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UV/Vis spectroscopy is a widely used analytical technique in laboratories of the pharmaceutical industry. Quality control and assurance analysis of pharmaceutical products are routinely performed under stringent quality guidelines established by international entities such as the United States Pharmacopeia (USP), thus ensuring fulfillment of Good Manufacturing Practices (GMP).
Periodical validation of spectrometer performance within the operational range is an essential part to ensure the reproducibility and reliability of the obtained results. The USP chapter <857> (43rd edition) describes performance qualification parameters for a UV/Vis spectrophotometer.
Analytik Jena’s SPECORD PLUS spectrophotometer series is fully compliant both in the device and software (Aspect UV) performance.
USP <857> Validation
Validation according to USP includes the following parameters:
1.Control of wavelengths and absorbance
2.NEW: Photometric response (linearity)
3.Limit of stray light
4.Control of resolution
USP <857> involves validation of the spectrometer in the operating range, i.e. covering the range in which routine or identity measurements are deployed. In order to assure compliance of the complete measuring range, both in the wavelength and in the absorbance, a wide range of possible validation methods, including certified reference materials (CRMs) and the corresponding target parameters and tolerance range(s) are unambiguously indicated. It is the responsibility of the user to select the suitable CRMs and methods to the required operating range. In addition to the general standards, alternative CRMs upon warranted certification are also tolerated by the USP <857>.
This allows validation of less common operational ranges (e.g., absorbance above the specified values).
Control of wavelengths
This qualification procedure ensures that the accuracy of wavelengths over the intended operational range is within the USP <857> specifications. Wavelength accuracy and precision have to be validated within the spectrometer’s operational range, therefore the USP provides a plethora of CRMs to suit user specifications (see Table 1 below).
400–780 nm ± 2 nm
Table 1: Certified reference material for the control of wavelength accuracy and precision according to USP <857>
Control of absorbance and photometric linearity
To ensure absorbance (or transmittance) accuracy, precision, and linearity of a given system within acceptable limits, the USP <857> specifies a clear procedure for this purpose (see Table 2). Different concentration ranges of CRMs are required to verify those parameters. K2Cr2O7 solution (from 20 mg/L to 200 mg/L), neutral density glass filters, as specified by the National Institute of Standards and Technology (NIST) are the accepted CRMs for this purpose. Photometric linearity, namely the range in which the absorbance-to-concentration correlation behaves linear, is validated by probing at least three concentration ranges in the intended operational range.
Table 2: Certified reference material for the control of absorbance and photometric linearity according to USP <857>
(235; 257; 313; 350 nm)
> 1 A: ± 1% A
> 1 A: ≤ ± 0.5% A
590 and 635 nm)
> 1 A: ± 0.8% A
Limit of stray light
Stray light might interfere with qualitative and especially quantitative UV/Vis measurements. Here, the USP
<857> stray light validation relies on cut-off filters, namely filters with a sharply defined spectrum. These block any light from passing through below a certain wavelength. Ideally, filters with a cut-off wavelength close to the required wavelengths are recommended (see Table 3).
The USP <857> stipulates two procedures for stray light determination. In the 10 mm vs. 5 mm method, a cell with 5 mm path length is first measured as a reference and then a 10 mm cell as a sample (both filled with the filter solution).
The resulting (absorbance) peak is used to determine the position and height of the maximum absorbance and the associated stray light value Sλ.
In the second method 10 mm vs. 10 mm H2O, the filter solution is measured against water as reference (both in a 10 mm cell). The measured absorbance at the certified wavelength (at the edge of an absorbance peak) must be greater than or equal to 2 A.
Table 3: Certified reference material for the control of stray light according to USP <857>
Requirement: A ≥ 0.7 A
Spectral resolution (or plainly resolution) is a measure to resolve two nearby standing peaks. In case of monochromator systems, the resolution is intimately related to the available spectral slit (or bandwidth). The USP <857> specifies the use of a 0.020% solution of toluene in hexane against the latter as reference (hexane). Herein, the absorption maximum and minimum (respectively at approximately 269 and 266 nm) are measured. Subsequently the resolving capacity is determined from the A269/A266 ratio.
The USP <857> states that a slit of 2 nm or smaller is necessary for most quantitative applications (monochromator system). Analytik Jena’s double beam spectrophotometer series SPECORD PLUS provides a clear solution for USP compliance. Herein the SPECORD 210 and 250 PLUS (0.5, 1, or 2 nm slit), as well as the SPECORD 50 and 200 PLUS devices (1.4 nm) meet the established USP requirements.
ASpect UV Software
Figure 1: ASpect UV Validation Support Software Module acc. to USP <857>
In addition to the suitable spectrometer, Analytik Jena provides solutions for all USP <857> software compliance requirements with the ASpect UV software. It provides powerful tools for flexible and compliant analysis in highly regulated environments as well as user-friendly spectrometer operation. With the comprehensive user management (FDA 21 CFR Part 11 compliant) and AJ file protect, data is protected against intentional and unintentional manipulation, thus guaranteeing data integrity (see Figure 1 for further details).
Standard measurements according to USP <857>
Table 4: Comparison of standard measurements of former versions up to USP 41 and the new versions from USP 42
Photometry range [Abs]
(1 slit width per validation)