이 작품은 국립 자연 과학 재단의 중국 (No. 91543117)에 의해 재정적으로 지원 되었습니다.

주의: 사용 하기 전에 모든 관련 물질 안전 데이터 시트 (MSDS)를 참조 하십시오. 사용 하십시오 적절 한 개인 보호 장비, 예를 들면, 실험실 외 투, 장갑, 고글, 전체 길이 바지와 폐쇄 발가락 신발).
1. 초-nanoESI 소스를 설정
<ol><li>SESI 과정, 즉에 따르면 Sec nanoESI 소스를 설정 호흡 가스는 분무 깃털 교차 고 충전된 물방울 (그림 1)에 의해 이온화. 개별 실험실에 내장 된 소스 사용 하는 질량 분석기24,25인터페이스에 따라 다릅니다. 여기, 상용 nanoESI 소스 (그림 2)에 따라 Sec nanoESI 소스를 설정 하 고 벤치탑 4 중 극 orbitrap 질량 분석기를 구현 합니다. 참고: 소스의 본체는 입방 스테인레스 스틸 챔버 (길이 25 m m, 아이디 13 m m) (그림 2b) 챔버로 nanoESI 모 세관을 소개 하는 입구 (아이디 4 m m)와 함께. 따라서, 챔버 하지 않습니다 완전히 봉인 (그림 2b).</li>
	<li>가스 배달에 대 한 챔버의 각 측면에 두 개의 스테인레스 스틸 튜브 (길이 8 m m, 마약 5 m m, 아이디 3 m m)를 설치 합니다.</li>
	<li>NanoESI 모 세관의 눈 또는 디지털 현미경 nanoESI 스프레이 팁의 위치를 확인 하는 챔버의 위아래에 두 석 영 창 (아이디 14 m m)를 장비.</li>
	<li>질량 분석기의 스윕 콘 챔버 용접. 참고: 디자인 개별 실험실에서 사용 하는 질량 분석기의 대기압 인터페이스의 특정 형상에 따라 변경할 수 있습니다.</li>
</ol>2. 악기 최적화
<ol><li>제조업체의 지침에 따라 모두 긍정적이 고 부정적인 이온 검출 모드에서 질량 분 서 계를 보정. 보정을 적용 하 여 렌즈 잠재력 검색 조건과 같은 질량 분석기 매개 변수를 지정 된 해상도 값에 좋은 감도 및 피크 모양에 게 최적화 되어 있다. 70000의 대량 해상도 여기에 사용 됩니다.<ol>
<li>상업적인 ESI 소스;를 사용 하 여 완전 한 Q Exactive 교정 수행 그러나, 사용자 지정을 포함 하 여 모든 호환 소스와 질량 보정을 수행할 수 있습니다.</li>
	</ol>
</li>
	<li>이온 이동 관 (ITT) 질량 분석기 > 100 ° c의 온도 설정 350 ° C에서 최고 온도 설정할 수 있습니다, 하지만 그것은 몇 가지 화합물의 분해 될 수 있습니다. 따라서, 150 ° C이이 실험에 사용 됩니다. 참고: ITT 대신 샘플링 구멍을 특징으로 하는 질량 분석기에 대 한 샘플링 구멍의 온도 설정 > 100 ° c.</li>
	<li>ESI 용 매 및 흐름 속도 대 한 용 매 (예를 들어, 극성 및 변동)과 대상된 화합물 (예를 들어, 양성자 선호도)의 속성에 근거 하 여 적절 한 ESI 용 매를 선택 합니다. 물과 다양 한 비율에서 메탄올의 혼합물은 ESI 용 매25으로 일반적으로 사용 되었습니다. 이 실험에서는 물이 용 매13,,1923보고 위한 높은 이온화 효율은 0.1% (v/v) 포 름 산을 포함 하. ESI 용 매 유량 0-1.5의 범위에서 설정 μ/min와 200 nL/분. 참고: 드가 ESI 사용 하기 전에 30 분 용 매.</li>
	<li>초-nanoESI 소스 매개 변수, 주로 nanoESI 전압 및 nanoESI 모 세관 팁 위치를 최적화 합니다. 전압 일반적으로 범위는 2.0에서 4.5 kV. 여기 사용 2.5 kV입니다. 참고: ESI 전압 높은 흐름 속도 증가으로 적용 됩니다. 팁과 질량 분석기 구멍 사이의 거리는 5 m m 1에서 조정할 수 있습니다. 최적화, 후 질량 스펙트럼에서 관찰 된 정규화 된 강도 수준 (NL) > 1 x 10 해야6 및 총 이온 크로마 (TIC)의 유사 모두 긍정적이 고 부정적인 이온 검출 모드에서 < 10% 이어야 한다. 질량 스펙트럼 및 TIC m/z 50-750의 질량 범위에서 얻을 수 있습니다.</li>
