MXPA99011840A - Method for enhancing the echogenicity and decreasing the attenuation of microencapsulated gases - Google Patents

Abstract

Se ha descubierto que micropartículas formadas de polímero natural o sintético con paredes más gruesas presentan ecogenicidad significativamente mejorada en comparación con micropartículas con paredes más delgadas. El efecto del espesor de las paredes ha sido determinado experimentalmente asícomo insertado en una fórmula para uso en la predicción de las condicionesóptimas. En la modalidad preferida, los polímeros son polímeros sintéticos, biodegradable y el espesor de la pared es entre 100 y 660 nm, aunque puede utilizarse un espesor de pared desde aproximadamente 20 nm hasta un exceso de 500 nm. Las micropartículas se fabrican con un diámetro adecuado para el determinado tejido el que se va a formar la imagen, por ejemplo, con un diámetro entre 0.5 y 8 micras para administración intravascular, y un diámetro de entre 0.5 y 5 mm para administración oral para imaginar el tracto gastrointestinal u otros lúmina. Los polímeros preferidos son polihidroxiácidos comoácido poliláctico-co-ácido glicólico, polilacturo o polilglicoluro, de mayor preferencia conjugados con un polietilén glicol u otros materiales que inhiban la captación por el sistema reticuloendotelial (SER). Las microesferas pueden ser utilizadas en una variedad de aplicaciones en formación de imágenes de ultrasonido, incluidas las aplicaciones en cardiología, aplicaciones en perfusión sanguíneas, asícomo para imágenes deórganos y venas periféricas.

Description

METHOD TO IMPROVE THE ECOGENICITY AND REDUCE THE ATTENUATION OF MICROENCAPULATED GASES BACKGROUND OF THE INVENTION The present invention is, in general, in the area of diagnostic imaging agents, and is directed particularly to microparticular contrast agents for ultrasound imaging that have increased echogenicity and decreased attenuation as a function of the thickness of the polymer membrane. When ultrasound is used to obtain an image of the internal organs and structures of a human or animal, the ultrasound waves, the waves of the sound energy at a frequency above "what is perceptible by the human ear, are reflected in accordance These pass through the body, different types of body tissue reflect the ultrasound waves differently and the reflections or reflections that are produced by the ultrasound waves reflecting the different internal structures are detected and converted electronically into a visual presentation. medical conditions, the obtaining of a useful image of the organ or structure of interest is especially difficult because the details of the structure are not adequately perceptible from the surrounding tissue in an ultrasound image produced by the reflection of the lacking ultrasound waves of a contrast enhancing agent. The physiological and pathological conditions can be substantially increased by improving the contrast in an ultrasound image by injecting or infusing an agent of an organ or other structure of interest. In other cases, the detection of the movement of the contrast enhancing agent itself is particularly important. For example, a distinct feature of blood flow that is known to result from specific cardiovascular abnormalities may be only discernible by infusing a contrast enhancing agent into the bloodstream and observing the dynamics of blood flow. Materials that are useful as ultrasound contrast agents work by having an effect on the ultra sonic waves as they pass through the body and are reflected to create the image from which the medical diagnosis is made. Different types of substances affect the ultrasound waves in different ways and to different degrees. In addition, certain effects caused by contrast enhancing agents are more easily measured and observed than others. To select an ideal composition for a contrast enhancing agent, the substance having the most drastic effect on the ultrasound wave as it passes through the body is preferred. Also, the effect on the ultrasound wave must be measured easily. There are two effects that can be observed in an ultrasound image: backscattering and beam attenuation. RE RODISPERSION: When an ultrasound wave that is passing through the body meets a structure, such as an organ or other body tissue, the structure reflects a portion of the ultrasound wave. Different structures within the body reflect the energy of ultrasound in different form and different intensities. This reflected energy is detected and used to generate an image of the structures through which the ultrasound wave has passed. The term "backscatter" refers to the phenomenon in which the ultrasound energy is dispersed back to the source by a substance with certain physical properties. For a long time it has been known that the contrast observed in an ultrasound image can be improved by the presence of substances known to cause a large amount of backscatter. When such substances are administered to a different part of the body, the contrast between the ultrasound image of this body part and the surrounding tissues that do not contain the substance is improved and contrasted. It is well known that, due to their physical properties, different substances are causative of backscattering in different degrees. Therefore, the search for contrast enhancement agents has focused on substances that are stable and non-toxic and that present the maximum amount of backscatter. The ability of a substance to cause backscattering of ultrasound energy depends on the characteristics of the substance as its ability to be compressed. When examining different substances it is useful to compare a particular measure of the ability of a substance to produce backscattering known as the "dispersion cross section". The dispersion cross section of a specific substance is proportional to the radius of the disperser, and also depends on the wavelength of the ultrasound energy and other physical properties of the substance, J. Ophir and KJ Parker, Contrast Agents in Diagnostic Ulstrasound , Ultrasound in Medicine &; Biology, vol. IS, n. 4, p. 319, 323 (1989). When assessing the usefulness of different substances of different substances as ultrasound contrast agents, ie gases, liquids or solids, it is possible to calculate which of the agents may have the highest dispersion cross section and, consequently, which agents may provide the largest contrast in an ultrasound image. It can be assumed that the compressive capacity of a solid particle is much smaller than the surrounding medium and that the density of the particle is greater. Using this assumption, the cross section of the dispersion of a solid particle contrast enhancing agent has been calculated as 1.75 (Ophir and Parker, supra, at 325). For a pure liquid dispersant, the adiabatic compression capacity and the density of the disperser and the surrounding medium are likely to be approximately equal, which would produce the result that the liquids would have an effective dispersion cross section of zero. However, liquids may exhibit some backscattering if large volumes of a liquid agent are present. For example, if a liquid agent passes from a very small vessel to a very large one so that the liquid occupies practically the entire vessel, the liquid may have edible backscattering. However, those skilled in the art will appreciate that pure liquids are relatively inefficient dispersants. The effective dispersion section of a gas is substantially different and greater than a liquid or solid, in part, because a gas bubble can be compressed to a much greater degree than a liquid or solid. In addition, free gas bubbles in a liquid exhibit oscillatory motion so that, at certain frequencies, the gas bubbles will resonate at a frequency close to that of the ultrasound waves commonly used in medical imaging. As a result, the effective dispersion section of a gas bubble can be a thousand times larger than its physical size. ATTENUATION OF THE BEAM: Another effect that can be observed from the presence of certain contrast enhancing agents is the attenuation of the ultrasound wave. The intensity of the ultrasound wave decreases as the wave passes through the volume of tissue or blood that contains the contrast agent. The decrease in wave intensity is the result of ultrasound that is backscattered by the agent as well as the dissipation of the wave as it interacts with the contrast agent. If the beam is too low, the power returned to the transducer from the distal regions for the contrast agent will be low, giving rise to a poor image depth. The use of differences in beam attenuation in different types of tissue has been tried as a method of image enhancement. The contrast of the image has been observed in conventional image formations due to localized attenuation differences between certain types of tissue. K. J. Parker and R. C. Wagg, "Measurement of Ulstrasonic Attenuation Within Regions selected from B-Scan Images ", IEEE Trans Biomed, Enar BME 30 (8), p 431-37 (1983); K. J. Parker, R. C. Wagg, and R. M. Lerner, "Attenuation of Ultrasound Magnitude and Frequency Dependence for Tissue Characterization", Radiology, 153 (3). p. 785-88 (1984). One hypothesis is that measurements of the attenuation of a tissue region taken before and after the infusion of an agent can produce an improved image. However, techniques based on the contrast of attenuation as a means to measure the improvement of the contrast of a liquid agent are not well developed and, even if they are fully developed, they may present limitations in terms of internal organs or structures with the which are techniques can be used. For example, it is very likely that a loss of attenuation due to liquid contrast agents can be observed in the cardiovascular system image due to the high volume of liquid contrast agent that would need to be present in a given vessel before it could be measured a substantial difference in attenuation. In summary, ultrasound for diagnosis is a powerful, non-invasive tool that can be used to obtain information about the internal organs of the body. The arrival of the gray scale and Doppler color imaging has greatly improved the scope and resolution of the technique. Although the techniques for performing ultrasound exams for diagnosis have improved significantly, such as the techniques for manufacturing and using contrast agents, there is still a need to improve the resolution of cardiac perfusion imaging and cardiac chambers, solid organs, perfusion renal, perfusion of solid organs and Doppler signals of the velocity and direction of blood flow during the formation of images in real time. The development of ultrasound contrast agents has focused on the use of biocompatible gases, as free gas bubbles or as gases encapsulated in natural or synthetic coating materials. To encapsulate a gas a variety of natural and synthetic polymers have been used, such as air for use as contrast agents for imaging. Schneide et al., Invest. Radiol., Vol. 27, pp. 134-139 (1992) describe polymeric particles filled with air of 3 microns. These particles were reported as stable in plasma and under applied pressure. However, at 2.5 MHz, its echogenicity is low. Another type of suspension of encapsulated ga micro bubbles has been obtained from voiced albumin.

