Abstract
A feasibility study was conducted for a sensitive and robust dye immersion method for the measurement of container closure integrity of unopened prefilled syringes using fluorescence spectrophotometry as the detection method. A Varian Cary Eclipse spectrofluorometer was used with a custom-made sample holder to position the intact syringe in the sample compartment for fluorescence measurements. Methylene blue solution was initially evaluated as the fluorophore in a syringe with excitation at 607 nm and emission at 682 nm, which generated a limit of detection of 0.05 μg/mL. Further studies were conducted using rhodamine 123, a dye with stronger fluorescence. Using 480 nm excitation and 525 nm emission, the dye in the syringe could be easily detected at levels as low as 0.001 μg/mL. The relative standard deviation for 10 measurements of a sample of 0.005 μg/mL (with repositioning of the syringe after each measurement) was less than 1.1%. A number of operational parameters were optimized, including the photomultiplier tube voltage, excitation, and emission slit widths. The specificity of the testing was challenged by using marketed drug products and a protein sample, which showed no interference to the rhodamine detection. Results obtained from this study demonstrated that using rhodamine 123 for container closure integrity testing with in-situ (in-syringe) fluorescence measurements significantly enhanced the sensitivity and robustness of the testing and effectively overcame limitations of the traditional methylene blue method with visual or UV-visible absorption detection.
LAY ABSTRACT: Ensuring container closure integrity of injectable pharmaceutical products is necessary to maintain quality throughout the shelf life of a sterile drug product. Container closure integrity testing has routinely been used to evaluate closure integrity during product development and production line qualification of prefilled syringes, vials, and devices. However, container closure integrity testing has recently gained industry attention due to increased regulatory agency scrutiny regarding the analytical rigor of container closure integrity testing methods and expectations to use container closure integrity testing in lieu of sterility tests in stability programs. Methylene blue dye is often used for dye ingress testing of container closure integrity, but we found it unsuitable for reliable detection of small breaches in prefilled syringes of drug product. This work describes the suitability and advantages of using a fluorescent dye and spectroscopic detection for a robust, sensitive, and quality control–friendly container closure integrity testing method for prefilled syringes.
1. Introduction
Prefilled glass syringes have been increasingly used for delivery of parenteral drugs and biological products (1, 2). Prefilled syringes function as a primary packaging component that provides protection and maintains efficacy and product sterility prior to use. Development of drug product using such syringes, and testing to demonstrate the sterile product packaging integrity, must follow regulatory agency requirements (3, 4). The U.S. Food and Drug Administration (FDA) requests the use of the USP sterility test as a part of the stability protocol for sterile products, with testing at initial release and at the stability end point (5). The FDA further provided guidance for industry to use container closure integrity testing (CCIT) as an alternative to sterility testing, performed throughout the product shelf life. In draft USP guidance, it is recommended to perform integrity testing at three phases throughout the life cycle of the sterile product: initial development of the product packaging, routine manufacturing, and shelf life stability assessment (6).
Many physical or chemical methodologies have been proposed and described for CCIT (7⇓–9). More detailed research and development work on CCIT has been published, including pressure/vacuum decay (10⇓⇓–13), trace gas permeation/leak tests (14⇓⇓–17), dye ingress tests (18⇓–20), electrical conductivity and capacitance tests (21⇓–23), and microbial challenge or immersion tests (19). These methods exhibit many advantages compared to conventional USP sterility testing in demonstrating the potential for product contamination over the product shelf life. For example, these methods may require less time than the sterility test method, detect a breach of the container/closure system prior to product contamination, and reduce the potential for false-positive results when compared to sterility tests. Some of the methods are non-destructive, which may allow sample to be conserved, and allow on-line inspection of container closure integrity for every vial of drug product, depending on the physical properties of the filled product (e.g., inerted with nitrogen, or under vacuum).
Among the many physical or chemical testing methodologies, dye ingress testing is the most commonly used method for CCIT. Dye ingress testing historically uses methylene blue dye. Besides a vacuum vessel, it does not require special instruments or technology. Detection is typically based on visual observation. A failure is determined when the dye is observed in the container, which proves ingress. This method is simple, inexpensive, widely accepted by industry and health authorities, and recommended by most compendia (24, 25). However, the dye ingress method is a limit test and not a quantitative approach. Traditional dye methods are also generally not as sensitive as some of the methods mentioned earlier using modern technologies (26).
