2021 Volume 69 Issue 4 Pages 352-359
Petrolatum ointment, which is an oleaginous ointment, is generally produced through manufacturing processes such as melting, mixing, and cooling. In this type of semisolid formulation, the manufacturing conditions of each process are empirically known to affect the quality of the resultant preparation; however, in many cases, the details of the factors are unclear. To clearly investigate the influence of the pharmaceutical properties of petrolatum ointments, we manufactured several ointments while changing the conditions of the mixing and cooling process after melting white petrolatum. As a result, the temperature at the termination was determined to influence the pharmaceutical properties of the final product. To investigate these phenomena, each petrolatum ointment sample was examined via digital microscopy and laser Raman analysis, and the distribution of the liquid–solid parts of samples was investigated. The internal structure of the ointment sample manufactured at a mixing–stop temperature of 40 °C, the needle crystals and the spherical aggregates surrounding them appropriately coexisted, while the structure exhibited a state wherein the two were linked in a semisolid phase. Meanwhile, for the ointment sample manufactured under the lowest mixing–stop temperature of 25 °C, the liquid part and the spherical aggregates were clearly separated, indicating that the liquid part was easily separated from ointments. In addition, the distribution of the hydrocarbons among the samples was measured via GC-MS; no significant difference in chemical structure was observed. In conclusion, the internal structure of the petrolatum ointment was changed by the manufacturing conditions, and this affected the pharmaceutical properties.
Oleaginous petrolatum ointment is one of the treatment forms that is commonly used as a topical formulation. Two types of ointments are listed in Japanese Pharmacopeia,1) oleaginous and water-soluble ointments; the former are the most widely used as a pharmaceutical external preparation in Japan.1–3) The reason for this frequency of use is that oleaginous ointments possess less irritancy and superior usability (good texture/elongation) than other topical formulations (e.g., ingredients), and they also exhibit superior skin protection and anti-inflammatory pharmacological effects.3) In addition, oleaginous ointments that are produced to be topically applied to the skin are divided into the “drug dispersion type,” in which the active drug ingredient (API) is uniformly dispersed in solid-state (crystal) petrolatum, and the “drug dissolving type,” in which API is dissolved. The latter is further classified into the “liquid droplet dispersion type,” in which API is dissolved in an appropriate solvent and then uniformly dispersed in petrolatum, and the “dissolution type,” in which API is dissolved in petrolatum.4,5) Drug-dispersion-type ointments have traditionally been used for API, including steroids or immunosuppressive drugs possessing strong medicinal effects; however, recently, drug-dissolving-type ointments obtained by dissolving API droplets with glycols or medium-chain fatty acid triglyceride (MCT) and uniformly dispersing the droplets in petrolatum have also been commercialized. Petrolatum is the base for any formulation-type of oleaginous ointment, and the property changes in the petrolatum will inevitably affect the pharmaceutical properties.
Petrolatum is an oleaginous semisolid that residues when the paraffin base crude oil in petroleum crude oil is distilled and refined. Moreover, it comprises solid and liquid hydrocarbons, while its physicochemical properties exhibit poor stability and poor reactivity to light and humidity.6) These hydrocarbons are linear, branched, and cyclic; have different lengths; and appear semisolid at room temperature, while solids and liquids coexist. As is well known, the rheology of petrolatum varies depending on the composition ratio of the hydrocarbons,7–14) and the plasticity fluctuates due to the inter-particle bonding force that internally forms the inter-network crystal structure. The melting point of petrolatum is between 38 and 60 °C,1) which is a wide temperature range in which the solid and liquid coexist, and the pharmaceutical properties of the petrolatum ointment are affected by the inter-particle bonding such that the string-like particles and the capillary attraction become intertwined. Recently, oleaginous petrolatum ointments have been manufactured using a variety of processes,15–22) including melting, cooling to room temperature through a mixing step, dispensing, and storing. In the manufacturing process, the cooling conditions for the molten petrolatum, that is, the cooling speed and the room temperature mixing conditions, presumably influence the pharmaceutical properties of the resultant ointments.
We previously researched the physical properties (hardness, bleeding ability, etc.) and API releasing property of Vitamin D3 petrolatum ointment containing 10% MCT and reported that these properties were greatly influenced when we altered the factors of the mix–cooling process (e.g., final mixing temperature and cooling rate to room temperature) during the manufacturing procedure.23,24) In addition, we clarified that the property changes were related to the internal structure of the ointments.