Feinstein et al., J. Am. Coil. Cardiol., Vol. 11, pp. 59-6 (1988). Feinstein describes the preparation of microbubbles that are suitably sized for the transpulonary country with excellent in vi tro stability. However, these microbubbles have a short life in vivo, with a half life in the order of a few seconds (which is approximately equal to one circulation step) because they dissolve rapidly in the subsaturated liquids, for example blood. Wible, J. H. et al., J. Al. Soc. Echocardiogr., Vpl 9, pp. 442-451 (1996). The air bubbles encapsulated in gelatin previously described by Carroll et al. (Carroll, BA et al., Invest. Radiol., Vol.15, pp. 260-266 (1980) and Carroll, BA et al., Radiology, vol.143, pp. 747-759 (1982)), but Due to their large sizes (12 and 80 μ) these probably do not pass through the pulmonary capillaries. Gelatin-encapsulated microbubbles have also been described in PCT / US80 / 00502 by Rasor Associates, Inc. These are formed by "coalescence" in the gelatin. Microbubbles of air stabilized by galactose microcrystals (SHU 454 and SHU 508) have also been reported by Fritzsch, T. et al., Invest. Radiol. vol. 23 (Suppl 1), pp. 302-305 (1988); and Fritzsch, T. et al., Invest. Radiol., Vol. 25 (Suppl 1), 160-161 (1990). The microbubbles last up to 15 minutes in vitro but less than 20 seconds in vivo. Rovai, D. et al., J. Am. Coil. Cardiol., Vol 10, pp. 125-134 (1987); and Smith, M. et al., J. Am. Coil. Cardiol., Vol. 13, pp. 1622-1628 (1989). The gas microbubbles encapsulated within a shell of a fluorine-containing material are described in WO 96/04018 by Molecular Biosystems, Inc. The application of European Patent No. 90901933.5 to Schering Aktiengesellschaft describes the preparation and use of microencapsulated gas or liquids. volatile for ultrasound imaging, where the microcapsules are formed of synthetic polymers or polysaccharides. The application of European Patent No. 91810366.4 of Synthetics S.A. (0 458 745 A1) describes air or gas microballoons attached by an interfacially deposited polymeric membrane that can be dispersed in an aqueous carrier by injection into a host animal or by oral, rectal or urethral administration for therapeutic or diagnostic purposes. SO 92/18164 of Delta Biotechnology Limited describes the preparation of microparticles by spray drying under highly controlled conditions such as temperature, spray speed, particle size and drying conditions, of an aqueous solution of protein to form hollow spheres with gas trapped in the same, for use in the formation of images. WO 93/25242 describes the synthesis of microparticles for ultrasound imaging consisting of a gas contained within a polycyanoacrylate or polyester shell. WO 92/21382 describes the manufacture of contrast agents in microparticles that include a covalently bound matrix containing a gas, wherein the matrix is a carbohydrate. US Patents Nos. 5,334,381, 5,123,414 and 5,352,435 to Unger describe liposomes for use as ultrasound contrast agents, which include gas, gas precursors, such as a gaseous precursor activated by pH or activated photo, as well as as other liquid or solid contrast enhancers. WO 95/23615 to Nycomed describes microcapsules for imaging which are formed by coacervation of a solution, for example a protein solution, with a perfluoro carbon content. PCT / US94 / 08416 from the Massachusetts Institute of Technology discloses microparticles formed of polyethylene glycol-poly (lactide-co-glycolide) block polymers having imaging agents encapsulated therein, including co- or air gases and perfluorocarbons. Although all the ultrasound contrast agents investigated to date, such as free gas bubbles or "encapsulated gas bubbles" are powerful backscatters, these agents also have a high degree of attenuation. image formation and loss of tissue images are distant from the contrast agent In many cases, information about the image can be completely lost beyond regions that have significant concentrations of the contrast agent, for example the left ventricle. Ultrasound contrast agents currently under investigation share this problem to some extent.To minimize the problem associated with the attenuation of contrast agents, researchers have resorted to various approaches.Most frequently the amount of contrast agent administered is decreases to allow more than one beam of ultrasound penis tre through the contrast agent. Although the attenuation is lower, the decrease in dose gives rise to less than an optimal contrast for many clinical indications. Otherwise, ultrasound contrast agents can be administered as a continuous infusion. This essentially reduces the local concentration of the agent and has the problem described above for dose reduction. Continuous infusion has the additional disadvantages of requiring a larger total dose over time and is not easy to perform in a clinical setting. To compensate for reduced doses, researchers have used harmonic imaging to improve the signal-to-noise ratio. However, the formation of the harmonic image is not standard at this time. It is important to note that these approaches do not lead to the rectification of the fundamental problem with the acoustic properties of the existing ultrasound contrast agents. Thus, in order for an ultrasound contrast agent to have high echogenicity, it is necessary to create a disperser which generates a high total return power in the receiver transducer from regions of interest at depths beyond the initial region containing the contrast agent. The return power will be regulated by backscattering and attenuation of the agent. Therefore, an object of the present invention is to provide icroparticle with significantly improved echogenicity. Another objective of the invention is to provide an ultrasound agent with high backscattering and low attenuation.