	<li>가스 소스에 적용 됩니다. 이 실내 공기에서 VOCs의 영향을 줄이는 목표로 단계는 선택적입니다. 고 순도 질소 가스 (N2, 99.99%) 또는 순수한 공기를 사용할 수 있습니다. 순수 가스의 존재와 질량 스펙트럼에서 관찰 하는 NL > 1 x 10 해야5 와 TIC의 변화 모두 긍정적이 고 부정적인 이온 검출 모드에서 < 10% 이어야 한다. 고 순도 N2 는 여기에 사용 되 고 0.8 L/분 전달. 참고: 순수한 가스와 호흡 가스의 총 유량 질량 분석기의 구멍을 통해 유량 보다 높아야 한다.</li>
</ol>3입니다. exhaled 호흡의 측정
<ol><li>실내 공기를 흡입 하 고 일정 한 유량에서 폐의 모든 공기를 밖으로 호흡 하는 정상적인 증발 기를 수행 합니다. 증발 기 유량을 압력 계 또는 유량 주제를 볼 수를 모니터링 합니다. 테 프 론 (PTFE) 튜브를 사용 하 여 호흡 가스23제공.<ol>
<li>튜브 안에 수증기의 응축을 방지 하기 위해, 80-100 oC7,,2327 의 온도에 튜브가 열 하거나29Nafion 드라이어28,사용 합니다. 이 실험에서 주제 exhaled 0.4 L/분의 유량 계에 의해 제어.</li>
		<li>PTFE 튜브 (길이 13 ㎝, 아이디 4 m m)에 exhaled 호흡에서 수증기를 제거 하 고 흐름 미터의 콘센트를 연결 하는 Nafion 튜브 (길이 60 cm)를 유량의 입구를 연결 합니다. 그것은 하나의 증발 기 측정에 대 한 < 30 s를 걸립니다.</li>
		<li>4-6 복제 측정28,29를 수행 합니다.</li>
		<li>혼란 효과 최소화 하기 위해 참가자 들이 먹고, 마시고, 그리고 측정23전에 그들의 이빨 적어도 30 분을 솔 질에서 있다. 참고: 실내 공기에서 VOCs의 영향을 최소화 하기 위해 실내 공기26대신 가스를 흡입 보고 되었습니다 그것은. Nafion 튜빙을 사용 하는 경우 일부 극 지 화합물 손실 될 수 있습니다.</li>
		<li>측정, 동안 이온 강도 여부는 악기의 선형 검출 한계를 초과 하는 경우 확인 하십시오. 채도 신호의 샘플에서 화합물에서 실질적으로 발생 하지 않는 한 유물 피크 발생할 수 있습니다. 코를 통해 흡입, 주변 Voc 및 입자의 일부 제거 될 것 이다; 그러나, 그것은 주목할 만한 비 강에 화합물도 검색 될 수 있습니다.</li>
	</ol>
</li>
</ol>4. 구하는 호흡 지문 및 화합물의 시간 추적
<ol><li>Chromatograms 및 질량 스펙트럼을 얻을. 소프트웨어 (예: Xcalibur) 기록 chromatograms 및 질량 스펙트럼을 사용 합니다. 때문에 이것이 직접 MS 분석 크로마 분리를 수행, 총 이온 크로마 (TIC) 실제로 질량 스펙트럼에서 검색 된 모든 신호의 시간 추적을 나타냅니다 추출된 이온 크로마 (EIC) 표시의 시간 추적 한 지정 된 화합물입니다. 참고: 다른 상업 질량 분석기에 대 한 chromatograms 및 질량 스펙트럼 얻어질 수 있다 해당 데이터 수집 소프트웨어에 의해.</li>
	<li>Exhaled 호흡 지문 exhaled 호흡 측정 때 TIC의 다양 한 검사를 선택 하 여 얻을. 이러한 검사의 평균을 대표 하는 소프트웨어에 의해 질량 스펙트럼을 얻을.<ol>
<li>호흡 지문에서 배경 봉우리를 제거 하려면 배경 빼기 도구를 사용 하 여 소프트웨어에. 제조업체에서 제공한 설명서를 참조 하십시오. 간단 하 게, 아니 숨 샘플 도입 되어 검사의 동일한 수를 선택 하 고 빼기 호흡 지문에서 배경 질량 스펙트럼. 참고:이 방법에서는, 호흡 지문에 기능을 식별 하기 위해 임계값 세 번 배경 신호의 표준 편차로 정의 됩니다. 다른 상업 질량 분석기에 대 한 해당 데이터 수집 소프트웨어에 의해 배경 빼기를 수행할 수 있습니다.</li>
	</ol>
</li>
	<li>지정 된 화합물의 시간 추적을 가져옵니다. 호흡 지문에 대상된 화합물의 피크를 선택 하 고 화합물의 시간 추적 소프트웨어에 의해 연속적으로 취득 된다.</li>
</ol>

