An edible, hardenable coating composition containing microcrystalline cellulose and carrageenan and at least one of a strengthening polymer, a plasticizer, a surface active agent or a combination thereof. The coating composition of the present invention may be applied to pharmaceutical and veterinary solid dosage forms, confectionery, seeds, animal feed, fertilizers, pesticide tablets, and foods and provides an elegant prompt release coating which does not retard the release of active ingredients from the coated substrate.

This invention relates to edible, hardenable, prompt release coating compositions comprising microcrystalline cellulose (MCC), carrageenan (CGN) and at least one of a strengthening polymer or a plasticizer. The coatings of the present invention can be applied to pharmaceutical, including nutriceutical, and veterinary solid dosage forms, confectionery, seeds, animal feed, fertilizers, pesticide tablets and granules, and foods, are readily dispersed in aqueous media, and, when applied as a coating and ingested by, for example, a human, do not significantly retard or extend release of active ingredient(s) from a substrate coated therewith.
It is a common practice to coat pharmaceutical and veterinary tablets to obtain several advantages. Among these are to mask unpleasant tasting active ingredients with a barrier coat, to improve the surface characteristics of tablets to make them easier to swallow, to reduce the absorption of water or moisture which can potentially degrade the active ingredient or promote some other undesirable change in the tablet structure, and simply to make a more elegant appearing tablet.
Another very important function of a pharmaceutical or veterinary tablet coating is to improve the integrity of the tablet itself. Uncoated tablets are often subject to being abraded or chipped, causing a loss of active ingredient in the process. More dramatically, they may break into two or more pieces. One measure of a useful coating is its ability to prevent any of these physical degradations of tablet structure. The effectiveness of a coating material to prevent abrading, chipping, or breakage of the tablet is determined by friability testing.
Confectionery and foods may be coated with a formulation to preserve the confection or food from deteriorating by contact with the oxygen and the moisture in the atmosphere. Coatings also can provide improved appearance and desirable organoleptic properties to the food as well as preventing loss of flavor.
Seeds may be coated to preserve the viability of the seeds by protecting against moisture. They may also be coated as a means for increasing particle size to facilitate mechanical planting. A dye can be included in the coating formulation to identify the seeds as to quality, type, or some other designation. Frequently, a pesticide, e.g., a fungicide, is incorporated into the coating formulation to protect both the seed itself and the seedling that results from germination of the seed. In all cases, this coating must not decrease the viability of the seeds or interfere with germination when the seeds are planted in the soil.
Animal feed may be coated to improve its flowability, appearance and its resistance to powdering or dusting. In such applications, the coating may be formulated to include vitamins, hormones, antibiotics, or the like, to benefit the livestock which will consume the feed.
Fertilizers, in either granular or tableted forms, may be coated to retain the integrity of the form and, especially, to protect the fertilizer from moisture which can cause agglomeration during storage, which could make rapid, even application to the soil difficult or inconvenient.
Coating of tableted pesticide formulations serves to maintain the integrity of the tablets or granules until they are placed in water where they rapidly disintegrate, forming a solution or slurry to be applied to the soil or plants. A second, and equally important, function of the coatings on tablets containing pesticides is to prevent human contact with the pesticide, thereby increasing safety for those handling and applying the pesticide.
Currently, most commercially available edible coatings utilize a synthetic cellulosic polymer such as hydroxypropylmethylcellulose (HPMC). Other synthetic film-formers which are commonly used include ethylcellulose, methylcellulose, polyvinylpyrrolidone, and polydextrose. These coating materials may be used alone or in combination with secondary film-formers such as sodium alginate or propylene glycol alginate. The foregoing are usually used in combination with other ingredients including fillers, for example, lactose or maltodextrin; plasticizers, such as polyethylene glycols, dibutyl sebacate, and triethyl citrate; surfactants; and often coloring materials such as a food dye or pigment, including opacifiers such as titanium dioxide and the like.
In the preparation of a coating formulation to be sprayed, the film former is usually dissolved or dispersed in a solvent, for example, water, along with the other ingredients of the formulation. In aqueous systems, since many polymers require significant time to become fully hydrated, the coating formulation must frequently be prepared in advance of the time it is to be applied to the tablets. A common procedure is to prepare these coating formulations the day preceding the coating operation in order to assure adequate hydration of the polymers used in them.
A particular disadvantage of coatings based primarily on HPMC is that the coating may harden over time and therefore increase tablet disintegration times. An increase in disintegration time delays the bioavailability of the active ingredient at least in proportion to the increase in disintegration time. Many other agents commonly used in coating compositions are also known to delay release of pharmaceutical agents, such as enteric coatings which use polymeric film forming materials which are insoluble in water, or gastric fluid, some of these being specifically selected to by-pass both the stomach and small intestine and provide colonic release.
The coatings of this invention meet U.S. Pharmacopoeia standards for rapid or immediate dissolution (U.S.P. monograph 23) of active ingredients from tablets or other solid dosage forms coated with them. They provide prompt release or dissolution consistent with the release rates which is normally obtained with the uncoated tablets or other substrates. Thus, they do not adversely impact or retard release of active ingredients from a substrate coated with them. Further, the coatings of this invention are readily dispersed and rapidly hydrated in aqueous media for application to a coating substrate, and provide elegant coatings which have all the benefits of coatings now in commercial use without the drawbacks that are common to them.
It has been found that these and other advantages may be achieved in accordance with the present invention by a coating composition which comprises a unique combination of materials specifically adapted for a prompt release when placed aqueous media or ingested, e.g., by a human. The coating composition of the present invention comprises microcrystalline cellulose, carrageenan, and at least one of a strengthening polymer and a plasticizer. More specifically, the present invention provides a prompt release, edible, hardenable coating composition comprising 3% to 40% microcrystalline cellulose, 9% to 25% iota carrageenan, and at least one of a strengthening polymer, a plasticizer, or a surface active agent, as well as to dry coatings and aqueous dispersions thereof.
The present invention also provides pharmaceutical, including neutriceutical, and veterinary solid dosage forms, confectionery, seeds, animal feed, fertilizers, pesticide tablets and granules, and foods coated with the prompt release edible, hardenable composition of this invention.
For purposes of this application, the term xe2x80x9cediblexe2x80x9d is intended to mean food grade materials which are approved by regulatory authorities for use in pharmaceutical or food applications. The term xe2x80x9chardenablexe2x80x9d used to describe the coating compositions of this invention is intended to include only those coating compositions that are capable of being dried from an aqueous solution or dispersion thereof into a solid coating which resists abrasive forces, i.e. a hardened coating, as distinguished from those xe2x80x9cenrobingxe2x80x9d coatings on confections which set up into a soft coating that can be handled and packaged but which do not resist abrasive forces significantly. The terms xe2x80x9cimmediatexe2x80x9d, xe2x80x9crapidxe2x80x9d or xe2x80x9cpromptxe2x80x9d release as applied to dissolution rates or times for the coating compositions of this invention or tablets coated with the compositions of this invention means that the coatings of this invention meet U.S. Pharmacopoeia standards (U.S.P. monograph 23) for rapid or immediate dissolution of active ingredients from tablets or other solid dosage forms coated therewith. Thus, they provide prompt release or dissolution consistent with the release rates which is normally obtained with the uncoated tablets or other substrate. They do not, consistent with the pharmacopeia standards above, when placed in aqueous media or ingested by, e.g., a human, significantly impact or retard release or dissolution of tablets or other solid dosage forms coated therewith. For example, coatings made in accordance with the present invention are substantially or completely disintegrated and/or dissolved within less than 10 minutes after being ingested or placed in aqueous media. Thus, when a pharmaceutical solid dosage form is coated with the coating of this invention and ingested by a human or other animal, the coating of this invention is dissolved or disintegrated prior to leaving the stomach. In the specification, examples and claims which follow, all percentages are by dry weight of the composition. These definitions are intended to apply throughout this application unless a contrary meaning is clearly indicated.
The microcrystalline cellulose, either coprocessed with carrageenan or simply blended therewith, interacts with the carrageenan to provide important film-forming characteristics required to provide an elegant coating which is particularly useful in, for example, coating pharmaceutical and veterinary tablets, caplets, granules, and spheres which contain active ingredients which require release promptly after being placed in aqueous media or ingested.
Microcrystalline cellulose is a purified, partially depolymerized cellulose that is generally produced by treating a source of cellulose, preferably alpha cellulose in the form of a pulp from fibrous plants, with a mineral acid, preferably hydrochloric acid. The acid selectively attacks the less ordered regions of the cellulose polymer chain, thereby exposing and freeing the crystallite sites, forming the crystallite aggregates which constitute microcrystalline cellulose. These are then separated from the reaction mixture and washed to remove degraded by-products. The resulting wet mass, generally containing 40 to 60 percent moisture, is referred to in the art by several names, including hydrolyzed cellulose, microcrystalline cellulose, microcrystalline cellulose wetcake, or simply wetcake. This microcrystalline cellulose wetcake may be used as such or may be further modified, for example, by attrition and/or drying, and utilized in accordance with the present invention.
Microcrystalline cellulose may also be produced for use in the present invention using a steam explosion treatment. In this process, wood chips or other cellulosic materials are placed in a chamber into which super-heated steam is introduced. After being maintained for a period of about 1-5 minutes, the exit valve is opened rapidly, releasing the contents explosively and yielding microcrystalline cellulose. No additional acid need be introduced into the reaction mixture, since it is believed that the acidic materials in the wood chips and the elevated temperature and pressure hydrolyze the cellulose and degrade it. In addition to the specific forms of microcrystalline cellulose, the present invention also contemplates the use of other cellulose derivatives, including microreticulated cellulose, also known as microreticulated microcrystalline cellulose, and powdered cellulose such as a commercial material sold as xe2x80x9cSolka Floc(copyright).xe2x80x9dAs discussed in greater detail below, the microcrystalline cellulose preferred for use in the present invention is microcrystalline cellulose which has an average particle size below about 100 microns, preferably microcrystalline cellulose which been attrited or has an average particle size in the range of 1 to 50 microns, preferably 1 to 30 microns.
