紫外可见分光光度法UV-VIS.ppt

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prohibit research analysts from being compensated for their involvement in investment banking transactions.,csfb.cs-DfsRoot$IBDHKGProjectDAQOExecutionManagement Presentation092308_Pre-IPO Investor Mgmt Pres_V11_DTP.ppt,Confidential,*,紫外可见分光光度法,UV-VIS,1,紫外-可见分光光度法,1,紫外,-,可见分光光度法原理,2,紫外,-,可见分光光度计,3,分析条件的选择,4,紫外,-,可见分光分度法的应用,紫外-可见分光光度法的特点,与其它光谱分析方法相比，其仪器设备和操作都比较简单，费用少 分析速度快,灵敏度高,选择性好,精密度和准确度较高,用途广泛,1,紫外,可见分光光度法原理,1,紫外,可见分光光度法原理,一、物质对光的选择性吸收,物质对光的吸收是,选择性,的，利用被测物质对某波长的光的吸收来了解物质的特性，这就是,光谱法的基础,。,物质的颜色是因物质对不同波长的光具有选择性吸收作用而产生的，是物质所吸收光的互补色。,透射光和吸收光互为补色光,1,紫外,可见分光光度法原理,M +h,M*,基态 激发态,E1,（,E）E2,E=E2 -E1=h,1,紫外,可见分光光度法原理,紫外,-,可见分光光度法是利用物质的分子或离子对某一波长范围的光的吸收作用，对物质进行,定性分析、定量分析,及,结构分析。,按所吸收光的波长不同，分为紫外分光光度法和可见分光光度法。,二、,吸收曲线,通过测定被测物质对不同波长的光的吸收强度（吸光度），以波长为横坐标，吸光度为纵坐标作图，得出该物质在测定波长范围的吸收曲线。,1,紫外,可见分光光度法原理,1,紫外,可见分光光度法原理,吸收曲线的作用,1,、选择测定波长,2,、初步定性分析,3,、判断共存组分对待测组分有无干扰,1,紫外,可见分光光度法原理,1,、,透光度,T,描述入射光透过溶液的程度,T,=,I,t,/,I,0,吸光度,A,与透光度,T,的关系,A,lg,T,I,0,：,入射光强度；,I,t,：,透射光强度,三、光的吸收基本定律,朗伯,比耳定律,1,紫外,可见分光光度法原理,2,、朗伯-比尔定律,当一束,平行单色光,通过含有吸光物质的,稀溶液,时，溶液的吸光度与吸光物质浓度、液层厚度乘积成正比。,T:,透射率,e,:,摩尔吸光系数,b,:,光程长,C:,浓度,吸收,A =log,=,e,C,b1,T,3,、摩尔吸光系数,的讨论,1,紫外,可见分光光度法原理,吸收物质在一定波长和溶剂条件下的,特征常数,；,不随浓度,c,和光程长度,b,的改变而改变；,可作为定性鉴定的参数；,同一吸收物质在不同波长下的,值是不同的。在最大吸收波长,max,处的摩尔吸光系数，常以,max,表示。,max,表明了该吸收物质最大限度的吸光能力，也反映了光度法测定该物质可能达到的最大灵敏度。,10,4,强吸收，高灵敏,10,3,10,4,较强吸收,10,2,10,3,中吸收,四、偏离朗伯,比耳定律的原因,1,紫外,可见分光光度法原理,入射光为非单色光,溶液的不均性,光程的不一致性,2,紫外,可见分光光度计,可见分光光度计,2,紫外,可见分光光度计,紫外-可见分光光度计,一、基本结构,2,紫外,可见分光光度计,光源,单色器,吸收池,检测器,显示,样 品,2,紫外,可见分光光度计,1,、光源,作用：提供激发能,要求：可以发射连续光谱，具有足够的辐射强度、较好的稳定性、较长长的使用寿命。,类型,热辐射光源,用于可见光区，如钨灯和卤钨灯（,360-2500nm),气体放电光源,用于紫外光区，如氢灯和氘灯,(185-400nm),。,氘灯,紫外,卤钨灯,-,可见,2,、单色器,将光源发射的复合光分解成连续光谱并可从中选出任一波长单色光的光学系统。,包括狭缝、色散元件、透镜等,2,紫外,可见分光光度计,3,吸收池,用于盛放待测溶液和决定透光液层厚度的器件。,玻璃,:,400,760nm,石英,:200,760nm,要求：匹配性,2,紫外,可见分光光度计,4,、检测器,将光信号转变为电信号。,光电池,光电管,光电倍增管,二极管阵列检测器,2,紫外,可见分光光度计,5、记录装置,讯号处理和显示系统,2,紫外,可见分光光度计,记录装置,二、分光光度计类型,按其光学系统可分为,单波长分光光度计,和,双波长分光光度计,1、单波长单光束分光光度计,简单，价廉，适于在给定波长处测量吸光度或透光度，一般不能作全波段光谱扫描，要求光源和检测器具有很高的稳定性,。