In this study, oleaginous ointment samples are produced with only petrolatum, and the property changes in the internal structure are considered to be based on the material petrolatum. The internal structure of the petrolatum ointment samples manufactured under different conditions was assessed via GC-MS, microscope observation, and laser Raman analysis. A number of new findings were obtained, which are reported below.
The petrolatum used was Japanese Pharmacopoeia (JP) white petrolatum (PF grade),1) manufactured by Kozakai Pharmaceutical Co., Ltd. (Tokyo, Japan), and standard reagents of linear saturated hydrocarbon were supplied by GL Sciences Inc. (Tokyo, Japan). The solvents and the other reagents used for the physicochemical assessment were commercially available special grade solvents or those suitable for the manufacturer’s standards.
Ointments Manufacturing MethodA high-speed mixing vacuum emulsifying machine (Agi-homo-mixer: Model 2M-2/5, PRIMIX Co., Ltd., Hyogo, Japan) equipped with a high-speed shearing homo-mixer as well as a paddle mixer for total mixing and a jacket structure container with an attached temperature sensor were used for manufacturing the petrolatum ointment samples on a 5 kg scale. The manufacturing flow of the samples is shown in Fig. 1. First, white petrolatum (material) was heated and melted at 80 °C and was kept at a product temperature of 65 °C under reduced pressure. After mixing for 15 min with the paddle mixer at 30 rpm and the homo-mixer at 2000 rpm (mixing process), only the homo-mixer was stopped. In the subsequent mix–cooling process, the petrolatum sample (intermediate product) was cooled using a paddle mixer (30 rpm) until the product temperature reached 25 °C at the cooling speed of −50 °C/h. In parallel, at the point where the product temperature reached 65 or 40 °C, the ointment sample was collected/dispensed into a container before being cooled to a temperature of 25 °C at a cooling speed of −7.5 °C/h.
Physical property measurements for the petrolatum ointment samples were performed in terms of rheometer hardness, discoloration (bleeding ratio), and GC-MS. Furthermore, to evaluate the internal state of the samples, observations via digital microscopy and Raman imaging were conducted. The experimental numerical values obtained were all expressed as mean ± standard deviation (Mean ± S.D.). The difference in the mean values between the two groups was determined using the Student’s t-test at a significance level of 5%.
Measurement Method for Rheometer HardnessApproximately 8 g of each petrolatum ointment sample was dispensed into a plastic container and then stored in a thermostatic chamber at 25 °C for 12 h. The hardness was measured using a rheometer (CR-3000-EX-L, Sun Scientific Co., Ltd., Tokyo, Japan), with the conditions of Adapter No. 25 (acrylic cylinder diameter = 20 mm; measuring speed = 120 mm/min, entrance distance = 2.5 mm). After bringing the adapter into contact with the sample surface, the adapter was advanced at a 2.5-mm depth from the surface, and the load (rheometer hardness: N) applied against the adapter was measured (n = 3).
Measurement Method for Bleeding AbilityApproximately 10 g of each petrolatum ointment sample was poured into a bleeding cone (100 meshes, opening clearance = 149 µm) and then placed in a 30 mL glass beaker (the mass of which was measured beforehand; see Fig. 2). After two weeks of storage at 25 °C/60% relative humidity (RH), the mass of the glass beaker separated liquid component was collected and measured without the bleeding cone. From Eq. (1), the bleeding ability (bleeding ratio, %) was calculated for the separation resistance index (ease of bleeding of the liquid portion, n = 3).
 | (1) |
An appropriate amount of each petrolatum ointment sample was dissolved/diluted in hexane to provide a test solution, which was analyzed using a gas chromatograph mass spectrometer (QP2010 Plus, Shimadzu Corporation, Kyoto, Japan). For the standard solutions, linear saturated hydrocarbons (single product; C17–C40, a total of 24 standard reagents) diluted in hexane were used. The measurement conditions used for the GC-MS were as follows: column = DB-1 (Agilent Technologies Japan, Ltd., Tokyo, Japan), injection mode = split-less, oven temperature = 60 °C (hold = 2 min)→heating (15 °C/min→200 °C; hold = 0 min)→heating (3 °C/min)→350 °C (hold = 10 min), injection volume = 2 µm. Meanwhile, the mass spectrometry conditions were as follows: measurement mode = SIM, CH m/z: 69, 83, 85, 97.1, and 111.1.