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Breath Res. 5 (1), 016002-016012 (2011).</li><li>Mart&#237;nez-Lozano Sinues, P., Zenobi, R., Kohler, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+the+exhalome+a+diagnostic+tool+of+the+future.">Analysis of the exhalome a diagnostic tool of the future.</a> Chest. 144 (3), 746-749 (2013).</li><li>Mart&#237;nez-Lozano Sinues, P., Fernandez de la Mora, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Direct+analysis+of+fatty+acid+vapors+in+breath+by+electrospray+ionization+and+atmospheric+pressure+Ionization-Mass+Spectrometry.">Direct analysis of fatty acid vapors in breath by electrospray ionization and atmospheric pressure Ionization-Mass Spectrometry.</a> Anal. Chem. 80 (21), 8210-8215 (2008).</li><li>Mart&#237;nez-Lozano Sinues, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Breath+analysis+in+real+time+by+mass+spectrometry+in+chronic+obstructive+pulmonary+disease.">Breath analysis in real time by mass spectrometry in chronic obstructive pulmonary disease.</a> Respiration. 87 (4), 301-310 (2014).</li><li>Mart&#237;nez-Lozano Sinues, P., Kohler, M., Zenobi, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monitoring+diurnal+changes+in+exhaled+human+breath.">Monitoring diurnal changes in exhaled human breath.</a> Anal. Chem. 85 (1), 369-373 (2013).</li><li>Chen, H. W., Zenobi, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neutral+desorption+sampling+of+biological+surfaces+for+rapid+chemical+characterization+by+extractive+electropray+ionization+mass+spectrometry.">Neutral desorption sampling of biological surfaces for rapid chemical characterization by extractive electropray ionization mass spectrometry.</a> Nat. Protoc. 3 (9), 1467-1475 (2008).</li><li>Li, X., Hu, B., Ding, J., Chen, H. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+characterization+of+complex+viscous+samples+at+molecular+levels+by+neutral+desorption+extractive+electrospray+ionization+mass+spectrometry.">Rapid characterization of complex viscous samples at molecular levels by neutral desorption extractive electrospray ionization mass spectrometry.</a> Nat. Protoc. 7 (6), 1010-1025 (2011).</li><li>Gordon, S. M., Szidon, J. P., Krotoszynski, B. K., Gibbons, R. D., O'Neill, H. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Volatile+organic+compounds+in+exhaled+air+from+patients+with+lung+cancer.">Volatile organic compounds in exhaled air from patients with lung cancer.</a> Clin. Chem. 31 (8), 1278-1282 (1985).</li><li>Ding, J. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+extractive+electrospray+ionization+ion+trap+mass+spectrometry+in+vivo+breath+analysis.">Development of extractive electrospray ionization ion trap mass spectrometry in vivo breath analysis.</a> Analyst. 134 (10), 2040-2050 (2009).</li><li>Basum, G., Dahnke, H., Halmer, D., Hering, P., M&#252;rtz, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Online+recording+of+ethane+trances+in+human+breath+via+infrared+laser+spectroscopy.">Online recording of ethane trances in human breath via infrared laser spectroscopy.</a> J. Appl. Physiol. 95 (6), 2583-2590 (2003).</li><li>T&#248;ien, &. #. 2. 1. 6. ;. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automated+open+flow+respirometry+in+continuous+and+long-term+measurements:+design+and+principles.">Automated open flow respirometry in continuous and long-term measurements: design and principles.</a> J. Appl. Physiol. 114 (8), 1094-1107 (2013).</li><li>Huang, L., Li, X., Xu, M., Huang, Z. X., Zhou, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+relatively+high+molecular+weight+compounds+in+human+breath+using+secondary+nano+electrospray+ionization+ultrahigh+resolution+mass+spectrometry.">Identification of relatively high molecular weight compounds in human breath using secondary nano electrospray ionization ultrahigh resolution mass spectrometry.</a> Chem. J. Chinese U. 38 (5), 752-757 (2017).</li></ol>