UV-visible (UV-vis) spectrophotometry has been applied to the detection of dye ingress for CCIT in order to overcome the limitations of visual detection. Jacobus et al. (18) developed a spectrophotometric dye immersion test method using an absorption maximum of Quinizarin Green dye at 648 nm for the CCIT of an oil-based product in glass vials. The sensitivity of the method was 0.1 μg/mL, corresponding to an absorbance of 0.002 AU. Burrell and coworkers (19) used a photo diode array detector to measure the absorbance of FD&C Red dye solutions at 506 nm for the CCIT of 5 mL tubing glasses. They reported that the dye solution spectrophotometric detection limit was similar to the visual detection limit, approximately at 0.0025 μL of dye/mL, which corresponds to an absorbance of 0.002 AU.
UV-vis spectrophotometric detection is more robust and typically offers lower detection limits in comparison with visual analysis. However, significant challenges were encountered in our laboratories when applying methylene blue dye immersion with UV-Vis spectrophotometric detection to drug product prefilled glass syringes (20). Due to the small diameter of the syringe (6.35 mm internal diameter) and possible low concentration of the dye intruded, direct spectrophotometric scanning of the intact syringe could not detect a signal comparable to that seen in glass vials and a similar detection limit could not be reached. An alternative method was evaluated in which the sample solution in the syringe was transferred to a cuvette to increase the effective pathlength. While the measurement in the cuvette improved sensitivity, the transfer procedure was labor-intensive and required multiple extra steps to reduce the potential for contamination, which increased the complexity, variability, and false-positive risk of the CCIT measurement. Consequently, this approach was not desirable for routine use.
In this study, fluorescence spectrophotometry was evaluated for the feasibility of developing a sensitive and robust method for the dye ingress CCIT of prefilled glass syringes. The evaluations include dye selection, optimization of operational parameters, comparison with visual and UV-Vis detection, and specificity with actual drug products. The objectives were to enable the fluorescence measurement of the unopened prefilled syringes after dye immersion without liquid transfer, develop a sensitive method with a better limit of decision compared to visual and UV-Vis methods, and to simplify the testing procedure to fit the needs for quality control (QC) and stability studies.
2. Materials and Methods
2.1. Materials
The study used prefillable glass syringes that have a staked needle with a rubber needle shield; the outside diameter of the syringes was 8.15 ± 0.10 mm and the syringe fill volume was 1 mL (Forma 3s, Schott Pharmaceutical Systems, Emlsford, NY); see Figure 1. The syringes are equipped with a plunger stopper made from a fluropolymer (BD Hypack, Becton, Dickinson and Company, Franklin Lakes, NJ).
Prefilled glass syringe. GB: glass barrel; BN: barrel/needle; TL: total length; NS: needle shield; PS: plunger stopper; ID: inside diameter; OD: outside diameter.
Methylene blue (3,7-bis(dimethylamino)phenazathionium chloride), certified by the Biological Stain Commission, was purchased from Sigma Aldrich (St. Louis, MO). Rhodamine 123, (2-(6-amino-3-imino-3H-xanthen-9-yl)benzoic acid methyl ester), BioReagent, was also purchased from Sigma Aldrich.
For assessment of the specificity of the fluorescence method, several injectable drug products were purchased: lidocaine HCl injection, USP, 20 mg/mL (Hospira Inc., Lake Forrest, IL), phenytoin sodium injection, USP, 50 mg/mL (Baxter, Deerfield, IL). Bovine serum albumin solution (A1595), 10% was purchased from Sigma-Aldrich.
2.2. Methods
2.2.1. CCIT by Dye Ingress:
0.1% methylene blue dye in water was placed into a vacuum vessel. Test samples were prepared with the glass syringes filled with testing medium (water for injection [WFI] or drug product). The prefilled syringes were then placed into the vacuum vessel and immersed. The timed pressure and vacuum cycles stated by USP (see Table I) were applied to the immersed syringes to simulate possible transportation environment changes. After the treatment, the samples were removed, rinsed, and analyzed.
Dye Ingress Requirements for Aseptic Drug Product in Vials
2.2.2. Detection of Dye Ingress by Visual Observation:
An aliquot of the testing medium was transferred into a glass syringe, which was used as a negative control. A series of standard solutions of methylene blue in WFI at different concentrations from 0.005 to 1.00 μg/mL were prepared and filled in a set of glass syringes. Through visual comparison by qualified analysts for the standard solutions with the negative control, a limit of detection (LOD) was determined; this is the lowest concentration at which coloration of the solution due to the dye is consistently detected by visual observation. Accordingly, a limit of decision (LD) should be determined for the method that is above the LOD. The LD must be at or below the concentration of dye that results from a small breach (typically a 5–10 μm capillary) and clearly distinguishable from a sample with no dye. The glass syringe filled with the solution of methylene blue in WFI at this LD concentration was used as a positive control.