Carrageenan is used in combination with microcrystalline cellulose to form the elegant prompt release coatings of the present invention. Carrageenan for use in the present invention is a naturally derived carrageenan, including the grades further defined below as iota, kappa, and lambda carrageenan. The preferred type of carrageenan, a polysaccharide which is comprised of repeating galactose units and 3,6-anhydrogalactose units, that is suitable for the compositions of this invention is referred to as iota carrageenan. A rich source of iota carrageenan is the seaweed Eucheuma spinosum. The approximate content of anhydrogalactose units in iota carrageenan is 30% whereas kappa carrageenan has 34% anhydrogalactose units and lambda carrageenan is essentially devoid of these units. Carrageenans are also characterized by the amount of ester sulfate groups that are present on both the galactose and anhydrogalactose units. The ester sulfate content of iota carrageenan may range from about 25% to 34%, preferably about 32%. This is intermediate between kappa carrageenan which has a 25% ester sulfate content and lambda carrageenan which has a 35% ester sulfate content. The sodium salt of iota carrageenan is soluble in cold water, but different grades of iota carrageenan require heating water to different temperatures to dissolve them. The iota carrageenans which are suitable for the microcrystalline cellulose/iota carrageenan material of this invention are soluble in water heated up to 80xc2x0 C. (176xc2x0 F.). Preferred grades of iota carrageenan are soluble at lower temperatures, for example, at 50xc2x0 C. (122xc2x0 F.), including but not limited to sodium iota carrageenan.
In the coating compositions of this invention, a film forming amount of carrageenan must be employed. A suitable film forming amount of carrageenan is generally in the range of about 9% to about 25% by dry weight of the coating composition, advantageously in the range of about 10% to about 20% of the dry weight of the composition.
The microcrystalline cellulose and carrageenan may be coprocessed or may be blended in any suitable manner, such as dry blending.
Coprocessed microcrystalline cellulose/iota carrageenan is rapidly peptizable. Peptization means that the dry agent can readily be dispersed in water in a colloidal state. Peptization of a dry agent in aqueous media allows the functionality of the agent to be restored to a level near or preferably at the level observed before the agent was dried. Rapidly peptizable dry agents can be dispersed (peptized) in a colloidal state with minimal agitation. Thus, the novel coating formulations in which the coprocessed microcrystalline cellulose/iota carrageenan is incorporated can be hydrated in as little as 0.5 hour, but more preferably require 1 to 3 hours. The common practice of preparing the coating formulation on the preceding day may be avoided, although it may also be continued, if this is preferable, without harming the formulation. If the formulation is prepared on one day and then used the next day, prior to being used, the formulation must be stirred for a short while to restore it to its flowable state.
The coprocessed microcrystalline/iota carrageenan compositions useful in this invention may be prepared by first attriting hydrolyzed cellulose wetcake, such that the average particle size of the wetcake particles is generally not more than about 20 microns, preferably less than about 10 microns, dispersing the attrited wetcake in water heated to a temperature above the temperature at which the particular grade of iota carrageenan being used dissolves, adding the dry carrageenan to the dispersion of microcrystalline cellulose, mixing the components, preferably homogenizing the mixture to assure intimate mixing, and drying the dispersion. Spray-drying is normally used to prepare the dried materials useful in this invention, but other methods of drying the dispersion may equally be acceptable.
It is possible to prepare the coatings directly, that is, before the drying of the wetcake, from a dispersion of microcrystalline cellulose wetcake and the carrageenan by accounting for the water present in the wetcake and adding the other ingredients in the formulation to this dispersion. Although this method of operation may be preferred for some coating operations, it is usually preferable to use the spray-dried, or otherwise dried, material because transportation costs for a dispersion would be less economical. Furthermore, drying by any method may enhance the association of the microcrystalline cellulose with the carrageenan, which may result in a more satisfactory prompt release coating.
Dry blended microcrystalline cellulose (e.g., Avicel(copyright) PH-105, average particle size 20 microns) and iota carrageenan, has been found to provide coating compositions that are at least equal to, and in some cases, superior to, coating compositions prepared from coprocessed microcrystalline cellulose/carrageenan.
Carrageenan by itself is known to be a natural film forming hydrocolloid when an aqueous dispersion thereof is spread on a surface and allowed to dry. However, the film is considered to be too weak for pharmaceutical tablets as shown by the results in Comparative Example A and therefore requires the presence of microcrystalline cellulose for satisfactory results.
A dry, physical blend of iota carrageenan and microcrystalline cellulose (Avicel(copyright) PH-102, average particle size 100 microns) also yielded what appear to be commercially unsatisfactory results in Comparative Example B. Thus, for commercial purposes, it is believed that the average particle size of the microcrystalline cellulose used in a dry blend with the natural, film forming hydrocolloid should be below 100 microns, advantageously below about 50 microns, preferably in the range of about 1-50 microns, more preferably, about 1-30 microns. Elegant, high performance coating formulations within the scope of this invention may be prepared from such dry, physical blends of microcrystalline cellulose and carrageenan.
A wide range of weight ratios of microcrystalline cellulose to carrageenan of from 90:10 to about 15:85 may be employed in this invention. In those applications which require a higher ratio microcrystalline cellulose to carrageenan in the MCC:CGN weight ratio generally ranges from about 90:10 to about 60:40, particularly from about 85:15 to about 65:35, more particularly, approximately 70:30. In certain embodiments of this invention it has been found that relatively lesser amounts of microcrystalline cellulose can be employed, such that the ratio of MCC:CGN can be in the range of from about 15:85 up to 60:40, preferably up to 55:45, and most preferably up to 50:50. A particular advantage for the dry, physical blends is that the ratio can be easily changed by simple blending techniques rather than manufacturing different ratios of coprocessed material. Thus, the dry, physical blends provide significantly greater flexibility for specific applications having different requirements. Pharmaceutical and veterinary solid dosage forms containing certain active ingredients may require increased carrageenan content in the composition to ideally coat the tablets. For these pharmaceutical and veterinary applications, a preferred weight ratio of microcrystalline cellulose to carrageenan is in the range of about 75:25 to about 25:75.
Regardless of whether the composition is based on coprocessed microcrystalline cellulose/carrageenan or a dry, physical blend of microcrystalline cellulose and carrageenan, a strengthening polymer, preferably, hydroxyethylcellulose (HEC) or polyvinylpyrrolidone (PVP), a plasticizer or both a strengthening polymer and a plasticizer and/or a surface active agent may be present in the coating formulation of this invention.
Other strengthening polymers which can provide the same benefit and may be used instead of HEC include HPMC, hydroxypropylcellulose, ethylcellulose, and methylcellulose; however, care must be exercised in the use of such alternative materials to avoid significantly retarding release of active ingredients and/or bioavailability. The preferred amount of strengthening polymer is less than the total amount of microcrystalline cellulose and carrageenan present in the composition. Depending on the desired hardness of the coating, the strengthening polymer may be employed in the composition at a level of about 0.5% to about 30%, to provide strength and improved appearance to the coating. This strength can be demonstrated by casting films of coating formulations on a flat, non-adherent surface, cutting strips of uniform width from the casting, and subjecting the strips to tensile testing on, e.g., an Instron Tensile Tester. The results of these tests show a very significant increase in tensile strength and decreased brittleness of the film when HEC or another strengthening polymer is included in the formulation. Strengthening polymers suitable for use in this invention and which will not significantly retard release from tablets or other solid dosage forms, are those polymers having a viscosity equal to or less than 20 mPaxc2x7s in a 2% aqueous solution at 20xc2x0 C. When a strengthening polymer is employed in the formulation in absence of a plasticizer, it is generally employed at about 5% to about 30% by dry weight coating composition.
In a preferred composition a conventional plasticizer is also included in the coating composition. Suitable plasticizers include polyethylene glycol, advantageously a high molecular weight polyethylene glycol, triacetin, dibutyl sebacate, propylene glycol, sorbitol, glycerin, and triethyl citrate. Of these, polyethylene glycol is preferred. These plasticizers may be employed in the coating compositions of the invention at a level of 18% to about 36% by dry weight of the coating composition, most preferably at a level of 31% to 35% by dry weight of the coating composition.
The prompt release coating compositions of the invention may include at least one filler. Such fillers may include, for example, calcium carbonate, dicalcium phosphate and carbohydrates, such as starch, maltodextrin, lactose, mannitol and other sugars. Of these, maltodextrin, mannitol and lactose are preferred fillers.
The prompt release coating compositions of the invention may include at least one surfactant. Such surfactants include either anionic or nonionic surfactants. Useful surfactants may be, e.g., sodium lauryl sulfate, hydroxylated soy lecithin (lecithin), polysorbates, and block copolymers of propylene oxide and ethylene oxide. Lecithin has been found to be particularly desirable in certain coating compositions of this invention and is suitably utilized at a level of about 2% to about 10% by dry weight of the formulation.
Coloring agents and opacifiers may also be used in these coatings or added to a suspension thereof including aluminum lakes, insoluble pigments, water-soluble dyes, titanium dioxide, and talc. Stearic acid or a salt or ester thereof, may be included at a level of about 1% to about 5% by dry weight of the composition to increase gloss of the coating, particularly when a plasticizer is not employed in the composition. Likewise propylene glycol alginate, may be used in small quantities (about 5% to about 10% by dry weight of the composition) to increase the gloss of the coating, as shown in Example 31.
A coating formulation of this invention may be sold as a dry powder formulation or as a ready-to-use dispersion in water. For aqueous dispersions it is preferred that these be prepared under aseptic conditions. Heating the water to an elevated temperature, for example, 85xc2x0 C., prior to preparation of the dispersion has shown that bacteria, mold, and yeast growth are prevented for at least 48 hours on agar pour plates. Therefore, if the containers for the dispersion are properly sanitized and then kept closed after being filled until they are used, there is little likelihood of bacteria, mold, or yeast growing in the dispersion. Alternatively, if a formulation is to be sold as an aqueous dispersion to be stored for a period of time, a preservative may be added. A combination of methyl paraben and propyl paraben has been found to be useful in this regard.
On a dry weight percentage basis a preferred composition of this invention comprises at least about 43%, suitably about 45% to about 75% of microcrystalline cellulose and carrageenan powder combined, more preferably about 45% to about 60%; about 0.5% to about 30% of strengthening polymer, more preferably about 7% to about 22%; and about 25% to about 40% of plasticizer, more preferably about 31% to about 35%; and an inert filler at about 2% to about 28%. Optionally, about 1% to about 30% of the formulation may comprise edible coloring agents and opacificiers such as talc or titanium dioxide, including from 1% to about 8% of coloring component such as a food dye or pigment, preferably about 1% to about 3%. Other optional ingredients may include a surfactant at about 0.5% to about 10%, advantageously 0.5 to about 7%, preferably 1.25% to 3% when a filler such as maltodextrin or mannitol is present. When no filler is employed higher amounts of surfactants such as lecithin may be employed at a level of about 5% to about 20%. Preservatives, such as methyl paraben at 0.75% to 1.50% and/or propyl paraben at 0.075% to 0.15% may also be present in the formulation. When maltodextrin is the filler, it is generally employed at about 2% to about 7% by dry weight of the composition, whereas when mannitol is the filler it is generally employed at about 10% to about 25% by dry weight of the formulation. These fillers may be employed alone or in combination within the ranges specified above.
The low level of fillers present in these coating formulations, particularly when the opacifier is titanium dioxide, enables the formulator to utilize relatively small amounts of coloring agent. Since coloring agents are quite costly, this provides a significant cost reduction from those formulations requiring from 6% to about 16% to effectively color prior art coating formulations.
