仪器确认附件1HPLC确认中英文doc

PA/PH/OMCL (11) 04 - OMCL Guideline on qualification of HPLC equipment (Annex 1)

ANNEX I 附件1

Level III. Periodic and motivated instrument checks

第三级 定期性的主动仪器检查

This Annex contains practical examples of tests and their associated tolerance Limits for several parameters related to the performance of the different modules of a HPLC. 本附件包括一些实用几个与HPLC不同模块相关的参数检测举例及其相应的可接受限度。

These examples can be considered by the OMCLs as possible approaches to perform the Level III of the equipment qualification process: ―Periodic and motivated instrument checks‖. 这些例子可以认为是OMCL实验室可能的进行第三级仪器确认的方法―周期主动仪器检查‖

HPLC SOLVENT DELIVERY SYSTEM 高压液相溶剂传输系统

The following tests are proposed for the periodic and motivated check of the HPLC solvent delivery system: flow rate and gradient test.

以下测试作为HPLC溶剂传输系统周期性主动检查：流速和梯度检查

FLOW RATE 流速

Materials: 材料

Volumetric flask of 5 or 10 ml 5ml或10ml的容量瓶

Calibrated chronometer 校正过的气压计

Settings: 设置

Mobile phase: degassed water 流动相：脱过气的水 No column (open end)* 无柱（开放式）*

Flow rate: adjusted between 0.5 and 3.0 ml/min 流速：调节在0.5到3.0ml/min

If high-pressure mixing systems are installed, this test has to be done on each solvent channel.

如果有安装高压混合系统，本测试应在每一溶剂通道进行。

* For certain equipment, e.g. in the case of low flow rates, the check would be performed by using a column or a backpressure regulator.

* 对于一定的仪器，例如，流速较低时，采用一根柱子或背压调整器进行检查

Method方法:

Set the flow rate at an appropriate level and measure the time needed to fill the volumetric flask up to the mark. Record the time needed.

将流速设定在一个合适的水平，测量将容量瓶装满至刻度的时间。记录所用的时间。

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PA/PH/OMCL (11) 04 - OMCL Guideline on qualification of HPLC equipment (Annex 1) ＝＝(V×60)/t

f ................. measured flow rate [ml/min] 测量的流速

t ................. elapsed time to fill up to mark [s] 充满至刻度所花的时间 V ............... volume of the volumetric flask [ml] 容量瓶的体积 D＝100×(f－F)/F

D ............... deviation [%] 偏差

F ............... adjusted flow rate [ml/min] 调节后的流速 f ................. measured flow rate [ml/min] 测量的流速 Limit限度s: ±5%

GRADIENT COMPOSITION AND RIPPLE 梯度组成和波动

Settings: 设置

Stainless steel capillary e.g. 2000 x 0.12 mm installed instead of a column 不锈钢毛细管，例如2000X0.12mm，代替色谱柱安装于仪器 Detection: UV-Detector adjusted to 265 nm 检测器：紫外检测器调节至265nm Mobile phase A: degassed water 流动相A：脱气水

Mobile phase B: degassed water containing 0.5% acetone 流动相B：含0.5%的脱气水 Flow rate: 1.0 ml/min 流速：1.0ml/分钟

Method方法:

The test is carried out in the following way by using a gradient program depending on the number of solvent channels and the configuration of the system:

按下列方法进行测试，根据通道数量不同使用下列不同系统参数和梯度进行洗脱 A-B

A-B and A-C A-C, A-B and B-D % mobile phase B % mobile phase A time [min] %流动相B (water) 时间【分钟】 (water-acetone mixture) %流动相A（水相） （水-丙酮混合物） 0.0 100 0 0.1 90 10 10 90 10 10.1 50 50 20 50 50 20.1 10 90 30 10 90 30.1 0 100 40 0 100 1st July 2011 Page 6 of 19

PA/PH/OMCL (11) 04 - OMCL Guideline on qualification of HPLC equipment (Annex 1) 40.1 100 0 Start the test by pumping water for at least 10 min to equilibrate the system. 开始测试前，先用水平衡系统至少10分钟。

The zero % value at the start of the test is the baseline. All steps are measured at the

beginning of the horizontal part of the line either by software or manually on the paper print using a liner. The height of the 100% water/acetone mixture is used as the 100% value in the following calculation.