2.2.3. Detection of Dye Ingress by UV-Vis Spectrometry:
An aliquot of the testing medium was used as a negative control. A series of solutions of methylene blue in WFI at different concentrations was prepared for determination of the method sensitivity. An Agilent 8543 UV-vis spectrophotometer (Agilent Technologies, Santa Clara, CA) was used for the UV-vis measurement at 635 nm. The samples were measured either through a 10 mm quartz cuvette or through the barrel of the filled syringe. The testing results were statistically evaluated to define the instrument baseline and variation for the medium blank. The LOD for the UV-vis measurement was determined based on the baseline variation of the spectroscopic measurement, with the LOD defined as being equal to 3 times the baseline noise. The LD for the UV-vis method was set as being equivalent to the signal from the concentration of dye in the syringe that resulted from the 5 μm capillary breach. The syringe samples filled with the dye solution or treated by dye immersion were either directly scanned by UV-vis through the barrel, or analyzed after transferring the liquid in the syringe into the cuvette. For the dye immersion–treated syringe samples, the syringe was subjected to six rinse cycles using fresh deionized water for each cycle. To transfer the liquid in the syringe into the cuvette, the syringe needle shield was carefully removed, and the first several drops of the liquid in the needle were discarded to avoid dye carryover. A CCIT failure was identified when the detected absorption value was greater than that of the LD.
2.2.4. Development of a Fluorescence Spectrometric Method for Detection of Dye Ingress:
A Cary Eclipse fluorescence spectrophotometer (Agilent Technologies, Santa Clara, CA) was used for the detection of the dye ingress in the prefilled glass syringes. It has the following wavelength ranges: excitation 200–900 nm and emission 200–900 nm. The wavelength accuracy and resolution were ±1.5 nm. The data collection and processing were performed by using the Cary Eclipse Scan software (Agilent Technologies, Santa Clara, CA). Both methylene blue and rhodamine 123 (see Figure 2a and 2b) were evaluated as fluorescence indicators for dye ingress CCIT of prefilled syringes. A syringe holder was designed and applied to hold the syringe in the sample compartment of the spectrophotometer to improve the repeatability of the fluorescence measurement (see Figure 3). Key operational parameters were optimized including the voltage of the photomultiplier tube (PMT)—PMT standard voltage settings of high (800 V), medium (600 V), and low (400 V)—and the width and shape of the excitation and emission slits (1.5, 2.5, 5, 10, and 20 mm rectangular; 10 mm round).
(a) Methylene blue; (b) Rhodamine 123.
Cuvette syringe holder: Left, holder; center, syringe; right, syringe inserted into the holder.
2.2.5. Assessment of Fluorescence Spectrometric Method:
The fluorescence spectrometric method using rhodamine 123 was assessed with respect to specificity, sensitivity, accuracy, precision, linearity, and range.
For the method specificity study, two types of test were performed for the commercial drug products and the biologic samples. In the first test, 1 mL of the lidocaine HCl injection, phenytoin sodium injection, or the bovine serum albumin solution (diluted for 1:10 in WFI to give a 1% w/v protein solution) was filled into a syringe. Fluorescence was measured using the optimized method parameters to confirm the absence of fluorescence at the emission wavelength. In the second test, each of the drug product samples was spiked with rhodamine 123 at the level of 0.005 μg/mL. Fluorescence was then measured for each spiked sample using the optimized method parameters to confirm that there was no interference to the measurement at the emission wavelength.
3. Results and Discussion
3.1. Sensitivity of Detection Methods for Dye Ingress
3.1.1. Visual Observation Method:
Use of dye ingress for CCIT has been widely accepted by the industry and regulatory agencies for decades. A number of dyes have been utilized as the indicator of the test, including Quinzarin Green (18) and FD&C Red (19), but the most commonly used and recommended by USP is methylene blue (24, 25). Traditionally, the test has relied on visual observation, and it requires no special instruments or technology. The test article can be checked in its original packaging container, such as a vial or syringe, after being treated with the dye immersion cycle. Figure 4 shows methylene blue in the prefilled glass syringes. The pass or fail decision for the test is determined based on whether dye ingress is seen in the treated container. The dye ingress test is relatively simple and efficient, and it is recommended by most compendia. However, this test typically suffers from several disadvantages: (1) there is no standard procedure or guideline for the test; (2) the visual analysis may be subject to human error and analyst-to-analyst variation if not properly implemented; (3) the test sensitivity is limited.