저자는 공개 없다.

Sec-nanoESI 소스 상용 nanoESI 원본을 기반 구축, 이온화 효율 ESI 소스30을 사용 하 여 보다 높은 수준 이다. 또한, 이온화 효율은 더욱 향상 닫힌된 챔버에 프로세스 분리로 주위 배경 공기에서와 같은 시간을 용이 하 게 가스 샘플 스프레이 깃털 사이 혼합. 초-nanoESI를 사용 하 여 덜 매개 변수는 ESI 소스, 쉽게 설치, 응용 프로그램 및 유지 보수에 비해 최적화가 필요 합니다.
<p class=...

현대 분석 기술의 빠른 개발, 휘발성 유기 화합물 (Voc)의 수백 인간의 exhaled 호흡1에서 확인 되었습니다. 이러한 Voc 치경 공기 (건강 한 성인에 대 한 ~ 350 mL)에서 주로 발생 하 고 해부학 죽은 공간 공기 (~ 150 mL)2, 신체 대사3,,45,6,7에 의해 영향을 받는 ,8 , 환경 오염9, 각각. 결과적으로, 식별,이 Voc 질병 진단과 환경 노출에 대 한 비-침략 적 방식으로 생체로 사용 될 것을 약속 하는.
가스 크로마토그래피 질량 분석 (GC-MS)은 가장 널리 사용 되는 기술 exhaled Voc2의 정성 및 정량 분석을 위해, 실시간 호흡 분석을 위해 개발 되었습니다, 직접 MS 기술을의 장점 높은 시간 해상도 및 간단한 샘플 사전 준비입니다. 직접 MS 기술, 양성자 이전 반응 MS (PTR-MS)10, 같은 선택 이온 흐름 관 석사 (선별-석사)11, 보조 분무 이온화 MS (SESI-MS)12,13 (또한 추출 물 분무로 라는 MS, MS EESI14,15이온화) 추적 대기 가스 분석기 (타가)16 및 플라즈마 이온화 MS (PI-MS)17 는 조사에서 최근 몇 년 동안.
모든 직접 MS 기술, 중 SESI 보편적인 부드러운 이온화 기술19,20,21;로 유명 하다 그리고 소스는 쉽게 사용자 지정 하 여 질량 분석기, 예를 들면, 시간의 비행 질량 분 서 계8,15의 다른 종류를 결합, 이온 트랩 질량 분석기14 그리고 orbitrap 질량 분석기12 ,18. 지금까지, SESI MS는 성공적으로 사용 되었다 호흡기 질환22, 진단 circadian 리듬3,,623, 약 동학7,8, 측정 및 대사 경로4, 등을 밝히고 있다. 가장 최근에, 상업 SESI 소스 사용할 수 있게 했다.
이 연구에서 손쉬운 소형 보조 nanoelectrospray 이온화 소스 (초-nanoESI) 설정 하 고 고해상도 질량 분석기를 결합 했다. 호흡에 exhaled Voc의 실시간 측정을 제시 했다.

<table><tbody><tr><td>Ultrapure water</td><td>Merck Millipore, USA</td><td>MPGP04001</td><td>Resistance &gt;18.2 M&amp;Omega;&amp;middot;cm</td></tr><tr><td>Formic acid</td><td>Sigma-Aldrich, USA</td><td>F0507</td><td>Corrosive to the respiratory tract.</td></tr><tr><td>Nitrogen gas</td><td>Guangzhou Shiyuan Gas Co. Ltd., China</td><td>N.A.&lt;sup&gt;a&lt;/sup&gt;</td><td>Purity &gt;99.99%</td></tr><tr><td>Q Exactive hybrid quadrupole-orbitrap mass spectrometer</td><td>Thermo Scientific, USA</td><td>02634L(S/N)</td><td>Beware of high voltage and high temperature</td></tr><tr><td>NanoESI source</td><td>Thermo Scientific, USA</td><td>ES002373(S/N); ES071(P/N)</td><td>Beware of high voltage and high temperature</td></tr><tr><td>Nano LC pump</td><td>Thermo Scientific, USA</td><td>5041.0010A(P/N)</td><td>/</td></tr><tr><td>Xcalibur software (Version 3.0)</td><td>Thermo Scientific, USA</td><td>BRE0008596</td><td>/</td></tr><tr><td>Dino-Lite Digital Microscope</td><td>Tech Video System (SuZhou) Co.Ltd., China</td><td>CQ401833R(S/N)</td><td>/</td></tr><tr><td>Nafion tubing</td><td>Perma Pure LLC, USA</td><td>ME60</td><td>/</td></tr><tr><td>PTFE tubing (I.D. 4 mm)</td><td>Dongguan Hongfu Insulating Material Co. Ltd., China</td><td>N.A.</td><td>Beware of the possible loss of polar compounds</td></tr><tr><td>Mass flow controller</td><td>Line-Tech, Korea</td><td>M15122007 (S/N)</td><td>/</td></tr><tr><td>Flow meter</td><td>Yuyao Industrial Automation Meter Factory, China</td><td>40784</td><td>/</td></tr><tr><td>&lt;sup&gt;a&lt;/sup&gt;N.A.: not available.</td><td></td><td></td><td></td></tr></tbody></table>

real-time breath analysis by using secondary nanoelectrospray ionization coupled to high resolution mass spectrometry

비-침략 적 방식으로 질병 진단과 환경 노출에 대 한 생체로 사용할 수 있습니다 이후 exhaled 휘발성 유기 화합물 (Voc) 상당한 관심을 자극 했습니다. 이 작품에서 선물이 고 분해능 질량 분석 (초-nanoESI-HRMS)에 결합 하는 보조 nanoelectrospray 이온화를 사용 하 여 실시간으로 exhaled Voc를 특성화 하는 프로토콜. 수 제 초-nanoESI 소스로 쉽게 세워졌다 기반 상용 nanoESI 소스에. 봉우리의 수백 exhaled 호흡의 배경 뺀 질량 스펙트럼에서 관찰 되었다 그리고 대량 정확도 값은-4.0-13.5 ppm와-20.3-긍정적이 고 부정적인 이온 검출 모드에서 1.3 ppm 각각. 봉우리는 정확한 질량 동위 원소 패턴에 따라 정확한 원소 구성으로 할당 되었다. 30 s 한 증발 기 측정에 사용 되 고 6 복제 된 측정에 대 한 약 7 분 소요.