开始时的0%值是基线值。所有步骤均在开始时测量基线的水平部分，由软件或手动在纸上用衬垫打印。100%水/丙酮混合物的高度在以下计算中作为100%值。

＝100* h/H

%H ............ calculated composition 计算组成

h ................ height of the measured line 测量线的高度

H ............... height of the 100% water/acetone mixture line (mobile phase B)

100%水/丙酮混合物线的高度（流动相B） ＝%H－G

d ................ deviation 偏差

G ............... gradient composition adjusted [% acetone/water solution mixture = mobile phase

B]

梯度组成调节【%丙酮/水溶液混合物 = 流动相B】

Limit限度s:

Absolute deviation: ±2 of the adjusted value 绝对偏差： 调整值的±2

The ripple of the gradient composition is the percentage of noise of the 50% line from the gradient program.

梯度组分的波动为梯度程序噪声基线的50%

＝100* N /h50

%R ............ ripple 波纹

h50 ............. height of the 50% line 50%的线高度

N ............... height of the noise line, measured during 1 minute in the linear region 噪声线高度，在线性区域1分钟测量的结果

Limit限度s: ≤ 0.2 %

HPLC INJECTOR 进样器

Volume precision and carry-over are the tests proposed for the periodic and motivated check of the HPLC injector.

建议在HPLC进样器周期性主动检查中进行体积精密度和残留检查 VOLUME PRECISION AND CARRY-OVER 体积精密度和残留 Solutions: 溶液

Solvent A: methanol : water R, 60 : 40.

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PA/PH/OMCL (11) 04 - OMCL Guideline on qualification of HPLC equipment (Annex 1) 溶液 A：乙醇：水（试剂） = 60：40

Reference solution (a): dissolve 15.0 mg methyl-, ethyl-, and propylparabene in 100.0 ml of solvent A.

对照液（a）：取15.0mg乙基、甲基和尼泊金丙酯溶于100.0ml溶剂A中

Reference solution (b): Dilute 1.0 ml of reference solution (a) to 10.0 ml of solvent A. 对照液（b）：取1.0ml对照液（a）用溶于10.0ml溶剂A中

Reference solution (c): Dilute 1.0 ml of reference solution (b) to 100.0 ml of solvent A. 对照液（c）：取1.0ml对照液（b）用溶于100.0ml溶剂A中

Settings: 设置

Column: Lichrospher 100 RP8, 5um, 125 x 4 mm, without precolumn 色谱柱：Lichrospher 100 RP8, 5 um, 125 x 4 mm, 无衬管 Mobile phase: methanol : water = 60 : 40 流动相：乙醇：水 = 60：40 Flow rate: 1.0 ml/min 流动速度：1.0ml/分钟 Detection: 254 nm 检测器：254nm

Injection volume: 20 μl 进样体积：20ul

Method方法:

Injection scheme: 进样序列

— 6x reference solution (b) 对照液（b）6针

— 1x reference solution (a) 对照液（a）1针

— 1x solvent A (blank injection 1) 溶剂A（空白）1针 — 1x reference solution (b) 对照液（b）1针

— 1x solvent A (blank injection 2) 溶剂A（空白2）1针 — 1x reference solution (c) 对照液（c）1针

Limit限度s:

Repeatability of peak areas: The relative standard deviation of the peak areas of all peaks in the chromatogram obtained with the reference solution (b) should be ≤1.0 %. 峰面积重复性：对照液（b）色谱图中所有峰的峰面积相对标准偏差应≤1.0 %

Carry-over: The percentage of the peak area corresponding to propylparabene in the blank injection 1 does not exceed 0.5% of 10 times the peak area of the propylparabene peak in the chromatogram obtained with the reference solution (b) injected after the blank injection.

残留：尼泊金丙酯在空白图谱中的峰面积不超过在空白后进样的对照液（b）中尼泊金丙酯的对应峰面积的10倍的0.5%

The percentage of the peak area corresponding to propylparabene in the reference solution (c) is 0.9 – 1.1% of the peak area of the propylparabene peak in the chromatogram obtained with

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