Internal Observation via a Digital MicroscopeAn appropriate amount of each petrolatum ointment sample was dispensed into a slide glass, and internal observations were performed using a digital microscope (VHX-1000, Keyence Corporation, Osaka, Japan). The magnification was conducted with the condition of 1000 (n = 3).
Raman Imaging via Laser Raman AnalysisAn appropriate amount of each petrolatum ointment sample was dispensed into a slide glass, and, using a Raman analyzer (RAMANtouch, Nanophoton Corporation, Osaka, Japan), the Raman spectra of the sample surfaces were measured under the following conditions: excitation wavelength = 532 nm, diffraction grating = 300 g/mm, magnification = 20× and 50× (two conditions). The standard reagents of linear saturated hydrocarbon were measured using the same method and conditions, with liquid samples transferred into small insert containers and sealed with parafilm for measurement. From the obtained Raman spectrum, Raman imaging was performed by binarizing the microscopic image (coloring = red/green) based on the peak intensity at a Raman shift of 2750–3000 cm−1.
Figure 3 shows the results of the rheometer hardness and the bleeding ratio of each petrolatum ointment sample manufactured by varying final mixing temperatures of the paddle mixer in the mix–cooling process. We previously reported the relationship between final mixing temperature and rheometer hardness/bleeding ratio in relation to petrolatum ointment containing 10% MCT.23) To more precisely clarify the physical properties of petrolatum ointment, the petrolatum ointment samples in the current report were manufactured using only material white petrolatum. However, the results were almost the same as those previously reported, that is, the rheometer hardness of the petrolatum ointment sample mixed at a final temperature of 40 °C was the highest, that of the sample mixed at a final temperature of 65 °C was the second highest, and the sample where the mixing was halted at 25 °C was the lowest. Note that this relationship (Fig. 3a) demonstrated no significant change, even in the samples stored at room temperature for eight months (data not shown). The manufacturing condition wherein the petrolatum ointment with the highest hardness was produced was that where the final mixing temperature of the paddle mixer in the mix–cooling process was 40 °C; the resulting rheometer hardness (1.5–2.0 N) was within an appropriate range in terms of texture usability.
Furthermore, the results for the bleeding ratio (Fig. 3b) were affected by the final mixing temperature of the paddle mixer. The bleeding ratio of the sample with a final mixing temperature of 40 °C was the lowest, while it was slightly higher for the sample with a final mixing temperature at 65 °C, with the highest ratio exhibited by the sample with a final mixing temperature of 25 °C. As shown in Figs. 3a and 3b, a descending relationship exists between rheometer hardness and bleeding ratio, indicating that the bleeding ability of the liquid portion in the petrolatum ointment samples was closely correlated with the hardness. With regard to the pharmaceutical properties of oleaginous ointments, the bleeding ratio should preferably be small. In this study, the manufacturing condition for obtaining the lowest bleeding ratio is considered to be that used to produce the ointment sample with the highest rheometer hardness, that is, where the final mixing temperature of the paddle mixer was 40 °C in the mix–cooling process. The results for the material petrolatum (the untreated manufacturing process), which are shown in Fig. 3a, were used for comparative control. The hardness and bleeding ratio data for the untreated material petrolatum were closest to the results for the 65 °C final mixing temperature sample.
While few studies have addressed the relationship between the manufacturing conditions of semisolid preparations and their pharmaceutical properties, the research on the solidifying ability of oil wax gels reported that the crystal phase formation in the manufacturing process depends on the hardness of the preparation manufactured, and this is related to the internal structure changes in terms of the load.25–27) Given the results of our experiments, to obtain physically stable petrolatum ointments with a high hardness and a low bleeding ratio, stopping the mixing at a temperature close to 40 °C, the lower limit of the melting point in the mix–cooling process, is important, considering the wide-ranging melting point of white petrolatum (38–60 °C).