SUMMARY OF THE INVENTION It has been found that thicker walled microparticles formed of natural or synthetic polymers have significantly improved echogenicity and less attenuation compared to microparticles having thinner walls. The effect of the thickness of the wall has been determined theoretically and the optimum thickness of the wall has been predicted. The microparticles that have these thicknesses were produced. In the preferred embodiment, the polymers are biodegradable, synthetic polymers, and the wall thickness is between 50 and 660 nm, although wall thicknesses of about 30 n a in excess of 800 nm can be used. The thickness of the cover will depend on the target tissue from which the image will be formed will depend on the blood volume and tissue volume of the target organ. The microparticles are manufactured with a diameter suitable for the target tissue to be imaged, for example, with a diameter between 0.5 and 8 μ for intravascular administration, and a diameter between 0.5 and 5 nm for oral administration for training of images of the gastrointestinal tract or other lumina. Preferred polymers are polyhydroxy acids, such as psi-lactic acid-co-glycolic acid, polylactide polyglycolide or polylactide-co-glycolide. These materials can be conjugated to polyethylene glycols or other materials that inhibit uptake by the reticuloendothelial system (RES). The microspheres can be used in a variety of ultrasound imaging applications including cardiology applications, blood perfusion applications as well as for imaging peripheral organs and veins.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a graph of the calculations of the effect of wall thickness on the effective cross-section of the total dispersion per unit volume as a function of the acoustic frequency for a representative size distribution of the microencapsulated octafiuoropropane in FIG. the synthetic polymer in a dilution of 1/1620 assuming wall thicknesses of 11 Q nm and 0.0034% C3F8 (fraction of the total volume of the gas), 165 nm and 0.0032% of C3F8, 220 nm 0.0029% C3F8, 330 nm and 0.0025% of C3F8, 440 nm and 0.0021% C3F8, 660 nm and 0.015% C3F8, 880 nm and 0.0010% C3F8 and 1100 nm and 0.007S of C3F8. Figure 2 is a graph of the calculations of the effect of wall thickness on the acoustic attenuation coefficient as a function of the acoustic frequency for a representative size distribution of the microencapsulated octafluoropropane in the synthetic polymer at a dilution of 1/1620 assuming thicknesses 110 nm and 0.0034% C3F8, 165 nm and 0.0032% C3F8, 220 nm and 0.0029% C3F8, 330 nm and 0.0025% C3F8, 440 n and 0.0021% C3F8, 660 nm and 0.015% C3F8, 880 nm and 0.0010% C3F8 and 1100 nm and 0.007% of C3F8. Figure 3 is a graph of the calculations of the effect of wall thickness on echogenicity (total return power per unit volume) as a function of the acoustic frequency for a representative size distribution of the microencapsulated octafluoropropane in the synthetic polymer in a dilution of 1/1620 assuming wall thicknesses of 110 nm and 0.034% C3F8, 165 nm and 0.0032% of C3F8, 220 nm and 0.0029% C3F8, 440 nm and 0.0021% C3F8, 660 n and 0.0015% C3F8. Figure 4 is a graph of the calculations of the effect of wall thickness on the effective section of total dispersion per unit volume as a function of the acoustic frequency for a representative size distribution of microencapsulated air in natural polymer at a dilution of 1 / 1620 assuming wall thicknesses of 40 nm and 0.0021% of air (fraction of total volume of gas), 80 nm and 0.0020% of air, 150 nm and 0.0019% of air, 300 nm and 0.0017% of air, 600 nm and 0.0013 % air, and 900 nm and 0.0010% air. Figure 5 is a graph of the calculations of the effect of the wall thickness on the acoustic attenuation coefficient as a function of the acoustic frequency for a representative size distribution of microencapsulated air with natural polymer at a dilution of 1/1620 assuming thicknesses of 40 ny wall and 0.0021% air, 80 nm and 0.0020% air, 150 nm and 0.0019% air 300 nm and 0.0017% air, 600 nm and 0.0013% air, and 900 nm 0.0010% air. Figure 6 is a graph of the calculations of the effect of wall thickness on echogenicity (total return power per unit volume) as a function of the acoustic frequency for a representative size distribution of microencapsulated air in natural polymer a dilution of 1 / 1620 assuming wall thicknesses of 40 n and 0.00211 of air, 80 nm and 0.0020% of air, 150 nm and 0.0019% of air 300 nm and 0.0017% of air, 600 nm and 0.0013% of air, 900 nm and 0.0010% of air. Figure 7 is a graph of the calculations of wall thickness effect on echogenicity as a function of the acoustic frequency for a representative size distribution of microencapsulated air in natural polymer at a dilution of 1/5400 assuming wall thicknesses of 15 nm and 0.0006% air, 40 nm and 0.0006% air, 80 nm and 0.0006% air 150 nm and 0.0006% air, 300 nm and 0.0005% air.

DETAILED DESCRIPTION OF THE INVENTION A method is described for maximizing echogenicity as a function of the wall thickness of natural or synthetic polymeric microparticles. Microparticles are useful in a variety of ultrasound imaging applications for diagnosis, particularly in ultrasound procedures such as blood vessel imaging and echocardiography. The increase in wall thickness significantly increases echogenicity compared to the same natural or synthetic polymeric microparticles with thinner walls.

