Report
Nanoparticle Detection Technology
T
he formation of clouds, fog, and rain droplets in the atmosphere does not occur directly by condensation of water molecules onto each other (homogeneous nucleation), but rather by condensation onto so-called Aitken nuclei, which are ttny aerosol particles always present in the atmosphere (2). Particles as small as a few nanometers in diameter may serve as nuclei for this heterogeneous nucleation and, because droplets can grow as large as raindrops the process may be regarded as a high-gain (but nonlinear) mass amplification As the droplets grow from diameters much smaller than visible light wavelengths (0 35-0 77 urn) to micrometer and larger diameters an equally
dramatic increase in the litrht scattering effiHenrv (the formation of a visible fog or clouds) occurs
process has been used to develop new methods of chemical analysis. Nucleation is largely independent of particle composition and, therefore, can provide little specific chemical information. However, coupled with a separation technique that provides a satisfactory level of chemical selectivity, techniques based on condensation nucleation principles could provide highly universal responses for even single molecules without the need for fluorescent radioactive or other labeling or derivatization. Condensation nucleation light scattering detection (CNLSD) electrospray with condensation nucleation particle counting (ES-CPO and gas-phase electrophoretic mobility molecular analysis
liquid-phase substance at T. S 1 for supersaturated vapor. In terms of the molecular number vapor concentration, S = N/N0, in which N is the actual molecular number vapor concentration in the carrier gas, and N0 is the molecular number vapor concentration in a saturated vapor at equilibrium. For a given supersaturation S, particles larger than DK nucleate droplet formation. In practice parameters, such astimespent in the vapor and details of the flow also influence the size dependence of the particle detection probability (4 5). A detection efficiency for a commercial ultrafine CPC is shown in Figure 1 D approximates the zero crossing point of the detection effi-
(GEMMA) have begun to realize this potential and are the suhiect of this Report
ciency curve As indicated in Fnuation 1 D decreases QO fVie local S increases However at ^nffirtpntW high S homogeneous nucleatinn can nrrur anrl Qiirn rnndih'nns art*
Growth by condensation nucleation, followed by light scattering, is the basis of a stan- Fundamentals dard method for measuring aerosol concentra- To characterize the mass transfer of vapor tions dating back to Aitken'sttme(2)) In mod- during the condensation nucleation proern instruments, this condensation nucleation cess, both condensation and evaporation process is used to count individual aerosol must be considered. From purely physical particles that have diameters as small as 3 nm, factors, the vapor pressure increases for the approximate size of a single molecule of a droplets and particles of decreasing diamesmall protein or of an aggregate of 30 sucrose ter. The diameter at which the condensaor 500 sodium chloride molecules. Viewed as tion and evaporation rates are equal (3), a "mass amplifier" this growth process repre- also known as the Kelvin equivalent diamesents astronomical gains—on the order of ter (/^K) , is given by 1010 and Recently, the condensation nucleation DK = 4Mo/RpTlnS (1) John A. Koropchak Salma Sadain Xiaohui Yang Lars-Erik Magnusson Mari Heybroek Michael Anisimov Southern Illinois University
Stanley L. Kaufman TSI, Inc. 386 A
Qwnirl^H tr» e n c n r e t h a t signal prnrlimn'rm is
The overall process of detection used with all three methods is depicted in Figure 2. The aerosol process begins with the conversion of a flowing liquid stream into aerosol droplets by a nebulizer. These solvent-laden droplets are then dried (desolvated) to remove the solvent from the particle phase (the solvent molecules are too small to serve as condensation nuclei and thus will not be detected). There are two ways to obtain concentrain which M, a, and p are the molecular tion-dependent response from the condenweight, surface tension (erg/cm2), and den- sation nucleation process. In the first sity (g/cm3) of the condensing fluid, respec- method, referred to as CNLSD, or the concentration-dependent particle size approach, tively; Tis the nucleation temperature in degrees Kelvin; R is the gas constant (8.3144 one can use the dependence of the dry resix 107 erg/K mole); and S is the vapor satura- due particle size on the analyte concentration given by tion ratio (P/P0), in which Pis the actual partial vapor pressure and P0 is the saturation vapor pressure in equilibrium with a Z)D =Z),(C/p) (2)
Analytical Chemistry News & Features, June 1, 1999
for Chemical Analysis in which DD is the diameter of the dried residue (desolvated) particle, Dx is the initial droplet diameter, C is the nonvolatile solute concentration (g/cm3), and p is the solute density (g/cm3). Generally, the wet aerosols are at least somewhat polydisperse (consisting of a range of sizes), and the dry size distributions are identical to the wet size distributions but scaled to smaller size, as shown in Equation 2. Because the concentration of the analyte affects particle size and particle size influthe detection efficiency of the densation nucleation process the response will be concentration-dependent The Dl values of the nebulizer clearly influence this process as well. Typically, pneumatic nebulizers coupled to an impaction plate system for droplet size reduction have been used for high sample-flow systems, while ES aerosol generation with charge neutralization is best suited to the flows and requirements of capillary separations. Furthermore, the size distributions are influenced by other processes on transport from the nebulizer to the CPC. Particles may diffuse to surfaces and be lost. This diffusion loss is strongly size-dependent as shown in the example of the screen penetration curve in Fitrure 1 The overlap of the size distribution which reaches the CPC with the detection effiHenrv m r v e determines the signal level