Several other embodiments of this invention may be specifically mentioned as being of special interest to this invention, in which there is provided an edible, hardenable, prompt release coating composition comprising about 3% to 40% microcrystalline cellulose in combination with about 9% to about 25% iota carrageenan and at least one of a strengthening polymer, a plasticizer, or a surface active agent, wherein said coating composition is rapidly hydratable and does not, when ingested or placed in an aqueous medium, significantly retard release of active ingredients from a substrate to which said coating is applied. Any of the strengthening polymers, plasticizers, surface active agents, coloring agents, etc, set forth herein, may be used in this embodiment.
For example, a first of these embodiments comprises an edible, hardenable, prompt release coating composition comprising about 5% to about 25%, more particularly about 5% to 10%, microcrystalline cellulose; about 10% to about 16%, more specifically about 14% to 16%, iota carrageenan. Such embodiments may also contain about 2% to about 10% of a surface active agent such as lecithin, about 35% to about 70% of a filler such as lactose as a filler and/or about 5% to about 10% propylene glycol alginate. These compositions my also contain 5% to 22% of a strengthening polymer such as polyvinylpyrrolidone or hydroxyethylcellulose. A mixture of strengthening polymers such as PVP and HEC may also be employed. In addition, from about 2% to 10% of a coloring agent may be included to provide pigmented coating composition. These compositions may also contain at least one filler such as maltodextrin, dicalcium phosphate, croscarmellose sodium and a mixture thereof. In this embodiment a reduced level of microcrystalline cellulose may be employed together with a small amount of surface active agent, with a high level of lactose used as the filler. Such a particular composition may comprise 5% to 10% microcrystalline cellulose, 14%to 16% iota carrageenan, 2% to 4% hydroxylated soy lecithin, 65% to 70% lactose, and may include from about 5% to about 10% propylene glycol alginate. The MCC:CGN weight ratio in this embodiment may be in the range of about 15:85 up to 60:40, or more particularly 15:85 to about 50:50; and the combined total of microcrystalline cellulose and carrageenan comprises from about 20% to about 40% of the coating composition. When PVP is used as the sole strengthening polymer, it is preferably employed at a level of about 5% to about 16%, whereas, when HEC is used as the sole strengthening polymer, it is preferably employed at a level of about 20% to about 22%. Specific examples of this embodiment are set forth as Examples 36-53 in Table 1 below.
A second of these embodiments of the coating composition of this invention comprises about 30% to about 40%, in particular about 33% to about 38%, microcrystalline cellulose; about 10% to about 20%, more specifically about 14% to about 17%, iota carrageenan, and about 12% to about 25%, more specifically about 12% to about 22% of a strengthening polymer such as hydroxyethylcellulose. These compositions may also further comprise about 5% to about 10%, more particularly 7% to 10%, of a surface active agent such as hydroxylated soy lecithin, about 8% to about 15% of a filler such as lactose, maltodextrin and a combination thereof, more particularly about 5% to about 10% lactose, and optionally from 5% to about 10% of a coloring agent. In this embodiment, if maltodextrin is the filler, it may be present at a level of about 3% to about 5%, specifically about 3%, in combination with a filler such as lactose at a level of about 7% to about 10%, more specifically about 8% to 9%. A suitable variation comprises compositions which, in addition to MCC, CGN and HEC, further comprise from about 30% to about 36, more particularly about 34% to about 35% polyethylene glycol and about 4% to about 5% coloring agent. In this group of compositions the weight ratio MCC:CGN may be in the range of about 60:40 to about 75:35, and the combined weight of MCC and CGN may be about 48% to about 60% by weight of the composition. It is also noted that this embodiment may contain a suitable coloring agent. This embodiment is illustrated in Examples 54-57 of Table 1 below.
Finally, a third embodiment of the composition of this invention comprises 30% to 40%, more specifically about 33% to about 38% microcrystalline cellulose, 10% to 20%, more specifically about 13% to about 16%, iota carrageenan, about 10% to about 15% of a strengthening polymer such as polyvinylpyrrolidone, and 30% to 40%, more specifically 33% to 36%, of a plasticizer such as polyethylene glycol. This embodiment may further comprise from about 2% to about 10% of a coloring agent. In this embodiment the weight ratio of MCC:CGN may be in the range of about 60:40 to about 75:25 and the combined weight of MCC and CGN comprises about 50% to about 60% by weight of the composition. A suitable coloring agent may be included in the compsotions of this embodiment. Example 58 of Table 1 illustrates this embodiment.
The viscosity of the hydrated formulation can be important. It ideally should be low enough to be pumped to a spray unit continuously and then sprayed evenly in a useful pattern onto the substrate being coated. A useful concentration of the dry ingredients in water on a weight percentage basis, therefore, may be about 6% to about 15%, advantageously 6.5% to 11%, preferably about 8% to about 11%. To assure uniformity of the coating composition, it may be preferable to maintain agitation of the aqueous dispersion during the entire period of its being sprayed onto the pharmaceutical or veterinary solid dosage forms, confectionery, seeds, animal feed, fertilizer, pesticide tablets, or food.
The preferred edible, hardenable, prompt release coating formulations of this invention may generally be prepared and used according to a simple procedure. A dry mixture of coprocessed microcrystalline cellulose/carrageenan powder or a dry blend of microcrystalline cellulose and carrageenan, and a strengthening polymer, such as hydroxyethylcellulose, polyethylene glycol or other acceptable plasticizer, optionally together with a solid filler such as maltodextrin, lactose, mannitol or the like, preservatives, and/or surfactants are blended to form dry coating composition. Addition of edible coloring agents, for example, a water-soluble dye or a pigment, may precede the hydration step required to prepare the final coating formulation. This dry mixture is then added slowly to the vortex of stirred, purified water. Stirring of this mixture is continued for a sufficient period to allow all of the components to be fully hydrated. If a colored coating material is required a water soluble dye or a pigment may also be added, preferably as a dispersion or solution, to the hydrated coating composition. Optionally surfactants, and/or plasticizers may also be added at this stage of the process.
In the formulations of microcrystalline cellulose and iota carrageenan, a simple propeller mixer provides adequate agitation for rapid hydration. The period of hydration may be as short as 0.5 hour. It may, and preferably should, be longer, but more than 3 hours is not believed to be necessary. Hydration can take place at room temperature or at elevated temperatures as high as 65.5xc2x0 C. (150xc2x0 F.), preferably at a temperature about 48.9xc2x0 C. (120xc2x0 F.). The time required for full hydration and the viscosity of the dispersion are both considerably reduced when the dispersion is prepared at an elevated temperature, but coating dispersions prepared at ambient temperature only require an increase in hydration time and a slight reduction in solids content to perform completely satisfactorily. As previously stated, these formulations may be prepared on the day preceding the coating operation, if that is more convenient; however, a period of mixing will be required to overcome the thixotropic behavior of a formulation which sets up during overnight storage. Unlike coating formulations based primarily on hydroxyalkyl ethers of cellulose, for example, HPMC, constant stirring of the microcrystalline and carrageenan-based formulations of this invention does not need to be continued throughout the coating procedure, but mixing may continue, if preferred.
Any commercial spray coater may be used to apply the coating. Examples of useful coaters are Vector High Coaters manufactured by Vector Corporation and Accela-Cota manufactured by Thomas Engineering. Equipment variables which one skilled in the art can manipulate to provide an elegant coating based on the microcrystalline cellulose and carrageenan materials, either coprocessed or dry blended, include inlet temperature, outlet temperature, air flow, speed of rotation of the coating pan, and the rate at which the coating formulation is pumped to the coater. It is important that the inlet and outlet temperatures be controlled so that they are high enough to efficiently dry the coating to prevent the tumbling action of the already-coated tablets from damaging the newly-applied coating before more coating is applied to the same tablets.
Hydroxyethylcellulose binds water more effectively than carrageenan does. Thus, the presence of the major amount of carrageenan in the formulations of this invention has a significant effect on the speed of drying of the edible coatings. Drying times are reduced considerably because of the presence of the carrageenan which dilutes the negative effect of HEC on drying time. Thus, in the case of low melting active pharmaceutical agents, for example, ibuprofen, the outlet temperature can be reduced and still provide short enough drying time to be commercially useful.
Hydroxyethylcellulose is particularly susceptible to clogging spray nozzles at high temperatures. An additional benefit provided by the formulations of this invention is the avoidance of clogging of the spray nozzles with dispersions being sprayed at high temperatures.
The level of coating applied to pharmaceutical or veterinary dosage forms is preferably between about 0.5% to about 4% by weight of the uncoated dosage form, more preferably about 2% to about 3.5%, by weight of the uncoated dosage form. This level of coating will provide an elegant, serviceable coating to a wide variety of dosage forms. To apply a heavier coating to tablets would not be economical, and it might adversely affect disintegration of the tablets or other properties. Too light a coating would not provide optimal properties normally expected from a coating, for example, improved friability or adequate taste masking.
For confections the coating level should be about 5% to about 10% by weight of the uncoated confection. Seed coatings should be in the range of about 3% to about 6% by weight of the uncoated seeds. Fertilizers and pesticide tablets and granules benefit from coating of 1% to about 3%, by weight of the uncoated granules or tablets.
From the following examples it has been shown that the coatings of the present invention may be applied successfully to tablets having a wide variety of active ingredients incorporated therein. For example, it has been reported that multivitamin tablets are difficult to coat because of the lipophilic surface properties of the vitamins. Similarly, ibuprofen is a challenging active ingredient to coat. Tablets comprising both of these difficult-to-coat active ingredients have been coated readily with the instant invention, providing elegant tablets. Additionally, the coatings have been applied to tablets which have been debossed with letters or a logo without bridging which would hide, or even obliterate, the debossed design.
An additional utility of the coating formulations of this invention is as a replacement for sugar coating of tablets. A sugar coating is applied primarily to increase the weight and/or size of the tablet, but this is an old art which presents numerous problems. It is, therefore, desirable to replace the traditional sugar coating with a more easily applied coating as shown in Example 26. This coating procedure has the additional advantage that no top coat is required to be applied as it is done with a sugar coating
Storage of coated tablets under ambient temperature and humidity and 40xc2x0 C. and 75% relative humidity for one to three months has demonstrated that no significant degradation has occurred. These tablets have disintegrated within the same length of time as the same batch of newly coated tablets did, and in each case provided dissolution rates and times substantially equal to those of the uncoated tablets used as a substrate for coating. This is an additional unexpected benefit of the coatings based on carrageenan and microcrystalline cellulose, and it differs from the known drawbacks of HPMC.
All components of the formulation are typically pharmaceutically acceptable, edible food grade materials.
The following examples, in which percentages are weight percent and tablet hardness is in Kiloponds (Kp), are provided to demonstrate the method of preparation and application of these elegant coatings, but they are not intended to be limiting as to amounts and the type of optional ingredients or the specific method of application of the tablet coating described herein.