MS Evaluation via Gas ChromatographyFigure 4a shows the mass spectrometry measurement results obtained via GC-MS in terms of the hydrocarbons/alkanes constitution (composition ratio) of each petrolatum ointment sample manufactured under each manufacturing condition with varying final mixing temperatures. The vertical axis (y-axis) of Figs. 4a and 4b are presented according to the calculated respective composition ratio (%) from the measured relative area data of the carbon chain (alkane) from C17 to C40 using linear saturated hydrocarbons. These figures clearly show that no significant differences/changes are present in the distribution of the hydrocarbons among the manufactured samples with different final mixing temperatures. Furthermore, no change was observed in comparison with untreated white petrolatum. Thus, this result clearly shows that no significant difference is present in the composition ratio and the distribution of each alkane in each petrolatum ointment sample manufactured through the melting, stirring, and cooling processes. Also, the distribution tendency of these hydrocarbons was consistent with the results of other analysis reported of raw materials of petrolatum.28)
Following this, a bleeding test was performed on the sample manufactured with a final temperature of 25 °C; the semisolid part on the bleeding cone and the bleeding liquid part that had passed through the cone were analyzed, while the alkane components were determined via GC-MS. Figure 4b shows the results of the measurements of the composition ratio of the hydrocarbons of their sample’s parts, which were compared with the same parts of the non-treated white petrolatum material. The figure clearly shows that the distribution of the hydrocarbons between the semisolid residing on the bleeding cone and the bleeding liquid part was different. The paraffin composition of the semisolid component on the cone with a distribution of higher alkanes with 26–32 carbon atoms was higher than that with lower alkanes. However, in the bleeding solution, the composition ratio and the distribution of the comparatively low alkanes with less than 25 carbon atoms were high. An alkane distribution difference between the sample with a final mixing temperature of 25 °C and the non-treated white petrolatum was not observed.
Manufacturing Conditions and Internal Structure of Petrolatum Ointment1. Appearance Observation Using a Digital MicroscopeAs shown in the previous section, the manufactured petrolatum ointment samples have different hardness and bleeding ratios depending on their manufacturing method/conditions. Therefore, to investigate the factors that differed in the pharmaceutical properties of the petrolatum ointment samples manufactured under different conditions, the state of the internal structure of each sample was observed using a digital microscope. Figure 5 shows the microscopic observations of the samples manufactured by changing the final mixing temperature of the paddle mixer in the mix–cooling process under the same cooling speed condition (−7.5 °C/h). In addition, the microscopic observations of the untreated material petrolatum are shown as a control. In these photographs, needle-like crystals could be observed in the sample with a final mixing temperature of 65 °C, which was also the case with the untreated petrolatum. Meanwhile, in both the samples with final mixing temperatures of 40 and 25 °C, a state wherein needle crystals coexist with spherical aggregates was observed. The petrolatum ointment samples differed in terms of the internal state due to the difference in the final mixing temperature: as the final mixing temperature decreased, needle crystals scattered (65 °C) and became spherical agglomerates (40 °C). Then, the needle crystals were incorporated into the spherical aggregates, and they became the dominant part (25 °C). In the 40 °C sample, the needle crystals and the spherical aggregates surrounding the crystals moderately coexisted in the liquid portion. Furthermore, the spherical aggregate formation was clearer in the sample with the final mixing temperature of 25 °C, after the mix–cooling process was performed up to the melting point temperature of petrolatum. This internal structure is thought to exist because the melted petrolatum was manufactured by mixing after transitioning from a molten state to a semisolid state in the cooling process. The reason why the sample with the final mixing temperature of 40 °C had the highest rheometer hardness and the lowest bleeding ratio is because the mix–stirring procedure was stopped near the lower limit (38 °C) of the melting point of petrolatum, at which point the petrolatum was in a semisolid state and spherical agglomerates formed in the solidified state alongside the needle crystals were observed in the molten state by mixing. This spherical aggregate coexistence (Fig. 5, 40 °C) presumably formed a kind of network structure, resulting in a stronger internal structure.
As described above, the factors that influenced the physical properties (rheometer hardness and bleeding ability) of our petrolatum ointment samples were determined using digital microscopic observation to ascertain how the internal structure of petrolatum differs. To analyze the internal crystallization and the network structure of petrolatum in more detail, Raman imaging was conducted as detailed below.