Equation 2 assumes that the mass of the nonvolatile solute is uniformly distributed through the initial solution phase, or, equivalency, that each droplet contains a large number of solute molecules. For sufficiently dilute solutions and/or small droplets, the number of analyte molecules in each droplet will be a small integer, and this assumption will not be valid. In this case, ,he dry size distribution will have a discrete Poisson dis-
When coupled with a selective separation technique, condensation nucleation could provide highlyuniversalresponse without the need for labeling orderivatization. tribution in which each macromolecule is an independent dry particle (6). If these macromolecules are large enough to act as nuclei for condensation and detection, the individual molecules can be counted to provide a second means for generating a concentration-dependent response. For the detection
efficiency curve in Figure 1, molecular weights greater than 5-10 kDa are required to provide individual particles of sufficient size (—3-4 nm) for efficient detection. We will refer to this as the macromolecule counting approach or ES-CPC detection. When the condensation process onto a
Analytical Chemistry News & Features, June 1, 1999 387 A
Report particle is initiated, the rate of mass accretion accelerates as the surface area increases, until growth is either slowed by local vapor depletion or stops when the particle leaves the condenser. In a typical instrument design, the final droplet diameters only weakly depend on the initial particle size (smaller particles travel farther before reaching a region of sufficient supersaturation to begin growth and thus spend less time growing) (7). Each droplet that is grown passes through a light beam (usually from a laser) and in doing so will scatter a nearly equivalent burst of light that be with the naked if visible light is used as the source beam The average intensity of this light scattering from the grown droplets can be monitored, or the pulses from individual droplets can be counted. In the latter case, the pulse rate and the known flow rate together yield the absolute number of particles larger than the design threshold diameter (e.g., ~DK) per unit volume. Commercial devices based on these principles are CPCs and are used, for example to determine aerosol particle
number densities in the atmosphere, clean rooms, and other environments. For CNLSD, it is useful to note that, with a fixed mass of analyte, a reduction in particle size of an order of magnitude increases the number of particles by a factor of 1000. Because volume (or mass) is a cubic function of diameter, if all of these particles were detected, then the sensitivity would increase 1000-fold in the smaller particle case. As a result smaller particles and thus smaller nebulizer droplets are generally advantageous for CNLSD. A caveat to this concept is that as particle sizes decrease diffusion coefficients increase in the low nanometer-size regime diffusion coefficients approach the values for molecular diffusion coefficients For sufficiently small particles this process can substantially influence the system response Although early condensation nucleationbased instruments generated supersaturation conditions by expanding discrete test volumes (2), the last two decades have seen the development of continuous-sample-flow CPCs, in which the aerosol flow is sampled and merged with a secondflowof gas, which
has passed over a heated reservoir of fluid and becomes saturated in the vapors of the fluid. The mixture becomes supersaturated when cooled in a condenser; the temperature difference between the heated saturator and the condenser establishes S. Typically, CPCs are operated with a temperature difference that provides S sufficient for heterogeneous nucleation but not for homogeneous nucleation. The continuous-flow CPCs described in the literature have been shown to respond to particles as small as 2 nm (5) and they provide a convenient means for continuous monitoring offlowinggas streams or dried aerosols produced from liquid streams whose composition varies with time Once initiated, condensation nucleation should be independent of the particle composition because further condensation occurs on a surface of the condensing liquid. Approximate composition independence for nucleation has been confirmed experimentally (8-10), although such studies have focused on a narrow range of materials. Nonetheless if one assumes substance-independent responses, then it is possible to design nebulization-desolvation CPC systems for monitoring extremely low levels of total dissolved solids in high-purity solvents (11-14) CNLSD, with discrete samples and flow injection analysis, was later described, and detection limits below the nanogram-permilliliter level were reported (15). However, as indicated in the introduction, to obtain more specific chemical information, all of these methods must be combined with a separation technique. The remainder of this Report will describe condensation nucleation for detecting chemical species separated by standard chemical separations before the spraying process or by gas-phase electrophoretic mobility after the spray.