Methylene blue in syringes. A blue color is clearly visible at 1 ug/mL, and faintly visible at 0.1 ug/mL. At 0.05 ug/mL, no blue coloration is visible (shadowing in the image causes the syringes to appear slightly darker).
In this study, the visual LOD for the methylene blue in WFI in syringes was determined by visual comparison of the dye solutions in the syringe with the syringe filled with WFI blank. The LOD was 0.1 μg/mL. At this level, the dye is visible in the syringe in comparison with the syringe filled with WFI blank. However, the dye is not visible in the syringe filled with 0.05 μg/mL methylene blue. The small diameter of the syringe and the strong curvature of the glass barrel reduce the effectiveness of the visual inspection.
3.1.2. UV-Vis Spectrometric Method:
UV-sis has been adopted for the detection of dye ingress in CCIT of glass vials (18, 19). However, the conventional spectrometric dye method for CCIT of vials was shown not to be robust for a QC environment when applied to syringes. Two approaches were evaluated in this lab (20) using UV-vis spectrometry for the measurement of the dye ingress in prefilled glass syringe: (1) direct scan through the syringe barrel, and (2) analysis after transferring the liquid in the syringe into a cuvette. Results from the scan through the barrel showed significant noise that was ∼10 times higher than that from the scan through the cuvette. This is likely due to the small diameter barrel, which is difficult to position for measurement of UV transmittance, and the light scattering from the large curvature of glass barrel. Both of these factors generate high variability in the measurement.
While the use of the cuvette for the UV-vis measurement of dye ingress improves the sensitivity, it does have a significant drawback in that the sample solution needs to be transferred to the cuvette from the syringe after the dye immersion treatment. Thorough washing of the syringe with the needle shield attached is challenging. Some post-rinse samples had evidence of dye in the shield housing, and the dye occasionally transferred to the syringe tip as the needle shield was removed, resulting in dye carry-over and contamination during the solution transfer to the cuvette. The sample solution transfer adds extra operational steps that increase the complexity, variability, and risk of false positives from the test, making the procedure problematic for use in a QC environment.
3.2. Development of a Fluorescence-Based Method
Fluorescence spectrometry was considered for the development of a sensitive and robust method for the dye ingress CCIT of syringes because it generally has higher sensitivity and selectivity in comparison to UV-vis. It could enable the in situ measurement of prefilled syringes after dye immersion without liquid transfer, simplify the testing procedure to fit QC needs, as well as provide a better LOD and robustness compared to the visual method and the UV-vis method.
3.2.1. Methylene Blue as a Fluorescence Dye:
Methylene blue is the most common dye used for CCIT with visual or UV-vis detection due to its signature color from absorption at 664 nm with a molar extinction coefficient 95,000 cm–1/M (27). It has also been reported that methylene blue has fluorescence with excitation wavelength at 607 nm, emission wavelength at 682 nm, and quantum yield of 0.03 (28, 30). The fluorescence spectra of a series of the methylene blue in WFI solutions in a syringe are shown in Figure 5. The emission peak at 682 nm from the solution of 0.05 ug/mL dye can be differentiated from the blank. Table II shows the measurements from different syringes at three concentration levels, and demonstrates good reproducibility with small relative standard deviations (RSDs) ≤3% for concentrations of 0.05 μg/mL and above. Because high Rayleigh scattering was observed in these measurements, including for the blank sample, the detection of fluorescence from the methylene blue was compromised. Fluorescence of methylene blue does not provide an obvious improvement compared to absorbance detection, probably due to the low quantum efficiency for this compound.
Fluorescence spectra of methylene blue determined in a syringe.