3. Raman Spectrum of Linear Saturated Hydrocarbon Using Standard ReagentsTo establish the Raman imaging method for the petrolatum ointments, we followed the approach outlined below. First, we visually observed the linear standard hydrocarbons that constitute petrolatum, the standard characteristics of linear saturated hydrocarbons from C6 to C40. Here, the linear hydrocarbons from C6 to C17 were in a liquid phase, while those from C18 to C40 were in a solid phase on the pure hydrocarbon. Moreover, the boundary where the property changes from a liquid to solid phase was found to lie between the carbon numbers C17 and C18. Then, the Raman spectrum between 500 and 3500 cm−1 of the standard product of straight-chain saturated hydrocarbons from C6 to C40 was measured, and two characteristic peaks (Peak A = around 2800 cm−1 and Peak B = around 2900 cm−1) were confirmed.
Figure 6a shows the spectrum around Peak A and Peak B for a total of six standards of three types of liquid and solid hydrocarbons. The figure shows that in C10, C14, and C17, which pertain to the lower hydrocarbons exhibiting a liquid phase, the intensity of Peak A was higher than that of Peak B. Meanwhile, in C40, C30, and C18, which pertain to higher solid hydrocarbons, the intensity of Peak B was higher than that of Peak A. The number of carbons in which the intensity ratio of Peak A and Peak B was reversed clearly occurred between C17 and C18, which coincided with the boundary of the liquid–solid phase obtained via visual observation. The multiple peaks appearing near 2800–2900 cm−1 are derived from the expansion and contraction vibration of the CH group of the hydrocarbon,29–31) but depending on the state of solid or liquid, it affects the bonding state due to intermolecular interaction, and the expansion and contraction frequency changes. Therefore, it is considered that the vibration intensity is different between the solid state and the liquid state, and the peak intensity ratio is different.
Material petrolatum is a mixture of linear, branched, and cyclic hydrocarbons with different numbers of carbons.6) We examined whether the visual observation analysis was correlated with the liquid and solid phase determination based on the above Raman spectroscopic analysis using the comparison of the Peak A intensity and the Peak B intensity. The material petrolatum was prepared by storing it for two weeks under the conditions of 25 °C/60% RH using a bleeding cone (see Fig. 2). Then, the bleeding liquid under the cone (separation liquid part: Sample 1) and the residue on the cone (separation solid part: Sample 2) were obtained. Following a visual evaluation of the appearances of the two samples, Sample 1 clearly exhibited a liquid phase, while Sample 2 exhibited a solid phase. Therefore, the two samples were subjected to Raman spectroscopic analysis; Fig. 6b shows the representative spectra of Peak A and Peak B. From these spectra, Peak A with a higher intensity than Peak B was observed in Sample 1 (bleeding liquid from petrolatum, Fig. 6b-1). Meanwhile, in Sample 2 (residue; bleeding liquid removed), Peak A with a lower intensity than Peak B was observed (Fig. 6b-2). These results were in agreement with the results of the saturated linear hydrocarbon standards (C6–C17) for the liquid phase and the standards (C18–C40) for the solid phase. Consequently, the discrimination of the appearance property (liquid–solid phase) of petrolatum could be determined via analysis of the intensity ratio of Peak A and Peak B observed between 2800 and 3000 cm−1 from the Raman spectra.
Next, Raman imaging analysis (microscopic color mapping setup) of the surface of the bleed tested Sample 1 and Sample 2 was conducted via microscopic mapping measurement, and a color visualization of the appearance was also performed. The surface area where the peak intensity ratio of Peak A and Peak B obtained from the respective sections of the Raman microscopic photographs of the two separated samples was greater than 1 (Peak B > Peak A) was colored green, while the surface area where the peak intensity ratio was less than 1 (Peak B < Peak A) was colored red. Using these color mapping operations, the peak intensity ratios (ratio [B/A]: 2750–3000 cm−1) were image-analyzed from the peak data related to the bleeding liquid (Sample 1) and the residue on the cone (Sample 2). The results are shown in Fig. 7. Sample 1 was color-coded in red (liquid), while in Sample 2, many portions of higher intensity were present (Peak B), and the surfaces were color-coded in green (solid). In addition, almost the same areas of (Peak B = Peak A) were scattered on their surface, with these portions color-coded in black, which represented the boundary of both phases assumed to be a semisolid area.
The appearance of each petrolatum ointment sample manufactured under different conditions (final mixing temperature: 65, 40, and 25 °C) and untreated material petrolatum, as a control, were visually evaluated. The four petrolatum ointment samples were oleaginous semisolids containing a liquid portion, and no significant difference in appearance was observed. Figure 8 shows the results of the color-coding via Raman spectroscopy and microscopic Raman observation for the four petrolatum ointment samples. We also attempted to comprehensively clarify the internal structure of the sample ointments by combining these results with the digital microscope photographs (Fig. 5).