A keyless electronic combination lock is adaptable for placement on a door, and operates independently of any other locks and without a central control system. The lock has a keypad for entering combinations, a random access memory (RAM) into which the entered data is loaded, a microprocessor, and an electrically erasable programmable read only memory (EEPROM). One or more entry codes for opening the lock are stored in the EEPROM and loaded into the RAM when the lock is powered-up. To open the lock, one of these codes or a master entry code must be entered at the keypad. Each entry code can be individually prevented from opening the lock by entering a corresponding individual lock-out code. All of the entry codes can be locked-out by entering a master lock-out code, by entering all of the individual lock-out codes or by throwing a dead bolt. The master entry code works regardless of the lock-out status and regardless of the dead bolt. To change lock-out status, the lock-out code is simply reentered. All of the codes can be changed as desired by entering a programming code, followed by an identifying code to tell the microprocessor which code to change. The programming code can be installed in the lock at the time of manufacturing the lock, and it can be changed during use in the field.

The present invention relates to an electronic combination lock suitable for industrial, commercial or residential use and capable of operating independently of a central control system. More particularly, the present invention relates to an electronic combination lock with entry codes, lock-out codes and a programming access code (programming code) for entering a programming mode in which the entry codes and lock-out codes can be changed.
An electronic combination lock is desirable because the lock can be opened without a key. The problem of losing keys is eliminated, and the cost of producing keys is also eliminated. A basic electronic combination lock is disclosed in "Combination Lock," Hobby Electronics, vol. 3, no. 12, October 1981, pp. 21-23. To open the lock, a five digit combination is entered using numbered push buttons. If the buttons have been pressed in the correct order, and in fairly rapid succession, and if the entered code matches a predetermined code, the lock will open. However, this article does not disclose a way to change the code.
U.S. Pat. No. 4,532,507 (to Edson et al) discloses an electronic combination security system (such as an alarm) in which the codes for turning off the alarm can be changed. There is a keypad at which a temporary pass code or a permanent pass code can be entered to turn off the alarm. In addition, entry of the permanent pass code will allow the temporary pass codes to be changed by using function keys separate from the keypad. The function keys must be pressed within 45 seconds of entry of the permanent pass code. However, in the system of the Edson et al patent, the permanent pass code cannot be changed. Therefore, an unauthorized person who discovers the permanent pass code can always gain entry, no matter how often the temporary codes are changed. Moreover, if it is desired to prevent temporary codes from working for a period of time, e.g. during a temporary period of increased security, the codes must be deleted from the system and later re-entered into the system.