Reproducibility of Measurements Made from Different Syringes at Three Different Concentration Levels (Methylene Blue)
3.2.2. Rhodamine for Dye Ingress Testing:
Further studies were performed using rhodamine 123, a dye widely used in biological applications with stronger fluorescence. Rhodamine 123 has a molar extinction coefficient of UV-vis absorption (at 512 nm) of 85,200 cm–1/M that is similar to methylene blue. However, the fluorescence quantum yield of rhodamine is 0.9 (with excitation at 480 nm and emission at 525 nm) (30, 31), which is 30 times that of methylene blue. Greater sensitivity is thus expected from using rhodamine 123 for dye ingress detection. Figure 6 shows the rhodamine 123 fluorescence spectra of the syringe samples. The intensity of the fluorescence band at 525 nm is significantly higher than the methylene blue band at 682 nm (Figure 5) for equivalent concentration levels. This result shows that rhodamine 123 in the syringe can be easily detected by fluorescence as low as 0.01 ug/mL without optimization. The detected rhodamine 123 fluorescence signal was more intense (>10 fold) than methylene blue under the same measurement condition (PMT voltage: 600 V; excitation slit: 5; emission slit: 5).
Fluorescence spectra of rhodamine 123 determined in a syringe.
3.3. Optimization of the Fluorescence Detection Method
3.3.1. Design of Syringe Holder for In-situ Measurement:
To measure the dye ingress in situ through the syringe barrel, the intact syringe needs to be positioned in the measurement center of the sample compartment of the spectrometer, which allows illumination by the excitation light without blocking of the fluorescence emission and scattering. A syringe holder is needed to keep the syringe steady to ensure the repeatability of the measurements. Wide variability from the fluorescence measurements was observed in our preliminary experiments without use of a suitable syringe holder. To overcome the problem, a syringe holder was designed in-house (see Figure 3). Syringes were filled with different concentrations of dye, and each syringe was placed in the holder and re-positioned six times to take measurements. The results showed a measurement RSD of <3.0%. This indicates that irreproducibility due to positioning of the device could be reduced to acceptable levels by use of an appropriate sample holder.
3.3.2. Optimization of Instrument Parameters:
The Cary Eclipse fluorescence spectrophotometer allows selection of a number of operational parameters. The sensitivity of fluorescence detection for rhodamine 123 in prefilled syringes was further enhanced by optimization of the key instrument parameters, including the voltage of the PMT, and the width and shape of the excitation and emission slits. Figure 7 illustrates some of the spectra obtained for a sample of 0.01 μg/mL rhodamine in the syringe using the different instrument parameters during the method optimization. When signal-to-noise ratio (S/N) for rhodamine measurement was determined, the fluorescence intensity at 525 nm and the baseline noise at 650 nm (where no fluorescence signal was detected) were utilized. It was found that the S/N was greater when high PMT voltage (800 V) was applied in comparison with the use of the medium (600 V) and low (400 V) when the width and the shape of the excitation and emission slits were fixed. Testing was also performed using manual mode photomultiplier settings of 900, 950, and 1000 V. Results showed the fluorescence signal increases with the increased PMT voltage. However, the noise also increases, and the S/R becomes worse than at 800 V. Furthermore the dynamic range of the recorded signal becomes narrower when a higher photomultiplier voltage is used, and the detector is easily saturated. Our experience showed that the use of 800 V voltage setting is the optimal for these measurements on this specific type of fluorometer.
Fluorescence detection optimization. Significant changes in the fluorescence intensity are observed when using different slits.
When the PMT voltage was fixed, and the bigger the excitation and emission slits used, the higher the S/N obtained. In addition, the measurements using a round shape excitation slit (10 R) generated a better S/N than those using the rectangular excitation slit. However, the measurements using a round shape emission slit (10 R) generated a much poorer S/N than that using rectangular shape emission slit. Results showed that a suitable measurement was obtained using PMT voltage: high; excitation slit: 10 R; and emission slit: 10. Under such conditions, the S/N for the syringe with 0.001 μg/mL rhodamine 123 was determined as >24, and rhodamine 123 in the syringe can be detected at concentrations as low as 0.0001 μg/mL.
3.3.3. Method Assessment:
The specificity was assessed by measuring the spectra of the media and samples in the syringe using the fluorescence methodology: (1) WFI, (2) methylene blue in WFI solutions, and (3) rhodamine 123 in WFI solutions. As shown in Figures 5 and 6, the excitation and emission wavelengths (607 nm and 682 nm for methylene blue, 480 nm and 525 nm for rhodamine 123) were determined based on the wavelength of maximum fluorescence of the dyes at wavelengths with negligible background interference.