In the sample with the final mixing temperature of 65 °C, the surface was almost a completely green color-coded area, where the solid portion occupied most of the sample, and a slightly red liquid portion was observed. In addition, this petrolatum ointment sample had an internal structure in which the liquid petroleum was scattered/dispersed in the solid part, much like with the untreated material petrolatum. Material petrolatum is produced and commercialized in a solid state through a molten process performed at 60–80 °C before being placed in containers without a high sheer mix–cooling process. Therefore, sample with the final mixing temperature of 65 °C was considered to have a similar internal structure because the mix–cooling process had not been performed within the vicinity of the melting point at which petrolatum begins to solidify. Since both the sample with the final mixing temperature of 65 °C and the material petrolatum had internal structures that captured the red color-coded liquid portion within the mostly green color-coded parts of the solid portion, the physical properties of high hardness and low bleeding ratio were thought to be obtained. Meanwhile, in the sample with the final mixing temperature of 25 °C manufactured with a low final mixing temperature, a green solid portion corresponding to the spherical aggregate was clearly identified in the red-colored liquid portion. The color-coded photographs from the Raman imaging in Fig. 8 show that the spherical aggregates observed in the digital microscopic photographs (Fig. 5) were solid portions. In addition, the sample with the final mixing temperature of 25 °C was in a state in which the spherical aggregates (solid phase) were comparatively separated in the liquid portion, and the structure entanglement between the two phases (liquid and solid) was demonstrated to be small. Therefore, the liquid portion was easily separated in the petrolatum, and a petrolatum ointment sample with low hardness and high bleeding ability was manufactured. The sample with the final mixing temperature of 40 °C appeared different from the other two manufactured samples. The results of the previous digital microscope (Fig. 5) indicated that spherical aggregates localized needle crystals both inside and around; however, in the Raman imaging, the spherical aggregates (solid phase: green) and the liquid portion (red) were color-coded, and the semisolid (black) phase, which is a liquid–solid boundary, was observed. Thus, both the liquid and solid parts were joined by this semisolid part (color-coded black). Based on the observation of the Raman imaging, the inner network structure of the needle crystals and the spherical aggregates discussed above indicate that the liquid part and the spherical aggregates were linked in a semisolid phase and that the structure comprised captured liquid and solid phases in the inner network of petrolatum ointment.
In GC-MS result (Fig. 4a), there was no difference in the composition ratio of hydrocarbons in each sample, however in Raman imaging, there seemed to be a difference between those samples. It is considered the distribution and existence state of each hydrocarbon are different. That is, in the bleeding test, it was confirmed that the liquid component exuded from the solid part contained in petrolatum (Fig. 3b), however this liquid component also contained more long-chain solid hydrocarbons (Fig. 4b). Then, in Raman imaging (Fig. 8b) of the 65 °C sample, the solid part and the liquid part coexist in a disorderly manner, and some of the liquid components are contained within the solid component, so that it is green part (solid phase) was mainly observed. On the other hand, in the sample (Fig. 8d) in which stirring was continued until the temperature reached 25 °C, long-chain hydrocarbons agglomerated and part of them dissolved/leaked in the liquid part during the stirring process, so that both parts were separated. Namely, the liquid part (red colored) was present as a continuous phase around the solid part (green colored), and the liquid part mainly observed.
Based on the above consideration, in the sample with the final mixing temperature of 40 °C, a higher hardness and lower bleeding ratio was obtained compared to the sample with the final mixing temperature of 65 °C, which largely comprised the solid portion. Differences occurred in the hardness and bleeding ratio of the petrolatum ointment samples due to the following reason: the manufacturing conditions (final mixing temperature) differed in the mix–cooling process in which petrolatum solidifies, which caused different distribution of the parts constituting the ointment (e.g., the solid, liquid, and semisolid parts).
The following points were clarified regarding the relationship between the internal structure of the petrolatum ointment samples and their pharmaceutical properties as well as the influence of the difference in the final mixing temperature in the mix–cooling process.
We would like to thank the researchers from the CMC departments of Maruho Co., Ltd., Kyoto R&D Center for their assistance with the practical experiments.
The authors declare no conflict of interest.