그림 3 m/z 50-750 모두 긍정적이 고 부정적인 이온 검출 모드에서 기록의 대량 범위에 숨 지를 보여준다. 291 봉우리 (피크 강도 > 5.0x104) 173 봉우리 (피크 강도 > 3.0x104) 긍정적이 고 부정적인 이온 검출 모드에서 배경 빼고 숨 지문에 각각 관찰 되었습니다. 질량 스펙트럼에 봉우리를 식별 하려면 세부 정보12,</...

Watch this Scientific Journal Video about Real-time Breath Analysis by Using Secondary Nanoelectrospray Ionization Coupled to High Resolution Mass Spectrometry at JoVE.com

Real-time Breath Analysis by Using Secondary Nanoelectrospray Ionization Coupled to High Resolution Mass Spectrometry

고 분해능 질량 분석 시연 보조 nanoelectrospray 이온화를 사용 하 여 실시간으로 exhaled 호흡의 화학 성분 특성화에 대 한 프로토콜 결합.

고 분해능 질량 분석을 결합 하는 보조 Nanoelectrospray 이온화를 사용 하 여 실시간 호흡 분석

Exhaled volatile organic compounds (VOCs) have aroused considerable interest, since they can serve as biomarkers for disease diagnosis and environmental ...

real-time-breath-analysis-using-secondary-nanoelectrospray-ionization

Research

JoVE Journal

Chemistry

8.8K 조회수.  Jinan University. 고 분해능 질량 분석 시연 보조 nanoelectrospray 이온화를 사용 하 여 실시간으로 exhaled 호흡의 화학 성분 특성화에 대 한 프로토콜 결합.

비디오: Real-time Breath Analysis by Using Secondary Nanoelectrospray Ionization Coupled to High Resolution Mass Spectrometry

Untargeted Metabolomics from Biological Sources Using Ultraperformance Liquid Chromatography-High Resolution Mass Spectrometry (UPLC-HRMS)

Here we present a workflow to analyze the metabolic profiles for biological samples of interest including; cells, serum, or tissue. The sample is first separated into polar and non-polar fractions by a liquid-liquid phase extraction, and partially purified to facilitate downstream analysis. Both aqueous (polar metabolites) and organic (non-polar metabolites) phases of the initial extraction are processed to survey a broad range of metabolites. Metabolites are separated by different liquid chromatography methods based upon their partition properties. In this method, we present microflow ultra-performance (UP)LC methods, but the protocol is scalable to higher flows and lower pressures. Introduction into the mass spectrometer can be through either general or compound optimized source conditions. Detection of a broad range of ions is carried out in full scan mode in both positive and negative mode over a broad m/z range using high resolution on a recently calibrated instrument. Label-free differential analysis is carried out on bioinformatics platforms. Applications of this approach include metabolic pathway screening, biomarker discovery, and drug development.

Untargeted Metabolomics from Biological Sources Using Ultraperformance Liquid Chromatography-High Resolution Mass Spectrometry UPLC-HRMS

Untargeted metabolomics provides a hypothesis generating snapshot of a metabolic profile. This protocol will demonstrate the extraction and analysis of metabolites from cells, serum, or tissue. A range of metabolites are surveyed using liquid-liquid phase extraction, microflow ultraperformance liquid chromatography/high-resolution mass spectrometry (UPLC-HRMS) coupled to differential analysis software.

Ultraperformance 액체 크로마토 그래피 고해상도 질량 분석 (UPLC-HRMS)를 사용하여 생물 소스에서 타겟이 불분명 한 대사

Here we present a workflow to analyze the metabolic profiles for biological samples of interest including; cells, serum, or tissue. The sample is first ...

연구

화학

Characterizing Bacterial Volatiles using Secondary Electrospray Ionization Mass Spectrometry (SESI-MS)

Secondary electrospray ionization mass spectrometry (SESI-MS) is a method developed for the rapid detection of volatile compounds, without the need for sample pretreatment. The method was first described by Fenn and colleagues1 and has been applied to the detection of illicit drugs2 and explosives3-4, the characterization of skin volatiles5, and the analysis of breath6-7.
SESI ionization occurs by proton transfer reactions between the electrospray solution and the volatile analyte, and is therefore suitable for the analysis of hetero-organic molecules, just as in traditional electrospray ionization (ESI). However, unlike standard ESI, the proton transfer process of SESI occurs in the vapor phase rather than in solution (Fig. 1), and therefore SESI is best suited for detecting organic volatiles and aerosols.
We are expanding the use of SESI-MS to the detection of bacterial volatiles as a method for bacterial identification and characterization8. We have demonstrated that SESI-MS volatile fingerprinting, combined with a statistical analysis method, can be used to differentiate bacterial genera, species, and mixed cultures in a variety of growth media.8 Here we provide the steps for obtaining bacterial volatile fingerprints using SESI-MS, including the instrumental parameters that should be optimized to ensure robust bacterial identification and characterization.