I. Calculation of the optimal thickness of the polymer To allow a better understanding of the response of the encapsulated microbubbles for diagnostic ultrasound, a mathematical model was used (C. Church J. Acoustical Soc. Amer. 97 (3): 1510-1521, 1995) to calculate important quantities such as the backscatter and attenuation coefficients for the values of the physical parameters such as thickness and rigidity of the encapsuant cover. The cover can be a natural or synthetic material. The model consists of an equation similar to that of Rayleigh-Plesset (non-linear) for the case of spherical gas bubbles encapsulated by a cover that behaves collectively as an elastic solid, damped, continuous, incompressible. An analytical solution for this equation, which includes the first and second components of the lowest order harmonic, is used in the present to estimate the effect of the thickness of the roof on the dispersion cross section (the ratio of the power dispersed by the gas bubbles encapsulated at the intensity of the incident acoustic beam) and the attenuation coefficient (the velocity at which the gas bubbles eliminate the beam acoustic energy) from a suspension of encapsulated gas bubbles. These quantifiers are then used to estimate the total return power from a suspension of gas bubbles encapsulated to the ultrasound transducer by emitting the incident pulse. The Rayleigh-Piesset equation that describes the response of an encapsulated gas bubble to an incident acoustic pressure wave is as follows: (1) where Ri is the radius of the gas-filled cavity, Ui is the radial velocity of interface 1 (the interface between the gaseous interior and the encapsulating solid), R is the outer radius of the encapsulating material, pL is the density of the liquid that surrounds the bubble, L is the density of the encapsulating cover, PG, eq is the pressure of the gas in equilibrium within the bubble, R0? is the initial radius of the cavity filled with gas, P8 (t) is the pressure at infinity (including the acoustic activation pressure) and s2 are the interfacial tensions at the covered gas and liquid covered interfaces, respectively, μs and μ? , are the effective viscosities of the cover and the surrounding liquid, respectively, VS = R23-R? 3, Gs is the rigidity of the cover and Re? it is the equilibrium position without deformation of the gas-covered interface. An expression for the scattering cross section s = i of an encapsulated gas bubble can be found by assuming that the amplitude of the pulse Ro? X (t) is small and replacing Ri = R0? (L + x) and related expressions in Equation (i) of Church (1995) The resulting equation (2) is: (2) where w is the (radial) frequency of the incident acoustic wave, w0 is the resonance frequency of the encapsulated gas bubble and d ^ is the damping constancy of the encapsulated gas bubble; The representative units for the cross-section are given in parentheses after the equation. After the equation (2) is suitable for cases in which the response of a single encapsulated gas bubble is of interest. In diagnostic ultrasound it is more common to be interested in the responses of a suspension of many millions of encapsulated gas bubbles. When a collection of gas bubbles encapsulated with a range of sizes is present, the effective section of the total dispersion per unit volume can be estimated simply by adding the contribution of each gas bubble encapsulated in a volume representative of the suspension: (3) where f (R0?) dR0? Is the number of gas bubbles encapsulated per unit volume with radii between RQ? and Role + dRQ? . The attenuation coefficient of the suspension can be estimated using the method provided by K. W. Commander and A. Prosperetti, "Linear Pressure waves in bubbly liquids: Comparison between theory and experiments", J. Acoust. Amer. 85 (2): 732-746 (1989). When describing a medium full of bubbles in terms of suppression, density, average speed, etc., these authors obtained an expression for cm, the complex speed of sound in the suspension. For the case of encapsulated gas bubbles, (4) where the factor 8.686 is necessary to convert from neperios to dB. Equations (3) and (4) can be combined to produce the following relationship for the return power: (5) (W / cm where x is now the distance between the transducer and the volume of the sample, and the G factor represents the additional geometric factors that include the transducer aperture, the distance between the transducer and the volume of the sample, and the angle of the solid intercepted by the spherical wave dispersed from each bubble in the receiver transducer. To make use of these results, it is necessary to provide an encapsulated gas bubble size distribution and estimate the values for the physical parameters used in the model. Two cases are considered. The first is for microparticles produced from polyesters and the second is for microparticles produced from albumin. The size distribution for the synthetic particles used in this case is the one that was measured for the PLGA-PEG microparticles produced by spray drying, as described in US Serial No. 08 / 681,710 filed on July 29, 1996, teachings of which are incorporated into the present. The values of the population parameters that characterize this distribution, as determined by the Coulter Multisizer® analysis, are: total concentration: 2.4 x 10 particles / ml, numerical average diameter: 2.2 μ, average volumetric diameter: 4.6 μ and 6.5% of the volume fraction of gas. The calculations provided in the following were produced assuming a dilution of 1/1620. The corresponding concentration was 4.4 x 10 particles / ml while the volume fraction of the gas was approximately 0.01%. the values for the parameters used in the model are: internal gas: adequate values for perfluoropropane, external liquid: values suitable for water, cover density: 1.5 g / cm, cover viscosity: 30 poise, cover stiffness: 10 MPa and cover thicknesses: 22, 55, 110, 165, 220, 330, 440, 660, 880 and 1100 nm. The results of the calculations for cross-sectioning of the total dispersion at the driving frequency are shown in Figure 1 for the range of PEG-PLGA tire thicknesses employed. At the lower frequencies, the cross-sections are increased approximately as the fourth power of the frequency, as expected for small dispersers, ie, Rayleigh. At higher biomedical frequencies, the total dispersion increases only as the frequency at the 1.5 power. At even higher frequencies, the intensity of the dispersion is continuous and then declines. The effect of increasing the thickness of the cover is to decrease the cross section of the total dispersion by an amount approximately equal to or somewhat greater than the proportional change in thickness. Thus, the cross section of the total dispersion having a suspension of encapsulated gas bubbles can be controlled by varying the thickness of the cover. The results of the calculations for the attenuation coefficient as a function of the driving frequency at different tire thicknesses are shown in Figure 2.