,2,紫外,可见分光光度计,2、单波长双光束分光光度计,自动记录，快速全波段扫描。可消除光源不稳定、检测器灵敏度变化等因素的影响，特别适合于结构分析。仪器复杂，价格较高。,3、双波长分光光度计,将不同波长的两束单色光(,1,、,2,),快速交替通过同一吸收池而后到达检测器。产生交流信号。无需参比池。,=,12,nm,。,两波长同时扫描即可获得导数光谱。,2,紫外,可见分光光度计,2,紫外,可见分光光度计,2,紫外,可见分光光度计,三、分光光度计的校验,当光度计使用一段时间后其波长和吸光度将出现漂移，因此需要对其进行校正,1,、,波长校正,2,、吸光度校正,3,、杂散光的检正,2,紫外,可见分光光度计,四、日常维护及保养,1,、对仪器工作环境的要求,稳固的工作台；,温度,15,28,；,湿度,45%,65%,，不超过,70%,；,防尘、防震、防电磁干扰；,放腐蚀。,2,紫外,可见分光光度计,2,、仪器保养和维护方法,仪器工作电源,220V,10%,，配稳压器；,光源使用注意事项,不使用时不要开光源灯,(,延长使用寿命,）；,单色器使用注意事项,不宜拆开，及时更换干燥剂；,正确使用吸收池,及时清洗，保护光学面，,1:1(3mol/LHCl:,乙醇,),混合液洗涤,一、显色反应条件的选择,选择显色反应时，应考虑的因素：,灵敏度高,选择性好,生成物稳定,显色剂在测定波长处无明显吸收,3,分析条件的选择,1、显色剂用量,吸光度,A,与显色剂用量,C,R,的关系会出现如图所示的几种情况。选择曲线变化平坦处。,3,分析条件的选择,2.反应体系的酸度,在相同实验条件下，分别测定不同,pH,值条件下显色溶液的吸光度。选择曲线中吸光度较大且恒定的平坦区所对应的,pH,范围。,3,、显色时间,4,、显色温度,5,、干扰的消除,3,分析条件的选择,二、仪器测量条件的选择,3,分析条件的选择,1、选择适当的入射波长,一般应该选择,max,为入射光波长。但如果,max,处有共存组分干扰时，则依据,“,吸收最大，干扰最小,”,原则确定的入射光波长。,3,分析条件的选择,2,、控制适宜的吸光度（读数范围）,不同的吸光度读数，产生的误差大小不同。,用仪器测定时应尽量使溶液透光度值在,T,%=2065(,吸光度,A,=0.700.20),，可求出浓度相对误差最小时的透光度,T,min,为：,T,min36.8%,A,min0.434,讨论,控制吸光度的方法,三、选择合适的参比溶液,溶剂参比,：当溶液中只有待测组分在测定波长处有吸收，其它组分无吸收，用纯溶剂（水)作参比溶液；,试剂参比：,显色剂或其它所加试剂在测定波长处有吸收，而待测溶液无吸收，用不加试样组分的其他试剂作参比溶液；,试样参比,：待测溶液在测定波长处有吸收，而显色剂等无吸收，用不加显色剂作参比溶液；,若显色剂、试液中其它组分在测量波长处有吸收，则可在试液中加入适当掩蔽剂将待测组分掩蔽后再加显色剂，作为参比溶液。,3,分析条件的选择,讨论,紫外可见分光光度法误差的主要来源。,3,分析条件的选择,33,紫外可见分光光度法是对物质进行定性分析，结构分析和定量分析的一种手段，而且还能测定某些化合物的物理常数，配合比和稳定常数，以及酸碱电离常数。,一、定性分析,制作试样的吸收曲线并与标准紫外光谱对照。,4,紫外可见分光光度法的应用,二、定量分析,1,、单组份定量方法,标准曲线法,配制不同浓度的标准溶液，由低浓度至高浓度依次测定其吸光度，做一定波长下浓度与吸光度的关系曲线，即标准曲线，通过标准曲线可求得未知样品的浓度。,标准对比法,该法是标准曲线法的简化，即只配制一个浓度为,cs的标准溶液，并测量其吸光度，求出吸收系数,k,，然后由,Ax=,kcx,求出,cx,该法只有在测定浓度范围内遵守,L-B,定律，且,cx,与,cs,大致相当时，才可得到准确结果,。,4,紫外可见分光光度法的应用,4,紫外可见分光光度法的应用,例 铁含量的测定,1,，,10,菲啰啉分光光度法,1,、标准溶液的配制,0.50mg/L,、,1.00mg/L,、,2.00mg/L,、,3.00mg/L,、,4.00mg/L,、,5.00mg/L,2,、吸光度测定,3,、绘制标准曲线,4,、相同条件下测定样品吸光度，在标准曲线中找出其浓度,4,紫外可见分光光度法的应用,2,、多组分定量方法,由于吸光度具有加合性，因此可以在同一试样中测定多个组份。,若各组分的吸收曲线互不重叠，则可在各自最大吸收波长处分别进行测定。这本质上与单组分测定没有区别,。,若各组分的吸收曲线互有重叠，则可根据吸光度的加合性求解联立方程组得出各组分的含量。,The End,