Electronic combination of lock with changeable entry codes, lock-out and programming code


The present invention is an improved fitting and training system for a visual prosthesis. Fitting a visual prosthesis through automated means is challenging and fitting a visual prosthesis manually is tedious for clinician and patent, and provides great opportunity for error. A hybrid of computer controlled and manual fitting provides effective, efficient and controlled fitting process. The process includes testing a group of electrodes in random order by providing a prompt followed by stimulation and the patient responding if they saw a percept. After each set, a maximum likelihood algorithm is used to determine the next stimulation level, or if further stimulation is needed for each electrode.

Ever since 1755 when LeRoy passed the discharge of a Leyden through the orbit of man and caused a visual percept, there has been a fascination with electrically elicited visual perception. The general concept of electrical stimulation of retinal cells to produce these flashes of light or phosphenes has been known for quite some time. Based on these general principles, some early attempts at devising a prosthesis for aiding the visually impaired have included attaching electrodes to the head or eyelids of patients. While some of these early attempts met with some limited success, these early prosthetic devices were large, bulky and could not produce adequate simulated vision to truly aid the visually impaired.
In the early 1930's, Foerster investigated the effect of electrically stimulating the exposed occipital pole of one cerebral hemisphere. He found that, when a point at the extreme occipital pole was stimulated, the patient perceived a small spot of light directly in front and motionless (a phosphene). Subsequently, Brindley and Lewin (1968) thoroughly studied electrical stimulation of the human occipital (visual) cortex. By varying the stimulation parameters, these investigators described in detail the location of the phosphenes produced relative to the specific region of the occipital cortex stimulated. These experiments demonstrated: (1) the consistent shape and position of phosphenes; (2) that increased stimulation pulse duration made phosphenes brighter; and (3) that there was no detectable interaction between neighboring electrodes which were as close as 2.4 mm apart.
As intraocular surgical techniques have advanced, it has become possible to apply stimulation on small groups and even on individual retinal cells to generate focused phosphenes through devices implanted within the eye itself. This has sparked renewed interest in developing methods and apparatuses to aid the visually impaired. Specifically, great effort has been expended in the area of intraocular visual prosthesis devices in an effort to restore vision in cases where blindness is caused by photoreceptor degenerative retinal diseases such as retinitis pigmentosa and age related macular degeneration which affect millions of people worldwide.
Neural tissue can be artificially stimulated and activated by prosthetic devices that pass pulses of electrical current through electrodes on such a device. The passage of current causes changes in electrical potentials across visual neuronal membranes, which can initiate visual neuron action potentials, which are the means of information transfer in the nervous system.
Based on this mechanism, it is possible to input information into the nervous system by coding the information as a sequence of electrical pulses which are relayed to the nervous system via the prosthetic device. In this way, it is possible to provide artificial sensations including vision.
One typical application of neural tissue stimulation is in the rehabilitation of the blind. Some forms of blindness involve selective loss of the light sensitive transducers of the retina. Other retinal neurons remain viable, however, and may be activated in the manner described above by placement of a prosthetic electrode device on the inner (toward the vitreous) retinal surface (epiretial). This placement must be mechanically stable, minimize the distance between the device electrodes and the visual neurons, and avoid undue compression of the visual neurons.
In 1986, Bullara (U.S. Pat. No. 4,573,481) patented an electrode assembly for surgical implantation on a nerve. The matrix was silicone with embedded iridium electrodes. The assembly fit around a nerve to stimulate it.
Dawson and Radtke stimulated cat's retina by direct electrical stimulation of the retinal ganglion cell layer. These experimenters placed nine and then fourteen electrodes upon the inner retinal layer (i.e., primarily the ganglion cell layer) of two cats. Their experiments suggested that electrical stimulation of the retina with 30 to 100 uA current resulted in visual cortical responses. These experiments were carried out with needle-shaped electrodes that penetrated the surface of the retina (see also U.S. Pat. No. 4,628,933 to Michelson).
The Michelson '933 apparatus includes an array of photosensitive devices on its surface that are connected to a plurality of electrodes positioned on the opposite surface of the device to stimulate the retina. These electrodes are disposed to form an array similar to a "bed of nails" having conductors which impinge directly on the retina to stimulate the retinal cells. U.S. Pat. No. 4,837,049 to Byers describes spike electrodes for neural stimulation. Each spike electrode pierces neural tissue for better electrical contact. U.S. Pat. No. 5,215,088 to Norman describes an array of spike electrodes for cortical stimulation. Each spike pierces cortical tissue for better electrical contact.
The art of implanting an intraocular prosthetic device to electrically stimulate the retina was advanced with the introduction of retinal tacks in retinal surgery. De Juan, et al. at Duke University Eye Center inserted retinal tacks into retinas in an effort to reattach retinas that had detached from the underlying choroid, which is the source of blood supply for the outer retina and thus the photoreceptors. See, e.g., E. de Juan, et al., 99 Am. J. Ophthalmol. 272 (1985). These retinal tacks have proved to be biocompatible and remain embedded in the retina, and choroid/sclera, effectively pinning the retina against the choroid and the posterior aspects of the globe. Retinal tacks are one way to attach a retinal array to the retina. U.S. Pat. No. 5,109,844 to de Juan describes a flat electrode array placed against the retina for visual stimulation. U.S. Pat. No. 5,935,155 to Humayun describes a visual prosthesis for use with the flat retinal array described in de Juan.
In an implantable visual prosthesis system, the array attached to the retina is very rarely centered and perfectly oriented about the center of the visual field. When an implanted individual's eyes are at a neutral forward looking and level gaze, this is where the brain expects to see the image created by the electrical stimulation of the array. If the camera mounted on the subject's head is looking forward and level and the array is superior to the preferred location, this lack of correspondence between the real world scene and the location of perception will result in the image from the scene in the center of the camera image appearing to the subject as being inferior, which can cause confusion in the down stream visual processing systems such as the LGN (thalamus), superior colliculus, and visual cortex. Similar issues arise with temporal and nasal misalignment as well as rotation.
Further, imperfect position of the camera itself relative to the subject's head can cause the same problem.
In order to properly customize a visual prosthesis for each user, the amount of stimulation required to produce visual percepts (the stimulation threshold) must be measured for each electrode or group of electrodes. While many psychophysical methods for (general) threshold measurement exist, they suffer from known flaws, some of which become prohibitive when applied to threshold measurement for a visual prosthesis. For example, the Method of Constant Stimuli is an accurate but extremely inefficient method; its use is impractical for measuring stimulation thresholds for each individual electrode in a multi-electrode prosthesis. Methods of Adjustment suffer from habituation and anticipation errors; to mitigate the errors, multiple measurements must be made for each electrode, rendering the method impractical. Adaptive methods, in which the stimulus intensity is varied according to previous responses, can produce too narrow a range of stimulation intensity, resulting in a limited understanding of the true psychometric function.
Applicants have developed many methodologies for fitting an electrode array to a patient including: U.S. Pat. No. 8,271,091, for Visual prosthesis fitting, U.S. Pat. No. 8,195,301 Video configuration file editor for visual prosthesis fitting and related method; U.S. Pat. No. 8,190,267, for Fitting a neural prosthesis using impedance and electrode height; U.S. Pat. No. 8,180,454, for Fitting a neural prosthesis using impedance and electrode height; U.S. Pat. No. 7,908,011, for Visual prosthesis fitting; U.S. Pat. No. 7,818,064, for Fitting of brightness in a visual prosthesis; U.S. Pat. No. 7,738,962, for Fitting of brightness in a visual prosthesis; U.S. Pat. No. 7,493,169 for Automatic fitting for a visual prosthesis; U.S. Pat. No. 7,483,751, for Automatic fitting for a visual prosthesis. The preceding list includes both manual and automated fitting methods. Both have advantages and disadvantages. What is needed is a method that uses the best advantages of both manual and automatic fitting.