The effect of increasing the thickness of the roof is to decrease the attenuation coefficient by an amount approximately equal to or somewhat less than the proportional change in thickness. Thus, the attenuation coefficient can be controlled by varying the thickness of the cover. The fact that both the cross section of the total dispersion and the attenuation coefficient are increased approximately in proportion to the decrease in the thickness of the roof may seem to indicate that the variation in the thickness of the roof has no effect on the power expected total that will be backscattered to a transducer that emits an acoustic wave in a suspension of encapsulated gas bubbles. With another consideration of Equation 5, however, it is evident that the suspension of encapsulated gas bubbles having thicker covers will present greater total return power. This is shown in Figure 3. The reason for this is that while the total backscattered power is directly proportional to the total dispersion cross section, is also proportional to the exponential of the attenuation coefficient. Therefore, if the thickness of the roof decreases by a factor of 2, the effect of the increase in the cross section of the total dispersion will be to increase the total power by approximately 2 while the effect of the attenuation will be to "increase" the total power by a factor d approximately exp (-2) = 1 / 7.4, for a net decrease of approximately 73%. The total return power increases as the thickness of the cover increases. Similar results are predicted for microparticles produced from albumin as shown in Figures 4-6. The parameters used for albumin are as described in C. Church, J. Acousticai Soc. Amer., 97 (3) 1510-1521 (1995). The total return power for the synthetic polymeric microparticles (Figure 3) and for the microparticles of albumin (Figure 6) for a microparticle dilution factor of 1/1620. The optimum thickness of the roof (defined as the thickness that provides a maximum in the total return power at a depth of 2 cm in a suspension of encapsulated gas bubbles) will depend on the dilution of the encapsulated gas bubbles (ie, the concentration of the encapsulated gas). This is shown in Figure 7 for albumin microparticles with a dilution of 1/5400. As the suspension is diluted it is possible to use microparticles with thinner covers. This occurs because although the thinner covers cause greater attenuation and greater intensity of "bubble-to-bubble" dispersion, it is sufficiently displaced in a number of microparticles to produce maximum total return power.

The optimum thicknesses of the cover for three dilutions are summarized in the following table for albumin and PEG-PLGA microparticles.

For bubbles whose size distribution is relatively stable in vivo, the choice of optimum cover thickness will be based on the expected concentration of the particles in the target organ of interest. To illustrate how the thickness of the cover can be selected, the above-described synthetic microparticles are considered. If the microparticles are dosed at approximately 0.25 ml / kg and the blood volume is assumed to be 50 ml / kg, the microparticles would be diluted 1/200 after intravenous injection. In the myocardium, the blood constitutes 10% of the total volume of the compartment and the microparticles are further diluted in the compartment by a factor of 10. Thus, the final dilution would be approximately 1/2000. At this dilution, the optimum thickness of the cover can be extrapolated from the data in the table and is 200 nm. Thus, the optimum thickness for use as a perfusion agent in the myocardium for these types of microparticles is approximately 200 nm. Based on this information, thicker covers should be used to optimize the design of a specific gas encapsulating icroparticle, minimize attenuation and maximize backscattered return power, allowing the use of high doses of contrast agents of ultrasound with minimum attenuation. Methods for producing the microparticles with suitable wall thicknesses are described below.

II. Processes and reagents for making microparticles with different cover thicknesses As used herein, the term "microparticle" includes microspheres and microcapsules, as well as microparticles, unless otherwise specified. The microparticles may or may not be spherical in shape. The microcapsules are defined as microparticles that have an outer polymeric shell surrounding a core of another material, in this case, a gas. The microspheres are microparticles having a honeycomb structure formed by pores through the polymer or combinations of honeycomb or microcapsule structures that are filled with a gas for imaging purposes, as described hereinafter. The term "wall thickness" or "polymer thickness" refers to the diameter of the polymer from the interior of the microparticle to the outside. In the case of a microcapsule with a hollow core, the wall thickness will be equal to the thickness of the polymer. In the case of porous microparticles having channels or pores in a polymeric sphere, the wall thickness can be equal to one half of the diameter of the microparticle.

Polymers It is possible to use non-biodegradable and biodegradable matrices for the microencapsulation of gases, although biodegradable matrices are preferred, particularly for intravenous injection. Non-erodible polymers can be used for enterally administered ultrasound applications. The synthetic or natural polymers can be used to manufacture the microcapsules. Synthetic polymers are preferred because the synthesis is more reproducible and the degradation controlled. The polymer is selected based on the time necessary for stability in vivo, in other words, the time necessary for distribution to the site where the image is to be formed, and the time necessary for the formation of the image.

Representative synthetic polymers are: poly (hydroxy acids) such as poly (lactic acid) poly (glycolic acid) and poly (lactic acid-co-glycolic acid) polyglycoires, polylactides, polylactide-co-ylcholide copolymers and mixtures, polyanhydrides, polyorthoesters, polyamides, polycarbonates, polyalkylenes, such as polyethylene and polypropylene, polyalkylene glycols such as poly (ethylene glycol), polyalkylene oxides such as poly (ethylene oxide), polyalkylene terephthalates such as poly (ethylene terephthalate), polyvinyl alcohols, polyvinyl ethers, polyvinyl esters, polyvinyl halides such as polyvinyl chloride, polyvinylpyrrolidone, polysiloxanes, polyvinyl alcohols, polyvinyl acetate, polystyrene, polyurethanes and copolymers thereof, celluloses derived as alkylcellulose, hydroxyalkylcellose, cellulose ethers , cellulose esters, nitro celluloses, methyl cellulose, ethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, hydroxybutyl methyl cellulose, cellulose acetate, cellulose propionate, cellulose butylate acetate, cellulose acetate phthalate, carboxyethyl cellulose, cellulose triacetate, and the sodium salt of cellulose sulfate (mentioned collectively herein as "synthetic celluloses", polymers of acrylic acid, methacrylic acid or copolymers or derivatives thereof including esters, poly (methyl methacrylate), poly (ethyl methacrylate), poly (butyl methacrylate), poly (isobutyi methacrylate) , poly (hexyium methacrylate), poly (isodecyl methacrylate), poly (lauryl methacrylate), poly (phenyl methacrylate), poly (methyl acrylate), poly (isopropyl acrylate), poly (isobutyl acrylate), and poly (octadecyl acrylate) (together referred to herein as "polyacrylic acids"), poly (butyric acid), poly (valeric acid), and poly (lactide-co-caprolactone), copolymers and mixtures thereof same . As used herein, the "derivatives" include polymers having substitutions or additions of chemical groups, for example, alkyl, alkylene, hydroxylations, oxidations, and other modifications made routinely by those skilled in the art. Examples of the non-biodegradable polymers include ethylene vinyl acetate, poly (meth) acrylic acid, polyamides, copolymers and mixtures thereof. Examples of the preferred biodegradable polymers include polymers of hydroxy acids such as lactic acid and glycolic acid, polylactide, polyglycolide, polylactide-co-glycolide and copolymers with PEG, polyanhydrides, poly (ortho) esters, polyurethanes, poly (butyric acid), poly (valeric acid), poly (lactide-co-caprolactone), mixtures and copolymers thereof. Examples of the preferred natural polymers include proteins such as albumin, hemoglobin, fibrinogen, polyamino acids, gelatin, lactoglobulin, and prolamines, for example, zein, and polysaccharides such as alginate, cellulose, and polyhydroxyaicates, for example, polyhydroxybutyrate. The proteins can be stabilized by crosslinking with an agent such as glutaraldehyde or thermal denaturation. Bioadhesive polymers of particular interest for use in imaging mucosal surfaces, such as - in the gastrointestinal tract, include polyanhydrides, polyacrylic acid, poly (methyl methacrylates), poly (ethyl methacrylate), poly (methacrylate) butyl), poly (isobutyl methacrylate), poly (hexyl methacrylate), poly (isodecyl methacrylate), poly (lauryl methacrylate), poly (phenyl methacrylate), poly (methyl acrylate), poly (acrylate), isopropyl), poly (isobutyl acrylate), and poly (octadecyl acrylate).

