수있었다.이는 은이온의검출에매우유용하다.
[59] (d)은이온의검출웅답시간확인
[60] DNA-AgNCs에서은이온이검출되는웅답시간을확인하였다.이를위하여 DNA-AgNCs 0.1 μΜ은이은이각 50， 100, 250, 500, 1000 ηΜ인시료를각각 제조하고,은이온과반웅한시간 (1, 2， 3， 4， 5， 7.5， 10분)에따라 485 nm에서의형광 방출세기를측정하였다. [도 8]에나타난것처럼，은이온은 DNA-AgNCs에 의하여약 1분이내에검출이완료되며 10분전후까지도안정한검출능력을 보여줌을확인할수있었다.

Claims

청구범위
[청구항 1] 은이온에특이적형광증대현상을보이는디옥시리보핵산
기반의은나노클러스터.
[청구항 2] 제 1항에있어서，
상기디옥시리보핵산은사이토신이 8 ~ 20개를가지는 디옥시리보핵산기반의은나노클러스터 .
[청구항 3] 제 1항에있어서，
상기디옥시리보핵산기반의은나노클러스터는 3 nm이하인 디옥시리보핵산기반의은나노클러스터.
[청구항 4] 디옥시리보핵산과질산은을교반하는단계；및
상기흔합물에환원제를첨가하여은나노클러스터를제조하는 단계;
를포함하는은이온에특이적형광증대현상을보이는 디옥시리보핵산기반의은나노클러스터의제조방법 .
[청구항 5] 제 4항에있어서，
상기디옥시리보핵산과질산은의흔합물은 1:4ᅳ 1:8의몰비율을 가지는디옥시리보핵산기반의은나노클러스터의제조방법 .
[청구항 6] 제 4항에있어서，
상기환원제는질산은의 1 ~ 2배의몰비율로첨가되는디옥시리보 핵산기반의은나노클러스터의제조방법.
[청구항 7] 제 1항내지 3항에있어서선택되는어느한항에있어서，
디옥시리보핵산기반의은나노클러스터를이용한은이온감웅 센서.
PCT/KR2015/006672 2014-07-02 2015-06-30 은이온에 특이적 형광 증대 현상을 보이는 디옥시리보 핵산 기반의 은나노클러스터와 이의 제조 방법 및 이에 따른 은이온 감응 센서 WO2016003148A1 (ko)

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