RNA aptamers and methods for identifying the same are disclosed. The RNA aptamers selectively bind coagulation factors, E2F family members, Ang1 or Ang2, and therapeutic and other uses for the RNA aptamers are also disclosed.

Thrombin belongs to the group of serine proteases and plays a central part in the blood coagulation cascade as terminal enzyme. Both the intrinsic and the extrinsic coagulation cascade lead, via a plurality of amplifying stages, to the production of thrombin from prothrombin. Thrombin-catalyzed cleavage of fibrinogen to fibrin then initiates blood coagulation and aggregation of platelets that, in turn, due to the binding of platelet factor 3 and coagulation factor XIII and a large number of highly active mediators, enhance thrombin formation.
To elaborate, the mechanism of blood coagulation normally occurs in a cascade of two possible routes. One of the routes, the extrinsic blood coagulation, starts with the liberation of thromboplastin and the activation of factor VII. Activated factor VII (i.e. factor VIIa) in turn activates factor X, followed by an activation of factor V and factor II (prothrombin). Factor IIa (thrombin) converts fibrinogen into fibrin at the end of the cascade.
The other route, the intrinsic blood coagulation, occurs via an activation of factor XII by contact with and subsequent activation of factor XI, factor IX and factor X in the presence of calcium and factor VIII, followed by an activation of thrombin (factor II) to activated thrombin (factor IIa) which triggers coagulation by cleaving fibrinogen to fibrin. Thus, activated thrombin (factor IIa) plays a role in both routes of the blood coagulation cascade.
Hitherto, there has been an intensive search for anticoagulants that can particularly be utilized in the treatment of cardiovascular disease, e g. septic shock, thromboses, embolisms, atherosclerosis and cardiac infarctions, furthermore in case of blood transfusions or following surgery. One method of suppressing the coagulation of blood is the direct administration of substances that modulate thrombin or other coagulation factors. The identification of such substances thus represents a long-felt and ongoing need in the art.
E2F refers to a family of transcription factors (also referred to herein as the "E2F family", which includes but is not limited to E2F-1, E2F-2, E2F-3, E2-F4, E2-F5 and E2-F6), and E2F activity is plays a role in a wide variety of proliferative events. E2F activity is controlled as the end result of G1 cyclin dependent kinase regulatory cascades that involve the Rb family of proteins. See, e.g., Sherr, C. J., Cell, 73:1059-1065 (1993); Hunter, T., Cell 75:839-841 (1993); Nevins J. R., Science, 258:424-429 (1992); Helin, K. and Harlow, E., Trends Cell Biol. 3:43-46 (1993); La Thangue, N. B., Trends Biochem. Sci. 19:180-114 (1994); Sherr, C. J.; Roberts, J. M., Genes Dev. 9:1149-1143 (1995); Weinberg, R. A. Cell 81:323-330 (1995); Harbour, J. W. and Dean, D. C., Genes and Development 14:2393-2409 (2000); and Black, A. R. and Azizkhan-Clifford, J., Gene 237:281-302 (1999). Thus, ligands that selectively bind to an E2F family member would play a role in the control of cell proliferation, which is of central importance to the proper development of a multi-cellular organism, the homeostatic maintenance of tissues, and the ability of certain cell types to respond appropriately to environmental cues.
Tie2 is an endothelial receptor tyrosine kinase (RTK) that is required for both embryonic vascular development and pathological angiogenesis. Tie2 is unique among RTKs in that it has two ligands with apparently opposing actions. Angiopoietin-1 (Ang1) is an activating ligand while Angiopoietin-2 (Ang2) is thought to be a naturally occurring antagonist for Tie2. Mice lacking Tie2 or Ang1 die midway through gestation due to abnormalities of vascular morphogenesis characterized by deficient recruitment of supporting smooth muscle cells and pericytes. Moreover, Ang1 promotes endothelial cell survival and blocks the increases in vascular permeability induced by vascular endothelial growth factor (VEGF), supporting a role for Ang1 in the stabilization and maintenance of the adult vasculature. In contrast, Ang2 is required for VEGF-mediated angiogenesis, and in the absence of endothelial mitogens Ang2 may induce vascular regression.
The exact mechanism of action of the Angiopoietins remains to be elucidated. For example, high-dose Ang2 can induce downstream activation of Akt and endothelial cell survival, suggesting that it does not simply exert a dominant negative effect on Tie2. The need for improved understanding of these ligands' function is particularly important in the study of tumor angiogenesis, as several studies have now shown that inhibition of Tie2 with a soluble receptor blocks tumor growth, angiogenesis, and metastasis. However, it is unclear whether these effects are due to inhibition of the effects of Ang1 or Ang2, since a soluble receptor would bind both ligands. Specific inhibitors of these ligands have the potential to more precisely modulate Tie2 signaling and serve as valuable therapeutic agents.
Aptamers can comprise single-stranded or double-stranded nucleic acids that are capable of binding proteins or other small molecules. Aptamers that have therapeutic value would most likely bind proteins involved in regulatory cascades. The presence of the aptamer would act as a sink for the protein factors, preventing the factors from carrying out their normal functions. To date, only a few aptamers are known.
It would be desirable to identify novel aptamers that bind to factors in the coagulation cascade, to an E2F family member, or to an angiogenesis factor. Indeed, among other applications, such aptamers have utility in the modulation of coagulation, cell proliferation or angiogenesis, and would thus meet a long-felt and continuing need in the art.
SUMMARY OF THE INVENTION
An RNA aptamer that selectively binds a coagulation pathway factor, that selectively binds an E2F family member, or that selectively binds an angiogenesis factor (e.g. Ang1 and/or Ang2) is provided in accordance with the present invention. Preferably, the binding affinity for the RNA aptamer for the coagulation pathway factor is represented by a dissociation constant of about 20 nanomolar (nM) or less. Optionally, the coagulation pathway factor is selected from the group consisting of thrombin, activated thrombin, IXa, X, Xa, VII, VIIIa and combinations thereof. Most preferably, the RNA aptamers selectively bind activated coagulation factors.
A method of modulating the biological activity of a coagulation pathway factor is also provided in accordance with the present invention. The method comprises: (a) administering to a warm-blooded vertebrate in need thereof an effective amount of an RNA aptamer that selectively binds a coagulation pathway factor, the RNA aptamer having a dissociation constant for the coagulation pathway factor of about 20 nM or less; and (b) modulating the biological activity of the coagulation pathway factor in the warm-blooded vertebrate through the administering of the RNA aptamer in step (a).
A method of treating cardiovascular disease in a warm-blooded vertebrate is also provided in accordance with the present invention. The method comprises administering an effective amount of an RNA aptamer that selectively binds a coagulation pathway factor, the RNA aptamer having a dissociation constant for the coagulation pathway factor of about 20 nM or less, to a vertebrate subject suffering from cardiovascular disease, whereby cardiovascular disease in the vertebrate subject is treated.
A method of modulating E2F activity in a warm-blooded vertebrate in which said modulation is desired is also provided. The method comprises: (a) administering to the warm-blooded vertebrate an effective amount of an RNA aptamer that selectively binds an E2F family member, the RNA aptamer having a dissociation constant for the E2F family member of about 20 nM or less; and (b) modulating E2F in the warm-blooded vertebrate through the administering of the RNA aptamer of step (a).
A method of modulating Ang1 or Ang2 activity in a warm-blooded vertebrate in which said modulation is desired is also provided. The method comprises: (a) administering to the warm-blooded vertebrate an effective amount of an RNA aptamer that selectively binds Ang1 or Ang2, the RNA aptamer having a dissociation constant for Ang1 or Ang2 of about 20 nM or less; and (b) modulating Ang1 or Ang2 in the warm-blooded vertebrate through the administering of the RNA aptamer of step (a).
A method of identifying a ligand to a target from a candidate mixture of potential ligands is also provided in accordance with the present invention. Products, i.e., ligands, produced or identified via the method are also provided in accordance with the present invention.
In a preferred embodiment the method comprises: (a) preparing a candidate mixture of potential ligands; (b) contacting the candidate mixture with a target in a lower stringency buffer, wherein ligands having increased affinity to the target relative to the candidate mixture bind to the target; (c) removing unbound candidate mixture; and (d) collecting the ligands that are bound to the target to produce a first collected ligand mixture. More preferably, the method further comprises: (e) contacting the first collected ligand mixture with the target in a higher stringency buffer, wherein ligands having increased affinity to the target relative to the first collected ligand mixture bind to the target; (f) removing unbound ligands; and (g) collecting the ligands that are bound to the target to produce a second collected ligand mixture to thereby identify ligands to the target. Even more preferably, ligands in the first or second collected ligand mixture are enriched or expanded by any suitable technique, e.g. amplification, prior to contacting the first collected ligand mixture with the target in the higher stringency buffer, after collecting the ligands that bound the target in the higher stringency buffer, or both. Optionally, the contacting and expanding or enriching steps are repeated as necessary to produce a desired ligand. Thus, it is possible that the second collected ligand mixture can comprise a single ligand.
Another embodiment of a method of identifying a ligand to a target from a candidate mixture of potential ligands is provided in accordance with the present invention. Products, i.e., ligands, produced or identified via the method are also provided in accordance with the present invention.
The method preferably comprises: (a) providing a target selected from a first species of organism; (b) preparing a candidate mixture of potential ligands; (c) contacting the candidate mixture with the target, wherein ligands having increased affinity to the target from the first species of organism relative to the candidate mixture bind to the target from the first species of organism; (d) removing unbound candidate mixture; (e) collecting the ligands that are bound to the target from the first species of organism to produce a first collected ligand mixture for the target; (f) contacting the first collected ligand mixture with a target from a second species of organism, the target from the second species having at least a portion thereof that is substantially homologous to the same portion in the target from the first species, wherein ligands having increased affinity to the target from the second species relative to the first collected ligand mixture bind to the target; (g) removing unbound first collected ligand mixture; and (h) collecting the ligands that are bound to the target from the second species of organism to form a second collected ligand mixture thereby identify ligands to the target.
Preferably, ligands in the first or second collected ligand mixture are enriched or expanded by any suitable technique, e.g. amplification, prior to contacting the first collected ligand mixture with the target from the second species of organism, after collecting the ligands that bound the target from the second species of organism, or both. Optionally, the contacting and expanding or enriching steps are repeated as necessary to produce a desired ligand. Thus, it is possible that the second collected ligand mixture can comprise a single ligand.
It is therefore an object of the present invention to provide novel RNA aptamers that selectively bind coagulation factors, an E2F family member, Ang1 or Ang2. The object is achieved in whole or in part by the present invention.
An object of the invention having been stated hereinabove, other objects will become evident as the description proceeds, when taken in connection with the accompanying Figures and Laboratory Examples as best described herein below.