Condensation detection methods
Figure 1 . The detection efficiency for a CPC, the typical penetration efficiency curve for a diffusion screen, and the typical electrospray particle size distribution for dried electrospray droplets. Curves can differ significantly depending on the instrument model. 388 A
Analytical Chemistry yews & Features, June 1, 1999
CNLSD and ES-CPC have been used for detecting a wide range of substances separated by reversed-phase, ion-exchange, size-exclusion, and normal-phase chromatographies; CE; electrochromatography; and even by supercritical fluid chromatography. One direction for this work has been the development of CNLSD as a sensitive,
Figure 2. Basic processes of CNLSD, ES-CPC, and GEMMA. The liquid to be analyzed (GEMMA) or the effluent of an analytical separation system (CNLSD or ES-CPC) is dispersed into droplets and dried. The resulting aerosol particles, either immediately (in CNLSD and ES-CPC) or after separation by electrical mobility (GEMMA), are detected by condensation nucleation followed by light scattering.
general-purpose, universal detector for separating a wide variety of low-volatility species, independent of molecular weight. In this case, the concentration-dependent particle size approach to quantitation is used. On the other hand, ES-CPC has focused on macromolecule detection using the macromolecule counting approach to quantitation, particularly for proteins separated by capillary sizeexclusion or reversed-phase LC or CE. Concentration-dependent particle size quantitation
The process of CNLSD can be viewed as an extension of evaporative light scattering detection (ELSD), for which the direct light scattering by the desolvated particles (DD) is monitored without the condensation step (16)) ELSD response is considered highly universal, but is generally limited to concentrations >1 ug/mL. Based on analyte concentrations of 1 ug/mL and the droplet size distributions of typical pneumatic nebulizers for ELSD, the largest dry particles expected would be only —100 nm in diameter and would be small in number. However in accordance with Mie theory light scattering efficiency drops off dramatically for particles below the wavelength of liriit utilized meaning that these particles
would be inefficient light scatterers for UV-vis photons. The growth in size because of condensation greatly increases the light scattering from each particle. The response characteristics of ELSD and CNLSD have been directly compared for the detection of polyethylene glycols separated by aqueous size-exclusion chromatography (IT). Detection limits with CNLSD were in the 15-ng/mL level on average 130 times better than those for a commercial ELSD. While the response for ELSD is typically nonlinear, the linear response range for CNLSD was at least 3 orders of magnitude, particularly when a diffusion screen (described later) was added. One limitation of CNLSD is background from nonvolatile species also present in the mobile phase. After evaporation, most organic solvents leave residues in the microgram-per-milliliter range. In reversed-phase separations, these residues can be a problem (18), and increasing the organic content of the mobile phase can heighten the background and degrade detection. Purifying the solvent provided modest improvements. However, a more general approach to background reduction is the use of diffusion screens in the aerosol flow. Diffusion screens are fine mesh screens to which
smaller particles with higher diffusion coefficients can diffuse and be collected prior to the CPC. A typical penetration efficiency curve for a diffusion screen is indicated in Figure 1. The response observed using diffusion screen (s) is given by the product of the particle size distribution, the penetration curve of the diffusion screen, and the detection efficiency curve. In Figure 1, the position of the particle peak is determined using Equation 1 and the concentration of all the nonvolatiles including the analyte ,n the llquid. Ideally with zero analyte concentration the peak would be on the left side of Figure 1 where detection is strongly suppressed by the screen response Anv increase presumably because of analyte would be deas in the detected aero