Solvents As defined herein, the polymer solvent is a solvent that is volatile or has a relatively low boiling point or can be removed in a vacuum and that is acceptable for administration to humans in trace amounts, such as methylene chloride. , water, ethyl acetate, ethanol, methanol, dimethylformamide (DMS), acetone, acetonitrile, tetrahydrofuran (THF), acetic acid and dimethyl suiphoxide (DMSO), or combinations thereof. In general, the polymer is dissolved in the solvent to form a polymer solution with a concentration of between 0.1 and 60% by weight to volume (w / v), more preferably between 0.2-5 and 30%.

Gases Any biocompatible or pharmacologically acceptable gas can be incorporated into microparticles. The term "gas" refers to any compound that is a gas or is capable of forming a gas at the temperature at which the formation of the image is taking place. The gas may be composed of a single compound such as oxygen, nitrogen, zenon, argon, nitrogen, fi uorinated gases, or a mixture of compounds such as air. Fluorinated gases are preferred. Examples of the fluorinated gases include CF, C¿F, C3F8, C4F8, SF6, C2F / and C3F6. Perfluoro propane is particularly preferred because it is pharmacologically acceptable. Commonly, hollow, air-filled microparticles are produced by the methods described and the air within the particles can be exchanged with any of the biocompatible gases described. The gas is usually exchanged by a vacuum in the microparticles to remove the air and then apply an atmosphere of the biocompatible gas at a specific temperature and pressure. The temperature and pressure of the gas to be exchanged will depend on the properties of the microparticles.

Pore Forming Agents Pore forming agents can be microencapsulated to introduce internal voids. The pore-forming agent can be a liquid or a volatile or sublimable sai that can be removed during microencapsulation or can be removed after the microparticles are formed using vacuum drying or lyophilization. After the elimination of the pore-forming agent the created internal voids can be filled with the gas of interest. It is possible to use more than a pore-forming agent. The pore-forming agent or agents may be included in the polymer solution in an amount of between 0.01% and 90% by weight by volume, to increase pore formation. For example, in spray drying, the evaporation of solvents, a pore-forming agent, such as a volatile salt, for example, ammonium bicarbonate, ammonium acetate, ammonium chloride or ammonium benzoate or other freeze-dried sai can be Encapsulated as solid particles or as a solution. If the pore-forming agent is encapsulated with a solution, the solution containing the pore-forming agent is ulsified with the polymer solution to create droplets of the pore-forming agent in the polymer. The polymer solution containing the particles of the pore-forming agent or the emulsion of the solution of the pore-forming agent in the polymer is then spray-dried or taken through a solvent evaporation / extraction process. After the polymer has been precipitated, the hardened microparticles can be frozen and lyophilized to remove the residual pore-forming agent or the hardened microparticles can be dried in vacuo to remove the pore-forming agent.

Additives for stabilizing the encapsulated gas Lipids In general, the incorporation of the compounds during the production of the microparticles that are hydrophobic and, in an effective amount, limits the penetration and / or uptake of water by the microparticles by this means, is effective in the stabilization of the echogenicity of polymeric microparticles that have gas encapsulated in them, especially fluorinated gases such as perfluoro carbons.

Lipids that can be used to stabilize gas within the polymeric microparticles include, but are not limited to the following lipid classes: fatty acids and derivatives, mono-, di and triglycerides, phospholipids, sphingolipids, cholesterol and spheroidal derivatives, terpenes and vitamins The fatty acids and derivatives thereof may include, but are not limited to, saturated to unsaturated fatty acids, even and non-existent fatty acids, cis and trans isomers, and fatty acid derivatives including alcohols, esters, anhydrides, hydroxy fatty acids and prostaglandins. Saturated and unsaturated fatty acids that can be used include, but are not limited to, molecules having between 12 carbon atoms and 22 carbon atoms in any linear or branched form. Examples of the saturated fatty acids that may be used include, but are not limited to, lauric, myristic, palmitic and stearic acids. Examples of the unsaturated fatty acids that may be used include, but are not limited to, lauric acid, fisetérics [sic], myristoleic acid, palmitoleic acid, petrocelinic acid, and oleic acid. Examples of branched fatty acids that can be used, include, but are not limited to, isoluric, isyrimic, isopalmitic, and isostearic acids and the isprenoids. Fatty acid derivatives include 12 - (((7'-diethylaminocoumarin-3-yl) carbonyl) methylamino) -octadecanoic acid; N- [12- (({ '-dietilaminocumarin-3-yl) carbonyl..!) Ethylamino) -octadecanoil] -2-aminopalmítico, N-succinyl-dioleiifosfatidiietanolamina and palmitoyl-omocisteina; and / or combinations thereof. Mono, di and triglycerides or derivatives thereof that may be used include, but are not limited to molecules that have fatty acids or mixtures of fatty acids between 6 and 24 carbon atoms, digalactosyldiglyceride, 1,2-dioleoyl-sn -gliceroi; 1,2-dipalmitoii-sn-3 succinylglycerol; and 1,3-dipalmitoyl-2-succinylglycerol. Phospholipids which may be used include, but are not limited to phosphatidic acids, phosphatidyl cholines with saturated and unsaturated lipids, phosphatidyl ethanolamines, phosphatidylglycerols, phosphatidylserines, phosphatidylinositols, lysophosphatidyl derivatives, cardiolipin and beta-acyl-alquii and phospholipids. Examples of phospholipids include, but are not limited to, phosphatidylcholines such as dioleoylphosphatidylcholine, diiuiristoilfosfatidilcolina, dipentadecanoylphosphatidylcholine, dilauroylphosphatidylcholine, diptoilfosfatidilcolina (DPPC), distearoylphosphatidylcholine (DSPC), diarachidoylphosphatidylcholine (DAPC), dibehenoilfosfatidilcolina (DBPC), ditricosanoiolfosfatidilcolina (DTPC), dilignocerodilfosfatidilcolina ( DLPC); and phosphatidyethanolamines such as dioiefilfosf tidiiethanolamine or l-hexadecyl-2-palmitoyl glycerophosphoethanolamine. Synthetic phospholipids with asymmetric acyl chains (for example, with a 6-carbon acyl chain and another 12-carbon acyl chain) can also be used.

