Chromatographic column for HPLC

Abstract

The utility model discloses a chromatographic column for HPLC (High Performance Liquid Chromatography), which comprises a column body, a filler and an end cover, the inner diameter of the column body is 3.0-4.6 mm, the column length is 10-30 mm, the particle size of the filler is 1.7-2.7 mu m, and a bonding phase is C8 or C18 or C8 polar embedding or C18 polar embedding; after the column length, the inner diameter, the particle size of the filler, the bonding phase and the like of the chromatographic column are improved, when the chromatographic column is applied to an existing chromatographic instrument, the detection time is shortened, and the sensitivity, the precision and the linearity meet the requirements of preparation sample content determination.

Description

Technical Field

[0001] The utility model relates to the field of high performance liquid chromatography, in particular to a chromatographic column for HPLC. Background Art

[0002] In the quality control analysis of pharmaceutical preparations, high performance liquid chromatography (HPLC) is the most commonly used high-precision analysis method. Using the traditional high performance liquid chromatography analysis method, the commonly used chromatographic column specifications are 75mm in length, 4.6mm in inner diameter, and 5μm in filler particle size (the column efficiency of the chromatographic column is about 15000 evaluated by L / dp), and the column volume is usually about 850μL. Calculated at a conventional flow rate of 1.5mL / min, the dead time of the method is greater than 0.57min. In order to ensure that the main peak and impurities can maintain sufficient separation, the analysis time of a single sample is usually 15-20min (under a column pressure slightly higher than 200bar). For highly retained compounds, due to their late peak elution, large peak width, and low theoretical plate number N, in order to obtain sufficient sensitivity, higher requirements will be placed on the column efficiency of the chromatographic column. With the development of modern liquid phase technology, a major breakthrough has occurred in chromatographic performance, namely the development of sub-2 micron stationary phase fillers with narrow particle size distribution, and HPLC analysis has also evolved into UHPLC and UPLC analysis. Taking the commonly used UPLC column as an example, its length is 50mm, inner diameter is 2.1mm, particle size is 1.8μm (the column efficiency is about 27000 when evaluated by L / dp), and its column volume is usually 120μL. Since the column pressure will increase exponentially after the particle size of the column changes, the flow rate of the conventionally used column of this particle size is generally less than 0.4mL / min. At this flow rate, the system pressure is still 8 times that of the HPLC method. At this time, the dead time of the method is greater than 0.3min. Since the column efficiency increases exponentially after the particle size is reduced, the separation between chromatographic peaks is improved, and the analysis time of a single sample can be shortened to 4-5min accordingly. However, in order to adapt to the problem of increased system pressure caused by the reduction of column particle size, the requirements for UHPLC and UPLC instruments have also increased accordingly. If the back pressure exerted by smaller particles in the chromatographic column increases, pumps and pipe joints that can operate at higher pressures need to be used; to ensure a smaller system volume, the upgraded detector flow cell needs to have a very small internal volume; at the same time, the data acquisition rate needs to be increased accordingly to ensure that the chromatographic peak has enough data points. The above requirements for the instrument hardware system have led to a much higher purchase cost for UHPLC and UPLC instruments than for HPLC instruments. At the same time, as the instrument pipelines become thinner, the maintenance cost during use and the training cost of personnel also increase accordingly.

[0003] When the content of preparation samples is tested, since the inspection items such as dissolution, content uniformity and mixing uniformity only focus on the main peak, there are only requirements for the sensitivity, precision, linearity, etc. of the method, and the requirements for separation are relatively small. When HPLC is used, the detection speed is slow, and when UHPLC or UPLC is used, although the detection speed is improved, the detection cost also increases. Utility Model Content

[0004] The technical problem to be solved by the utility model is to provide a chromatographic column for HPLC, which can improve the detection speed when the content of the preparation sample is detected, and meet the requirements for sensitivity, precision and linearity at the same time.

[0005] In order to solve the above technical problems, the utility model provides a chromatographic column for HPLC, comprising:

[0006] The column body has an inner diameter of 3.0-4.6 mm and a column length of 10-30 mm;

[0007] Filler, the particle size of which is 1.7-2.7 μm, and the bonding phase is C8 or C18 or C8 polar embedded or C18 polar embedded;

[0008] End caps are installed at the top and bottom ends of the column body to seal the filler in the column body.

[0009] Preferably, the inner diameter of the column body is 4.6 mm and the column length is 20 mm.

[0010] Preferably, the matrix of the filler is fully porous or solid core silica gel.

[0011] Preferably, the matrix of the filler is solid core silica gel.

[0012] Preferably, the pore size of the matrix of the filler is

[0013] Preferably, the bonded phase is polar amide and diisopropyl C18 polar embedding.

[0014] Preferably, the carbon loading is 9-15%.

[0015] Preferably, the end caps are detachably mounted on the top and bottom ends of the column body.

[0016] Preferably, the end cap installed at the top end of the column body is provided with a liquid inlet, and the end cap installed at the bottom end of the column body is provided with a liquid outlet.

[0017] The utility model improves the column length, inner diameter, filler particle size, etc. of the chromatographic column, and when applied to an existing chromatograph, the detection time is shortened, and the sensitivity, precision and linearity meet the requirements for the content determination of preparation samples. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] In order to more clearly illustrate the technical solution of the utility model, the following briefly introduces the drawings required for use in the utility model. Obviously, the drawings described below are only some embodiments of the utility model. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0019] Figure 1 It is a schematic diagram of Embodiment 1 of the HPLC chromatographic column of the utility model;

[0020] Figure 2 This is a chromatogram of the caffeine test in Example 2 of the present utility model;

[0021] Figure 3A This is a sample test chromatogram of UPLC in Example 3 of the present utility model;

[0022] Figure 3B This is a sample test chromatogram of HPLC in Example 3 of the present utility model;

[0023] In the figure, 1-column body; 2-filler; 3-end cover. DETAILED DESCRIPTION

[0024] The following will be combined with the accompanying drawings to clearly and completely describe the technical solutions in the utility model. Obviously, the described embodiments are part of the embodiments of the utility model, not all of the embodiments. Based on the embodiments of the utility model, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the utility model.