An ultra wide band (UWB) pulse train generator, which includes a trigger signal controller for sequentially outputting n trigger signals for triggering generation of UWB pulse train in accordance with n bit data to be transmitted at a predetermined time period, a latch unit for loading in parallel the n bit data to be transmitted and sequentially toggling the loaded n bit data by one bit whenever the trigger signal is input from the trigger signal controller, and a pulse train generator for generating polarity pulses corresponding to a direction of an edge transition in accordance with toggling on each output signal of the latch unit.

1. Field of the Invention
The present invention relates generally to a UWB (Ultra Wide Band) communication method, and more particularly, to a UWB train generator.
2. Description of the Related Art
Conventionally, a UWB wireless communication technique was used only for military affairs under control of the U.S. Department of Defense. Recently, however, UWB has been opened to non-military organizations regulated by the FCC (Federal Communications Commission). Such a UWB wireless communication technique uses wide frequency band of GHz. In addition, as compared with that of wireless LAN (Local Area Network) of IEEE 802.11 or Bluetooth, the UWB technique has a faster high speed transmission and lower power consumption. Accordingly, the UWB wireless communication technique is considered as a core technique in an advanced network field.
In UWB wireless communication, a transmitter requires a UWB pulse train generator for generating UWB pulse train depending on a data to be transmitted. Additionally, in such a UWB pulse train generator, it is preferable to implement the UWB pulse train generator to be easily integrated with a digital SOC (system on chip).


A magnetic resonance imaging method comprises performing imaging where more than one polarizing magnetic field strength is used during scanning and processing at least one image resulting from the scanning to yield an enhanced contrast image.

Nuclear Magnetic Resonance (NMR) imaging, or Magnetic Resonance Imaging (MRI) as it is commonly known, is a non-invasive imaging modality that can produce high resolution, high contrast images of the interior of a subject. MRI involves the interrogation of the nuclear magnetic moments of a sample placed in a strong magnetic field with radio frequency (RF) magnetic fields. During MRI the subject, typically a human patient, is placed into the bore of an MRI machine and is subjected to a uniform static polarizing magnetic field B0 produced by a polarizing magnet housed within the MRI machine. Radio frequency (RF) pulses, generated by RF coils housed within the MRI machine in accordance with a particular localization method, are typically used to scan target tissue of the patient. MRI signals are radiated by excited nuclei in the target tissue in the intervals between consecutive RF pulses and are sensed by the RF coils. During MRI signal sensing, gradient magnetic fields are switched rapidly to alter the uniform magnetic field at localized areas thereby allowing spatial localization of MRI signals radiated by selected slices of the target tissue. The sensed MRI signals are in turn digitized and processed to reconstruct images of the target tissue slices using one of many known techniques.
When a substance, such as human tissue is subjected to the static polarizing magnetic field B0, the individual magnetic moments of the spins in the tissue attempt to align with the static polarizing magnetic field B0, but precess about the static polarizing magnetic field B0 in random order at their characteristic Larmor frequency. The net magnetization vector lies along the direction of the static polarizing magnetic field B0 and is referred to as the equilibrium magnetization M0. In this configuration, the Z component of the magnetization or longitudinal magnetization MZ is equal to the equilibrium magnetization M0. If the target tissue is subjected to an excitation magnetic field B1, which is in the x-y plane and which is near the Larmor frequency, the longitudinal magnetization MZ may be rotated, or "tipped" into the x-y plane to produce a net transverse magnetic moment MXY. When the excitation magnetic field B1 is terminated, relaxation of the excited spins occurs, with a signal being emitted that effects the magnitude of radiated MRI signals. The emitted signal is received and processed to form an image.
In particular, when the excitation magnetic field B1 is terminated, the longitudinal magnetization MZ relaxes back to its equilibrium. The time constant that describes how the longitudinal magnetization MZ returns to its equilibrium value is commonly referred to as the spin lattice relaxation time T1. The spin lattice relaxation time T1 characterizes the time required to reduce the difference between the longitudinal magnetization MZ and its equilibrium value M0 to zero.
The net transverse magnetic moment MXY also relaxes back to its equilibrium when the excitation magnetic field B1 is terminated. The time constant that describes how the transverse magnetic moment MXY returns to its equilibrium value is commonly referred to as the transverse relaxation time or spin-spin relaxation time T2. The transverse relaxation time T2 characterizes the time required to reduce the transverse magnetic moment MXY to zero. Both the spin lattice relaxation time T1 and the transverse relaxation time T2 are tissue specific and vary with concentration of different chemical substances in the tissue as well as with different microstructural features of the tissue. Variations of the spin lattice relaxation time T1 and/or the transverse relaxation time T2 from normal can also be indicative of disease or injury.
Like many diagnostic imaging modalities, MRI can be used to differentiate tissue types, e.g. muscles from tendons, white matter from gray matter, healthy tissue from pathologic tissue. There exist many different MRI techniques, the utility of each depending on the particular tissue under examination. Some techniques examine the rate of tissue magnetization, while other techniques measure the amount of bound water or the velocity of blood flow. Often, several MRI techniques are used together to improve tissue identification. In general, the greater the number of tests available the better chance of producing a correct diagnosis.
In some instances contrast agents may be used to emphasize certain anatomical regions. For example, a Gadolinium chelate injected into a blood vessel will produce enhancement of the vascular system, or the presence and distribution of leaky blood vessels. Iron-loaded stem cells injected into the body and detected by MRI, will allow stem cell migration and implantation in-vivo to be tracked. For a contrast agent to be effective the contrast agent must preferentially enhance one tissue type or organ over another.
Clinical MRI machines commonly utilize superconducting windings to produce a strong, static polarizing magnetic field B0 that remains at a fixed magnetic field strength. Because the magnetic field strength is fixed, these clinical MRI machines are unsuitable for use in tests that measure the dependence of parameters on the strength of the applied magnetic field. There are characteristics of certain tissues, materials or contrast agents that would be better identified or quantified if it were possible to measure the variation of the MRI properties of those tissues, materials or contrast agents over a range of polarizing magnetic field strengths, not just at a single polarizing magnetic field strength.
Accordingly, there is a continuing need for improvements in MRI. It is therefore an object to provide a novel system and method for producing image contrasts in magnetic resonance imaging.