Sphingolipids that can be used include ceramides, sphingomyelins, cerebrosides, gangliosides, suifatides and lysulfosulfatides. Examples of sphingoiipids include, but are not limited to, gangliosides GM1 and GM2. Spheroids that can be used include, but are not limited to, cholesterol, cholesterol sulfate, cholesterol emisuccinate, 6- (5-cholesterol-3-yloxy) hexyl-6-amino-6-deoxy-1-thio-aD- galactopyranoside, 6- (5-cholesten-3-teloxy) exyl-6-amino-6-deoxy-l-thio- -D-mannopyranoside and cholesteryl) 4'-trimethyl-35-ammonium) butanoate [sic]. Additional lipid compounds that can be used include tocopherol and derivatives, and derived oils and oils such as stearylamine. It is possible to use a variety of cationic lipids such as DOTMA, N- [1- (2, 3-dioleoyloxy (propyl-N, N, N-trimethylammonium, DOTAP, 1,2-dioleoyloxy-3- (trimethylammonium) propane; and DOTB, 1,2-dioleoyl-3- ('-rimetii-ammonium) butanoyl-sn-glyceryl.The most preferred lipids are phospholipids, preferably DPPC, DAPC, DSPC, DTPC, DBPC, DLPC and most preferably DPPC , DAPC and DBPC.The lipid content is in the range from 0.01-30 (p of lipid / p of polymer), more preferably between 0.1-12 (p lipid / p polymer) .The lipids can be added to the solution polymer before the formation of microparticles.

Other hydrophobic compounds Other preferred hydrophobic compounds include amino acids such as tryptophan, tyrosine, isoleucine, leucine and valine, aromatic compounds such as alkyl paraben, for example methyl paraben and benzoic acid.

Microparticles and methods for making them In the most preferred embodiment, the microparticles are produced by spray drying. The polymer and the pore-forming agent are atomized through a nozzle and the solvent of the polymer is evaporated by a heated drying gas. It is possible to use other techniques, such as solvent extraction, encapsulation of the hot melt and evaporation with solvents, to produce microparticles having a wall thickness of adequate diameter to optimize echogenicity. Pore forming agents are commonly used to create internal voids. The pore-forming agents are microencapsulated and eliminated after the formation of the microparticle by lyophilization or vacuum drying. The evaporation of the solvents is described in E. Mathiowitz, et al., J. Scanning Microscopy, 4, 329 (1990); L. R. Beck, ec al., Fertile. Steril., 31, 545 (1979); and S Benita, et al., J. Pharm. Sci., 73, 1721 (1984). Hot melt microencapsulation is described by? Mathiowitz, et al., Reactive Polymers, 6, 275 (1987). During the synthesis of the microparticles a variety of surfactants can be added. Exemplary emulsifiers or surfactants that can be used (0.1-5% in pe'so) include the most physiologically acceptable emuisifiers. Examples include natural and synthetic foams of bile salts or bile acids, conjugated with amino acids and unconjugated such as taurodeoxycholate, and cholic acid.

Size of the microparticle In a preferred embodiment for the preparation of injectable microparticles capable of passing through the pulmonary capillary bed, the microparticles should have a diameter between about 1 and 10 microns. Larger particles can clog the pulmonary bed, and smaller particles may not provide enough echogenicity. The larger microparticles are useful for administration by routes other than injection, for example the oral route (for evaluation of the gastrointestinal tract), application to other surfaces of the mucosa (rectal, vaginal, oral, nasal), or by inhalation. The preferred particle size for oral administration is between about 0.5 microns and 5 mm. The analysis of the particle size can be done in a Coulter counter, by light microscopy, electron scanning microscopy or electron transmittance microscopy.

Wall thickness control The preferred wall thickness is greater than 20 nm, more preferably in the range of 160 to 220 n to approximately 700 nm, at which point the advantage obtained by increasing the wall thickness begins to fall. For each of the microencapsulation techniques described above there are several ways in which the final thickness of the cover of the polymeric microparticles can be controlled.

Polymer concentration The final thickness of the polymeric coating can be increased by increasing the concentration of the polymer phase during the encapsulation process. This is applicable to synthetic polymers or natural polymers such as proteins or polysaccharides. For a given polymer droplet size, the use of a more concentrated polymer solution will give rise to more polymer per unit volume of droplets and thus a thicker cover.

The concentration of the polymer to obtain a certain cover thickness will depend mainly on the type of polymer, the solvent of the polymer, the solubility of the polymer and the solvent system and, the temperature at which the encapsulation is carried out. Polymer concentrations in the range from 0.1 to 60% can be used. The preferred polymer concentrations are in the range of between 0.5 and 30%. As already described, pore-forming agents, such as volatile or sublimable salts, can be used to produce microparticles with internal voids. The pore-forming agent can be microencapsulated as solids or as an aqueous solution or can be co-dissolved in the polymer solution. In the case of solid pore forming agents, the size of the solid particles and the amount of the encapsulated solid agent will control the final thickness of the polymeric shell. Thinner coatings of the microparticles will conform to the diameter of the solid pore forming particles increasing with respect to the phase of the polymeric droplets or as the weight of the solid pore forming agent increases with respect to the phase of the polymeric droplets . The diameter of the solid pore-forming microparticles [sic] is between 1 and 95% of the diameter of the phase of the polymeric droplets. The diameter of the solid pore forming agent can be adjusted to the appropriate diameter using standard techniques such as jet grinding. The weight of the solid pore forming agent to be encapsulated is between 1 and 50% (w / w of the polymer). In the case of a pore-forming agent that is dissolved in the polymer solvent, the concentration of the shell will be governed by the amount of the encapsulated pore-forming agent. As the total amount of the pore-forming agent increases, the final thickness of the cover will decrease. For a pore-forming agent microencapsulated as an aqueous solution, the final thickness of the polymeric sheath will be governed by the volume of the encapsulated pore-forming solution relative to the polymer phase, the weight of the microencapsulated pore-forming agent and the size of the pore-forming agent. the droplets of the solution of the pore-forming agent with respect to the size of the polymeric droplet. The final thickness of the polymeric coating will decrease as the volume ratio of the pore-forming solution increases with respect to the polymer phase. The volume ratio of the pore-forming solution relative to the polymer phase is between 0.002 and 0.5 with preferred ratios in the range from 0.01 to 0.1. For a volume ratio of the pore-forming agent, the thickness of the polymeric coating will decrease as the concentration of the pore-forming agent increases in the pore-forming solution to be encapsulated. The weight of the pore-forming agent to be encapsulated is between 1 and 50% (w / w of the polymer). As the droplet size of the pore-forming solution that is going to be encapsulated relative to the polymer solution decreases, the thickness of the cover of the final microparticle will be increased. The droplet size of the pore-forming solution can be controlled by the process used to create the droplets. The diameter of the droplets of the pore-forming solution is in the range of between 1 and 95% of the diameter of the phase of the polymeric droplets. If homogenization is used to create the pore-forming droplets, homogenization speed (500-20,000 rpm), homogenization time (0.1-10 minutes), homogenization temperature (4-50 ° C) and ei Type of blades (ie, slotted head, square head, circular head) used will regulate the final size of the droplets of the pore-forming solution. The homogenization conditions are adjusted to create the size of droplets of interest. If voicing is used to create the droplets of the liquid pore-forming solution in the polymer droplets, the type of voicing probe, the voicing time (0.1-10 minutes).