[0025] Embodiment 1

[0026] refer to Figure 1 , shows a chromatographic column for HPLC of the utility model, comprising:

[0027] The column body has an inner diameter of 3.0-4.6 mm, preferably 4.6 mm; the column length is 10-30 mm, preferably 20 mm;

[0028] Filler, the particle size of which is 1.7-2.7 μm; the bonded phase is C8 or C18 or C18 polar embedding, preferably polar amide and diisopropyl C18 embedding;

[0029] End caps are installed at the top and bottom ends of the column body to seal the filler in the column body.

[0030] In the embodiment of the utility model, by optimizing the inner diameter of the column body, the column length and the filler particle size, the chromatographic column provided by the present application can achieve fast peak generation and short detection time when applied to HPLC, and the method has high sensitivity, high precision and good linearity, which can at least meet the requirements of the content determination of the preparation sample. It should be noted that the void volume (i.e., the remaining volume after deducting the volume occupied by the filler from the column volume) of the chromatographic column of the utility model does not exceed 230 microliters. If it exceeds 230 microliters, the desired effect cannot be achieved.

[0031] In a specific embodiment, the matrix of the filler is fully porous or solid core silica gel, preferably solid core silica gel, and the pore size of the filler matrix is ​​preferably Preferably, the bonded phase is polar amide and diisopropyl C18 polar embedding.

[0032] In a specific embodiment, the carbon loading is 9-15%.

[0033] In a specific embodiment, the end cap is detachably installed at the top and bottom ends of the column body. It can be understood that the end cap installed at the top end of the column body has a liquid inlet for sample injection, and the end cap installed at the bottom end of the column body has a liquid outlet for the sample to flow out of the chromatographic column.

[0034] Embodiment 2

[0035] Based on Example 1, this embodiment provides a chromatographic column with the following specifications:

[0036] Column length: 20 mm;

[0037] Column diameter: 4.6mm;

[0038] Column packing: Bonus RP packing, particle size 1.8 μm, bonded phase is polar amide and diisopropyl C18 embedded, carbon loading is 9.5%;

[0039] Chromatograph: Agilent 1260 Infinity II;

[0040] Sample: Caffeine, CAS 58-08-2;

[0041] The chromatogram of the sample is Figure 2 As shown, the peak time is 0.471 min, the running time of the whole method is 1 min, the retention factor is 2.36, and the column efficiency is 3209.

[0042] In addition, after the chromatographic column of this embodiment was used to test the other 87 samples (as shown in Table 1), the peak times of the samples were all within 0.45-0.55 min, the running time of the entire method was within 1 min, the retention factors of the compounds under the corresponding method were all k≥2, and the tailing factors of 94% of the compounds were within the range of 0.8-1.5, and the column efficiencies of the chromatographic columns were all >1000.

[0043] Table 1

[0044]

[0045]

[0046]

[0047] Embodiment 3

[0048] The preparation samples were analyzed by ultra-high performance liquid chromatography (using a commercial chromatographic column BEH C18) and liquid chromatography (using the chromatographic column of Example 1 of the present invention). The results are shown in Tables 2 and Figure 3A and Figure 3B ,in, Figure 3A This is the chromatogram of UPLC. Figure 3B It is a chromatogram of HPLC.

[0049] Table 2

[0050]

[0051] It can be seen that compared with the commercially used UPLC, the liquid chromatograph using the chromatographic column of the utility model to analyze the sample has a shorter running time and a retention factor greater than 2, which meets the requirements of the FDA (Reviewer Guidance, Validation of Chromatographic Methods) guidance document; while when the UPLC method running time is set to 2 minutes, its retention factor k is less than 2, which does not meet the requirements of the FDA guidance document.

[0052] The above description is only a preferred embodiment of the present invention and is not intended to limit the scope of protection of the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A chromatographic column for HPLC, characterized in that: include: The column body has an inner diameter of 3.0-4.6 mm and a column length of 10-30 mm; Filler, particle size is 1.7-2.7μm, bonded phase is C8 or C18 or C8 polar embedded or C18 polar embedded; End caps, which are mounted on the top and bottom ends of the column body to seal the filler in the column body; The pore volume of the column does not exceed 230 μl.

2. The chromatographic column according to claim 1, characterized in that The inner diameter of the column body is 4.6 mm and the column length is 20 mm.

3. The chromatographic column according to claim 1, characterized in that The matrix of the filler is fully porous or solid core silica gel.

4. The chromatographic column according to claim 3, characterized in that The matrix of the filler is fully porous silica gel.

5. The chromatographic column according to claim 3, characterized in that The pore size of the matrix of the filler is 80-120Å.

6. The chromatographic column according to claim 1, characterized in that The bonded phases are polar amide and diisopropyl C18 polar embedding.

7. The chromatographic column according to claim 1, characterized in that The carbon loading is 9-15%.

8. The chromatographic column according to claim 1, characterized in that The end caps are detachably mounted on the top and bottom ends of the column body.

9. The chromatographic column according to claim 1, characterized in that The end cap installed at the top end of the column body is provided with a liquid inlet, and the end cap installed at the bottom end of the column body is provided with a liquid outlet.

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