Characterizing Bacterial Volatiles using Secondary Electrospray Ionization Mass Spectrometry SESI-MS

Secondary electrospray ionization mass spectrometry (SESI-MS) enables the detection of volatile organic compounds (VOCs) without the need for any sample pretreatment. This protocol provides instructions for the rapid (within minutes) characterization of bacterial VOCs using SESI-MS.

보조 전기 분무 이온화 질량 분석법 (SESI - MS)를 사용하여 특성화 세균 휘발성 물질

Secondary electrospray ionization mass spectrometry (SESI-MS) is a method developed for the rapid detection of volatile compounds, without the need for ...

생체공학

PTR-ToF-MS Coupled with an Automated Sampling System and Tailored Data Analysis for Food Studies: Bioprocess Monitoring, Screening and Nose-space Analysis

Proton Transfer Reaction (PTR), combined with a Time-of-Flight (ToF) Mass Spectrometer (MS) is an analytical approach based on chemical ionization that belongs to the Direct-Injection Mass Spectrometric (DIMS) technologies. These techniques allow the rapid determination of volatile organic compounds (VOCs), assuring high sensitivity and accuracy. In general, PTR-MS requires neither sample preparation nor sample destruction, allowing real time and non-invasive analysis of samples. PTR-MS are exploited in many fields, from environmental and atmospheric chemistry to medical and biological sciences. More recently, we developed a methodology based on coupling PTR-ToF-MS with an automated sampler and tailored data analysis tools, to increase the degree of automation and, consequently, to enhance the potential of the technique. This approach allowed us to monitor bioprocesses (e.g. enzymatic oxidation, alcoholic fermentation), to screen large sample sets (e.g. different origins, entire germoplasms) and to analyze several experimental modes (e.g. different concentrations of a given ingredient, different intensities of a specific technological parameter) in terms of VOC content. Here, we report the experimental protocols exemplifying different possible applications of our methodology: i.e. the detection of VOCs released during lactic acid fermentation of yogurt (on-line bioprocess monitoring), the monitoring of VOCs associated with different apple cultivars (large-scale screening), and the in vivo study of retronasal VOC release during coffee drinking (nosespace analysis).

Proton Transfer Reaction Time of Flight Mass Spectrometry allows high-sensitivity, rapid and non-invasive analysis of volatile organic compounds. To demonstrate its potential, we give three examples: lactic acid fermentation of yogurt (on-line bioprocess monitoring), different apple genotypes (large-scale screening), and retronasal space after drinking coffee (nosespace analysis).

자동 샘플링 시스템과 결합 된 PTR-ToF-MS 및 식품 연구를위한 맞춤형 데이터 분석 : 바이오 프로세스 모니터링, 스크리닝 및 코 공간 분석

Proton Transfer Reaction (PTR), combined with a Time-of-Flight (ToF) Mass Spectrometer (MS) is an analytical approach based on chemical ionization that ...

Leaf Spray Mass Spectrometry: A Rapid Ambient Ionization Technique to Directly Assess Metabolites from Plant Tissues

Plants produce thousands of small molecules that are diverse in their chemical properties. Mass spectrometry (MS) is a powerful technique for analyzing plant metabolites because it provides molecular weights with high sensitivity and specificity. Leaf spray MS is an ambient ionization technique where plant tissue is used for direct chemical analysis via electrospray, eliminating chromatography from the process. This approach to sampling metabolites allows for a wide range of chemical classes to be detected simultaneously from intact plant tissues, minimizing the amount of sample preparation needed. When used with a high-resolution, accurate mass MS, leaf spray MS facilitates the rapid detection of metabolites of interest. It is also possible to collect tandem mass fragmentation data with this technique to facilitate a compound identification. The combination of accurate mass measurements and fragmentation is beneficial in confirming compound identities. The leaf spray MS technique requires only minor modifications to a nanospray ionization source and is a useful tool to further expand the capabilities of a mass spectrometer. Here, fresh leaf tissue from Sceletium tortuosum (Aizoaceae), a traditional medicinal plant from South Africa, is analyzed; numerous mesembrine alkaloids are detected with leaf spray MS.

Leaf spray mass spectrometry is a direct chemical analysis technique that minimizes the sample preparation and eliminates chromatography, allowing for the rapid detection of small molecules from plant tissues.

잎 스프레이 질량 분석: 식물 조직에서 대사를 직접 평가 하는 급속 한 대기 이온화 기술

Plants produce thousands of small molecules that are diverse in their chemical properties. Mass spectrometry (MS) is a powerful technique for analyzing ...