The subsequent study using the commercial drug products and the protein sample further confirmed the method specificity. With the excitation at 480 nm, no fluorescence peak was detected at or close to 525 nm for any of the tested commercial drug products or the protein sample. In addition, the fluorescence peak was detected at 525 nm in all these rhodamine-spiked commercial drug products and protein sample. As demonstrated in Figure 8, the active drugs and protein did not show any interference to the rhodamine 123 fluorescence measurement.
Fluorescence detection specificity assessment. The spectra showed are from the water blank, drug product as is, and spiked with rhodamine at the level of 0.005 μg/mL: (a) lidocaine HCl injection, (b) phenytoin sodium injection, and (c) bovine serum albumin solution.
The sensitivity was evaluated using the optimized instrument condition. The fluorescence intensity of a WFI blank solution at 525 nm was less than 2 RFU (relative fluorescence units). The standard deviation of six blank solutions was measured at 525 nm. The LOD was defined as 3 times the RSD of these six measurements. The RSD of these measurements was 0.65 RFU, and the LOD for this method was 1.96 RFU. The fluorescence intensity of a diluted rhodamine 123 solution with a concentration of 0.00005 μg/mL was below the detection limit. However, the fluorescence intensity of a 0.0001 ug/mL sample was 4.5 RFU. With such a low LOD, this method can provide a sensitive and robust approach that can detect the dye in the syringe from a 5 μm breach as described by Burrell et al (19).
The accuracy and precision were assessed using two procedures for prefilled syringes containing rhodamine 123: (1) Measure 10 individual syringe samples, 0.05 μg/mL and 0.005 μg/mL, (each was measured once) and calculate the RSD of the fluorescence intensity. The results showed an RSD of 0.9% at a concentration of 0.05 μg/mL and 1.1% at 0.005 μg/mL (see Table III). (2) Measure one syringe sample from the lower level 10 times with re-positioning of the syringe, by rotating the syringe in the holder for each measurement, and calculate the RSD. The resulting RSD was 6.6%. Note that the measurement variation of 10 syringes from syringe to syringe is more significant than that of a single syringe being re-positioned 10 times, suggesting greater between-syringe variability. A similar assessment was also performed for prefilled syringes containing methylene blue. Results showed that the RSDs for the measurements of the methylene blue–containing syringes using the same two procedures are much larger than those of the rhodamine-containing syringe samples, even though the concentration of the methylene blue in the samples is much higher. It is also observed that the measurement variation of 10 methylene blue–containing syringes from syringe to syringe is more significant than that of a single methylene blue–containing syringe being re-positioned 10 times.
Fluorescence Measurements* Using Optimized Instrument Parameters
The linearity was evaluated over a rhodamine 123 concentration range from 0.001 μg/mL to 0.01 μg/mL. The results demonstrated a good linearity over this range with an equation of y = 66378x + 2.99 (x is the dye concentration, and y is the fluorescence intensity) and a correlation coefficient of R2 = 0.9948. The % y-intercept was 0.88. As demonstrated, the dye-ingress test with fluorescence detection is suitable for evaluation of container closure integrity of drug product in the prefilled syringe.
4. Conclusion
We successfully demonstrated in this study that it is feasible to develop a sensitive and robust CCIT method for unopened prefilled syringes by using dye ingress with in-situ fluorescence detection. The conventionally used dye for CCIT, methylene blue, could not provide the sensitivity needed for in-situ syringe measurements. The detection limit is at the level of 0.05 μg/mL. As an alternative, rhodamine 123 has stronger fluorescence and can meet the application need. Holding the syringe steady is a key for the in-situ detection. A sample holder was designed and applied, which significantly enhanced the repeatability of the fluorescence measurement. A detection limit of less than 0.0001 μg/mL or 0.1 ppb is feasible to detect dye in the syringe from a 5 μm breach and to meet the need for the CCIT application for prefilled syringes.
The method developed in this study using fluorescence detection is not only suitable for the WFI-filled syringes for component qualification and filling process validation, but is also applicable to testing injectable drug products for stability, especially for biologics and protein products. Further studies are underway to expand the method's application, including evaluation of the compatibility of rhodamine with specific drug product solutions, evaluating other fluorescent dyes, determination of practical LOD for the product in prefilled syringe, and validation of the CCIT procedures based on the practical detection limits.
Conflict of Interest Declaration
The authors declare that they have no competing interests.
Acknowledgements
The authors would like to thank Yong Quan and Thomas Haby for providing dyes and samples, and Jayshree Patel, Lili Lo, and Dipti Patel for their assistance in preparing syringes and testing samples.
- © PDA, Inc. 2016