The sounding temperature (4-40 ° C), the frequency of the probe and the power of the voicing can all be used to modify the size of the droplets.

III. Diagnostic Applications Usually, the microparticles are combined with a pharmaceutically acceptable carrier such as saline buffered with phosphate or saline or mannitol, then an effective amount for detection administered to a patient using a suitable route, usually by injection into a blood vessel (iv) or orally. Microparticles containing an encapsulated imaging agent can be used in vascular imaging, as well as in applications to detect liver and kidney diseases, in applications in cardiology, in the detection and characterization of tumor masses and tissues and in the measurement of the peripheral blood velocity. The microparticles can also be linked with ligands that minimize adhesion to tissues or direct the microparticles to specific regions of the body as previously described. The methods and compositions described above will also be understood with reference to the following non-limiting examples.

Example 1: Production of polymeric misroparticles having improved echogenicity. 3.2 grams of PEG-PLGA ^ 75: 25) (IV = 0.75 dL / g), 6.4 g of PLGA (50:50) (IV = 0.4 dL / g;, and 384 mg of diarachyidoylphosphatidylcholine were dissolved in 480 ml of chloride of methylene, 20 ml of 0.18 g / ml ammonium bicarbonate solution were added to the polymer solution and the polymer / salt mixture was homogenized at 10,000 RPM for 2 minutes using a Vircís homogenizer.The solution was pumped at a flow rate 20 ml / min and spray-dried using the Buchi Lab spray dryer. The intake temperature was 40 ° C and the discharge temperature was 20-22 ° C. The diameters of the particles were in the range from 1 -10 microns when the size was determined in a coulter counter with a numerical average [sic] from 2.0 microns.The electronic scanning microscopy showed that the particles were generally spherical with smooth surfaces and occasional surface crenulations. to electron transmission microscopy by incrusting them in white LR resin followed by polymerization under UV light. The thin sections were cut in an ultramicrotome LKB using a glass knife and observed in a Zeiss EM-10 TEM at 60 kv. The thickness of the cover of the microparticles is in the range of between 200 and 240 nm.

Claims (15)

1. A method for increasing the echogenicity of microparticles by encapsulating a gas for use in ultrasound imaging is to: determine the range of microparticle wall thicknesses that give rise to higher amounts of total backscatter power as a function of the material which forms the microparticles and the encapsulated gas, and produces the microparticles with a wall thickness in the range that originates the highest levels of the total backscattering power.

2. The method of claim 1, wherein the microparticles are formed of a synthetic polymer.

3. The method of claim 2, wherein the thickness of the microparticles is between 50 and 660 nm. The method "of claim 1, wherein the microparticles are formed of a natural polymer 5. The method of claim 4, wherein the natural polymer is a protein and the thickness of the microparticles is between 20 and 600 nm 6. An ultrasound imaging method consists of administering to an individual from whom the image is to be formed, polymeric microparticles encapsulating a gas, wherein the microparticles have a polymer wall thickness in the range that they originate from. higher levels of total backscattered power for microparticles of the same composition and size 7. The method of claim 6, wherein the microparticles are formed of a synthetic polymer 8. The method of claim 7, wherein the The thickness of the microparticles is between 50 and 660 nm and the microparticles are formed of a synthetic polymer 9. The method of claim 6, wherein the microparticles they are formed of a natural polymer 10. The method of claim 9, wherein the natural polymer is a protein and the thickness of the microparticles is between [sic] 11. The method of claim 6, wherein the gas is a fluorinated gas. 12. An ultrasound composition containing polymeric microparticles encapsulating an echogenic amount of a biocompatible gas, wherein the microparticles are selected from a population of microparticles based on polymeric wall thicknesses of the microparticle with maximum echogenicity and minimal attenuation. The composition of claim 12, wherein the polymer is a synthetic polymer with the exception of a block copolymer of polyethylene glycol and poly (iacturo-co-glycolide). 1

4. An ultrasound composition containing microparticles formed of a natural polymer, wherein the wall thickness of the polymer is greater than 20 nm. 1

5. The composition of claim 14, wherein the polymer is albumin. SUMMARY OF THE INVENTION It has been found that microparticles formed of natural or synthetic polymer with thicker walls have significantly improved echogenicity compared to microparticles with thinner walls. The effect of the thickness of the walls has been determined experimentally as well as inserted into a formula for use in the prediction of optimum conditions. In the preferred embodiment, the polymers are synthetic, biodegradable polymers and the wall thickness is between 100 and 660 nm, although a wall thickness of from about 20 nm to an excess of 500 nm can be used. The microparticles are manufactured with a diameter suitable for the particular tissue to which the image is to be formed, for example, with a diameter between 0.5 and 8 microns for intravascular administration, and a diameter between 0.5 and 5 mm for oral administration to imagine the gastrointestinal tract or other lamina. Preferred polymers are polyhydroxy acids co or polylactic acid-co-glycolic acid, polylactide or polylglycolide, more preferably conjugated with a polyethylene glycol or other materials that inhibit uptake by ei. reticuiocndothelial system (SER). The microspheres can be used in a variety of applications in ultrasound imaging, including applications in cardiology, applications in blood perfusion, as well as for peripheral and organ imaging.