생화학

Fizzy Extraction of Volatile Organic Compounds Combined with Atmospheric Pressure Chemical Ionization Quadrupole Mass Spectrometry

Chemical analysis of volatile and semivolatile compounds dissolved in liquid samples can be challenging. The dissolved components need to be brought to the gas phase, and efficiently transferred to a detection system. Fizzy extraction takes advantage of the effervescence phenomenon. First, a carrier gas (here, carbon dioxide) is dissolved in the sample by applying overpressure and stirring the sample. Second, the sample chamber is decompressed abruptly. Decompression leads to the formation of numerous carrier gas bubbles in the sample liquid. These bubbles assist the release of the dissolved analyte species from the liquid to the gas phase. The released analytes are immediately transferred to the atmospheric pressure chemical ionization interface of a triple quadrupole mass spectrometer. The ionizable analyte species give rise to mass spectrometric signals in the time domain. Because the release of the analyte species occurs over short periods of time (a few seconds), the temporal signals have high amplitudes and high signal-to-noise ratios. The amplitudes and areas of the temporal peaks can then be correlated with concentrations of the analytes in the liquid samples subjected to fizzy extraction, which enables quantitative analysis. The advantages of fizzy extraction include: simplicity, speed, and limited use of chemicals (solvents).

Fizzy extraction is a new laboratory technique for analysis of volatile and semivolatile compounds. A carrier gas is dissolved in the liquid sample by applying overpressure and stirring the sample. The sample chamber is then decompressed. The analyte species are liberated to the gas phase due to effervescence.

Chemical analysis of volatile and semivolatile compounds dissolved in liquid samples can be challenging. The dissolved components need to be brought to ...

Sample Preparation for Probe Electrospray Ionization Mass Spectrometry

Mass spectrometry (MS) is a powerful tool in analytical chemistry because it provides very accurate information about molecules, such as mass-to-charge ratios (m/z), which are useful to deduce molecular weights and structures. While it is essentially a destructive analytical method, recent advancements in the ambient ionization technique have enabled us to acquire data while leaving tissue in a relatively intact state in terms of integrity. Probe electrospray ionization (PESI) is a so-called direct method because it does not require complex and time-consuming pretreatment of samples. A fine needle serves as a sample picker, as well as an ionization emitter. Based on the very sharp and fine property of the probe tip, destruction of the samples is minimal, allowing us to acquire the real-time molecular information from living things in situ. Herein, we introduce three applications of PESI-MS technique that will be useful for biomedical research and development. One involves the application to solid tissue, which is the basic application of this technique for the medical diagnosis. As this technique requires only 10 mg of the sample, it may be very useful in the routine clinical settings. The second application is for in vitro medical diagnostics where human blood serum is measured. The ability to measure fluid samples is also valuable in various biological experiments where a sufficient volume of sample for conventional analytical techniques cannot be provided. The third application leans toward the direct application of probe needles in living animals, where we can obtain real-time dynamics of metabolites or drugs in specific organs. In each application, we can infer the molecules that have been detected by MS or use artificial intelligence to obtain a medical diagnosis.

This article introduces sample preparation methods for a unique real-time analytical method based on the ambient mass spectrometry. This method lets us perform real-time analysis of the biological molecules in vivo without any special pretreatments.

Mass spectrometry (MS) is a powerful tool in analytical chemistry because it provides very accurate information about molecules, such as mass-to-charge ...

Breath Collection from Children for Disease Biomarker Discovery

Breath collection and analysis can be used to discover volatile biomarkers in a number of infectious and non-infectious diseases, such as malaria, tuberculosis, lung cancer, and liver disease. This protocol describes a reproducible method for sampling breath in children and then stabilizing breath samples for further analysis with gas chromatography-mass spectrometry (GC-MS). The goal of this method is to establish a standardized protocol for the acquisition of breath samples for further chemical analysis, from children aged 4-15 years. First, breath is sampled using a cardboard mouthpiece attached to a 2-way valve, which is connected to a 3 L bag. Breath analytes are then transferred to a thermal desorption tube and stored at 4-5 &#176;C until analysis. This technique has been previously used to capture breath of children with malaria for successful breath biomarker identification. Subsequently, we have successfully applied this technique to additional pediatric cohorts. The advantage of this method is that it requires minimal cooperation on part of the patient (of particular value in pediatric populations), has a short collection period, does not require trained staff, and can be performed with portable equipment in resource-limited field settings.

This protocol describes a simple method for the acquisition of breath samples from children. Briefly, samples of mixed air are pre-concentrated in sorbent tubes prior to gas chromatography-mass spectrometry analysis. Breath biomarkers of infectious and non-infectious diseases can be identified using this breath collection method.

아이 들 질병 바이오 마커 발견에서 호흡 컬렉션

Breath collection and analysis can be used to discover volatile biomarkers in a number of infectious and non-infectious diseases, such as malaria, ...

