Particle size is an important detection index for nano-pharmaceutical preparations. All nano-pharmaceutical preparations must be tested for particle size and particle size distribution, including nano-emulsion, nano-crystals, nano-particles, and nanoplex. There are many particle size measuring instruments, including nanometer laser particle size measuring instruments and microscopes (such as transmission electron microscope and scanning electron microscope) based on dynamic light scattering technology.
Electron microscope technology
Transmission electron microscope (TEM) and scanningelectron microscope (SEM) are versatile electronic microscopy instruments. Theyare intuitive methods for particle size observation and measurement, and havehigh reliability. The size and shape of the nanometer drug delivery system canbe observed with an electron microscope, the particle thickness can beestimated based on the contrast of the image, and statistics can be combinedwith image analysis to give a particle size distribution. If the particles areembedded and sliced to make thin samples, the microstructure inside theparticles can also be analyzed.
In the electron microscope measurement, it should be notedthat: 1)The measured particle size may be the size of the aggregates, so whenpreparing the nanoparticle SEM sample,it should be fully dispersed; 2) The measurement result is not statisticalbecause the amount of the electron microscope sample is very small. As aresult, the particles in the observation range are not representative; 3) Theresult observed by electron microscope is particle size rather than grain size.
Dynamic laser scattering
Dynamic laser scattering (DLS), also known as photoncorrelation spectroscopy (PCS), is the most widely used method for analyzingparticle size of nanoparticles. This method obtains particle size informationby measuring the diffusion coefficient of nanoparticles in a liquid. Whennanoparticles are dispersed in a solvent, the particles diffuse in the solventdue to the Brownian motion of the nanoparticles. The velocity of the Brownmoving particles is related to the particle size, which is consistent with theStokes-Einstein equation: d (H) = kT / 3 πηD. In the formula, d (H) is theparticle size; k is the Boltzmann constant; T is the thermodynamic temperature;η is the viscosity; D is the diffusion coefficient.
According to the viscosity η of the solvent (dispersionmedium) and the dispersion temperature T, the particle diameter d can beobtained by measuring the diffusion coefficient D of the nanoparticles in thedispersion. The laser diffraction particle size analyzer is more accurate forsamples with a particle size of more than 5 μm; the dynamic light scatteringparticle size analyzer is accurate for nano and sub-micron particle sampleswith a particle size of less than 5 μm. In this method, it should be noted thatthe best particles are spherical and monodisperse. In fact, the measuredparticles are mostly irregular and polydisperse. The particle shape andparticle size distribution characteristics have a greater impact on theparticle size analysis results, and the more irregular the particle shape andthe wider the particle size distribution, the larger the error in the particlesize analysis results. Laser particle size analysis has the advantages of smallsample consumption, high degree of automation, fastness, good repeatability,and online analysis. Its disadvantage is that it limits the concentration ofthe sample and makes it difficult to analyze the particle size and particlesize distribution of the high concentration system. At present, there areadvanced instruments to relax the concentration range, but the sample withlower concentration is still more accurate than the sample with higherconcentration due to the small interparticle interference. When using a laserparticle size analyzer, you must have an understanding of the system particlesize range, otherwise the results may be biased.
Small-angle X-ray scattering method
Small-angle scattering refers to the phenomenon of coherentscattering near the reciprocal lattice origin (000) node in X-ray diffraction.Small-angle X-ray scattering (SAXS) technology can study a variety of particlesin the range of several nanometers to hundreds of nanometers. Analyzing thesmall-angle scattering pattern can obtain information on the long-periodstructure of the substance, or the shape, scale, or mass information of thesubmicron particles (or pores). Small-angle scattering is an extremely powerfultechnique or tool for analyzing the spatial correlation of diffuse objects,such as polymer chains and macromolecules in solution.
Specific surface area method
According to the specific surface area Sw of the unit masspowder, the diameter of the particles in the nanopowder can be calculated(assuming that the particles are spherical). The general measurement method ofSw isthe BET multilayer gas adsorption method. Nitrogen is the adsorbent most commonlyused by the specific surface area (BET) method, and the specific surface arearanges from 0.1 to 1000m2/g.The advantage of this method is that the equipment is simple and the test speedis fast, but it is only the specific surface area information of thenano-powder. After conversion, the average particle size is obtained, but theparticle size distribution cannot be understood.
Atomic force microscopy
Atomic force microscopy (AFM) scans the surface of a sampleby a tiny probe to convert the interaction between the probe and the surface ofthe sample into a surface topography and characteristic image. Its advantage isthat it can provide a three-dimensional and high-resolution image of thesurface, and has a high horizontal and vertical resolution. In addition tomeasuring the particle size, it can also describe the sample shape. Itsdisadvantages are small sample observation time and time consuming. Similartechnologies to AFM include scanning transmission microscopy (STEM) andscanning transmission x-ray microscopy (STXM).
Selection of particle size measurement techniques
Nano drug delivery systems consist of a series of particles of varying sizes. Depending on the purpose of the measurement, the overall particle size distribution can be expressed by the number or volume diameter. The particle size distribution is determined not only by the average particle diameter, but also by the method of evaluating the average particle diameter. In addition, the particle distribution shape greatly affects the result of particle distribution. The influence of large particles on the calculation results in DLS measurement is very significant, and the scattered light of core-shell double-layer particles and single-layer particles with the same particle size distribution is different at different wavelengths. In practical applications, generally based on the actual particle size range, the detection method is reasonably selected, and two or more methods can be used to separately measure and verify each other. The triptolide nanoparticles were characterized by TEM and AFM at the same time. It was found that the structure of the nanoparticles was round and the surface was smooth.
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