Fast and Accurate Exhaled Breath Ammonia Measurement

This exhaled breath ammonia method uses a fast and highly sensitive spectroscopic method known as quartz enhanced photoacoustic spectroscopy (QEPAS) that uses a quantum cascade based laser. The monitor is coupled to a sampler that measures mouth pressure and carbon dioxide. The system is temperature controlled and specifically designed to address the reactivity of this compound. The sampler provides immediate feedback to the subject and the technician on the quality of the breath effort. Together with the quick response time of the monitor, this system is capable of accurately measuring exhaled breath ammonia representative of deep lung systemic levels. 
Because the system is easy to use and produces real time results, it has enabled experiments to identify factors that influence measurements. For example, mouth rinse and oral pH reproducibly and significantly affect results and therefore must be controlled. Temperature and mode of breathing are other examples. As our understanding of these factors evolves, error is reduced, and clinical studies become more meaningful. This system is very reliable and individual measurements are inexpensive. 
The sampler is relatively inexpensive and quite portable, but the monitor is neither. This limits options for some clinical studies and provides rational for future innovations.

Ammonia is an important physiologic metabolite relevant to various disease and wellness states. It is also a difficult molecule to measure in breath, which demands particular precautions be taken to obtain accurate results. Not all factors influencing ammonia are known, but progress can be difficult without accounting for these factors.

빠르고 정확한 날숨 암모니아 측정

This exhaled breath ammonia method uses a fast and highly sensitive spectroscopic method known as quartz enhanced photoacoustic spectroscopy (QEPAS) that ...

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Versatile Dual-Inlet Sample Introduction System for Multi-Mode Single Particle Inductively Coupled Plasma Mass Spectrometry Analysis and Validation

Metal-containing nanoparticles (NP) can be characterized with inductively coupled plasma mass spectrometers (ICP-MS) in terms of their size and number concentration by using the single-particle mode of the instrument (spICP-MS). The accuracy of measurement depends on the setup, operational conditions of the instrument and specific parameters that are set by the user. The transport efficiency of the ICP-MS is crucial for the quantification of the NP and usually requires a reference material with homogenous size distribution and a known particle number concentration.
Currently, NP reference materials are available for only a few metals and in limited sizes. If particles are characterized without a reference standard, the results of both size and particle number may be biased. Therefore, a dual-inlet setup for characterizing nanoparticles with spICP-MS was developed to overcome this problem. This setup is based on a conventional introduction system consisting of a pneumatic nebulizer (PN) for nanoparticle solutions and a microdroplet generator (&#181;DG) for ionic calibration solutions. A new and flexible interface was developed to facilitate the coupling of &#181;DG, PN and the ICP-MS system. The interface consists of available laboratory components and allows for the calibration, nanoparticle (NP) characterization and cleaning of the arrangement, while the ICP-MS instrument is still running.
Three independent analysis modes are available for determining particle size and number concentration. Each mode is based on a different calibration principle. While mode I (counting) and mode III (&#181;DG) are known from the literature, mode II (sensitivity), is used to determine the transport efficiency by inorganic ionic standard solutions only. It is independent of NP reference materials. The &#181;DG based inlet system described here guarantees superior analyte sensitivities and, therefore, lower detection limits (LOD). The size dependent LODs achieved are less than 15 nm for all NP (Au, Ag, CeO2) investigated.

Here we provide a protocol for the use of a dual-inlet system for single particle inductively coupled mass spectrometry which allows for a standard independent nanoparticle characterization.

Metal-containing nanoparticles (NP) can be characterized with inductively coupled plasma mass spectrometers (ICP-MS) in terms of their size and number ...

Atmospheric-pressure Molecular Imaging of Biological Tissues and Biofilms by LAESI Mass Spectrometry

Ambient ionization methods in mass spectrometry allow analytical investigations to be performed directly on a tissue or biofilm under native-like experimental conditions. Laser ablation electrospray ionization (LAESI) is one such development and is particularly well-suited for the investigation of water-containing specimens. LAESI utilizes a mid-infrared laser beam (2.94 &mu;m wavelength) to excite the water molecules of the sample. When the ablation fluence threshold is exceeded, the sample material is expelled in the form of particulate matter and these projectiles travel to tens of millimeters above the sample surface. In LAESI, this ablation plume is intercepted by highly charged droplets to capture a fraction of the ejected sample material and convert its chemical constituents into gas-phase ions. A mass spectrometer equipped with an atmospheric-pressure ion source interface is employed to analyze and record the composition of the released ions originating from the probed area (pixel) of the sample. A systematic interrogation over an array of pixels opens a way for molecular imaging in the microprobe analysis mode. A unique aspect of LAESI mass spectrometric imaging is depth profiling that, in combination with lateral imaging, enables three-dimensional (3D) molecular imaging. With current lateral and depth resolutions of ~100 &mu;m and ~40 &mu;m, respectively, LAESI mass spectrometric imaging helps to explore the molecular structure of biological tissues. Herein, we review the major elements of a LAESI system and provide guidelines for a successful imaging experiment.

Laser ablation electrospray ionization (LAESI) is an atmospheric-pressure ion source for mass spectrometry. In the imaging mode, a mid-infrared laser probes the distributions of molecules across a tissue section or a biofilm. This technique presents a new approach for diverse bioanalytical studies carried out under native experimental conditions.