System and method for magnetic resonance imaging


A connector for connecting a pipe of small diameter in which chamber in communication with a flow bore of a cylindrical connection is are perforated axially on the side of the top end of a connector main body recessed windows are disposed to the circumferential wall of the chamber and the circumferential wall at the rear end thereof is formed as an engaging wall, the rear end of the chamber is engaged by the engaging wall and a separate receptacle member having a pair of finger walls protruded being opposed to each other in the inserting direction thereof, in which connection is made by engaging the finger wall to an annular bulged wall disposed near the connection end of a pipe incorporated to the inside of the connector main body in a state of connecting the pipe, wherein the receptacle main body 8 comprises an annular circumferential wall 7 to be attached on the outer circumferential surface at the rear part of the connector main body 1 and a pair of finger walls 5 each of a square block form being disposed to each other and protrudes substantially in perpendicular to the axial direction, and having a tapered surface 5' diverging at the protruding rear end which the formed to the inside at the top end in contiguous with a resilient arm 7' extending circumferentially in the circumferential wall along a slit 12.

1. Field of the Invention
The present invention concerns an improvement for a structure of a connector for connecting metal or synthetic resin tubes of a relatively small diameter of less than about 20 mm, which are often disposed and used as a pipe-line for supplying oils and gases generally in automobiles or various kinds of machineries and equipments (hereinafter simply referred to as a pipe).
2. Description of the Prior Art
The connector of the aforementioned type has been constituted so far, for example, as shown in FIG. 23 such that a receptacle member 22 having a pair of resilient finger walls 23 which are situated at a portion forward in the inserting direction of circumferential wall of around a through hole at the axial center of a connector main body 21 and protruded being inclined forward is engaged to a stepped portion 24 of a connector main body disposed to the rear annular circumferential wall 23' at an engaging wall 21 in the rear portion of the connector main body 21 and, in this state, a pipe P.sub.0 is connected by engaging the finger wall 23 to an annular bulged wall P.sub.0 ' of the incorporated pipe P.sub.0.
Further, a connector in Japanese Patent Publication Sho 64-10711, is constituted such that a retaining element is inserted into the housing (connector main body), a jaw end of the retainer element is urged to expand outward by the annular surface of a pipe as it is inserted thereby enabling the insertion of the pipe and, when the jaw end passes over the annular surface, the Jaw end returns to its inward home position and abuts against the engaging wall of the pipe to prevent it from slipping off.
Furthermore, a connector as shown, for example, in Japanese Utility Model Laid-Open Hei 1-136791 has such a constitution that incorporating at the inside of a connector main body upon pipe connection is visually observed reliably by the deformation of a sensing finger incorporated in the connector main body as an observation means.
However, in the prior art shown in FIG. 23, since the connector is engaged to the annular bulged wall P.sub.0 ' due to the snapping function of the finger wall 23 in the connection state of the pipe P.sub.0, the engagement between them is somewhat unreliable. Then, when tension is exerted on the pipe P.sub.0 due to the fatigue for long period of time of the finger wall 23, the top end of the finger wall is expanded to cause disengagement, thereby making the connection instable and tending to induce undesirable leakage.
Further, in the pipe connector as described in Japanese Patent Publication Sho 64-10711, since it is necessary that the diameter of the retainer is larger as compared with that of the pipe in order to obtain a snapping action of a resilient finger of the retainer upon inserting the pipe and the length of the retainer is also increased, the size of the connector main body is made larger which is not suitable to disposition in a narrow space. Further, since the clipping lug end for detaching the pipe is protruded, it involves such a problem that the pipe may be detached accidentally due to interference with other parts, etc.
Furthermore, in the prior art as described in Japanese Patent Utility Model Laid-Open Hei 1-136791, since the sensing finger is small, it is difficult to confirm connection in a narrow or dark place. In addition, since the sensing finger deforms to some extent even if the locked state is insufficient and it leads to a problem of erroneously recognizing this as complete locking.

Connector for connecting a pipe of small diameter with block expansion prevention surfaces


A cutting insert includes an upper surface, a lower surface opposite to the upper surface, a first side surface and a second side surface configured to connect the upper surface and the lower surface, a mounting hole formed to penetrate the upper surface and the lower surface, a first cutting edge formed at an edge of the upper surface that meets the first side surface, and a second cutting edge formed at an edge of the upper surface that meets the second side surface. The lower surface includes a ridge portion, and a first inclined surface and a second inclined surface positioned on both sides of the ridge portion as a boundary. When looking at the lower surface, the first inclined surface and the second inclined surface have asymmetric shapes with respect to the ridge portion.

An end mill (also referred to as an 'end milling cutter') is known as an example of a rotary cutting tool used for milling. Unlike a drill used only for perforation, the end mill has cutting edges on its periphery and tip, and may be used for groove cutting, side cutting and the like of a workpiece. Among end mills, the ball nose end mills having a round front end are divided into a solid ball nose end mill in which an end mill body and a ball nose are integrally formed and an indexable ball nose end mill in which a cutting insert is replaceably mounted on an end mill body (or a tool body), depending on whether the cutting insert is mounted.
In the indexable ball nose end mill, the cutting insert is usually coupled to the tool body by a screw. The screw provides only a fastening force in a direction perpendicular to the cutting insert (e.g., a direction in which the cutting insert is pressed against the tool body as the screw is tightened). As such, the screw does not sufficiently support the reaction force generated by the cutting force. In particular, the cutting insert used in the ball nose end mill has an arc-shaped edge for curved surface cutting. Accordingly, if the cutting insert is fixed to the tool body only with a screw, then the cutting insert rotates around the screw due to the reaction force generated by the cutting force during machining. This leads to a problem wherein the cutting insert cannot be firmly fixed to the tool body. That is, there is a problem in that the deformation of the screw or the breakage of the cutting insert may be caused.
To solve such conventional problems, there is disclosed a ball nose end mill in which a cutting insert and a mounting structure of an insert pocket provided in a tool body are changed. According to such ball nose end mill, a concave groove is formed on the lower surface of the cutting insert, and a protrusion having a shape corresponding to the concave groove is formed on the bottom surface of the insert pocket on which the cutting insert is seated. The cutting insert is fixed in a direction perpendicular to the insert pocket by a screw, and is firmly fixed so that its rotation on the insert pocket is prevented by the engagement between the protrusion and the concave groove. However, in the case of the ball nose end mill having such mounting structure, an additional manufacturing process is required to form the protrusion on the bottom surface of the insert pocket, and a grinding process is additionally required to precisely machine the concave groove on the lower surface of the cutting insert. This may result in an increase in the manufacturing cost of the cutting insert. Moreover, the cutting vibration or cutting force is concentrated to these coupling portions in a state in which the protrusion is coupled to the concave groove. As such, there are drawbacks since the fatigue increases and the cutting insert is likely to be damaged.

Cutting insert and rotary cutting tool including same


The present invention discloses a thermal block unit for thermal treatment of samples comprising temperature regulating units, temperature sensors for measuring temperature at different locations of the thermal block unit, a converter for converting signals from the temperature sensors into digital signals and a thermal block interface for communicating with an instrument.

Devices for the thermal treatment of samples or reaction mixtures in a controlled way are used in several fields of chemistry and biochemistry. For example, it is known that chemical reaction rates are proportional to temperature. Also, the working time or shelf life of a biological samples or laboratory reagents can be increased by keeping the substance at an optimal temperature. Since labor time as well as reagents are expensive) it is desirable to increase the throughput of production and analysis, while at the same time, to minimize the necessary reaction volumes. In general such devices or instruments have a thermal block made of e.g. metal, composite, ceramic or the like, that is in thermal contact with the sample under investigation so that the temperature of the sample is affected by the temperature of the thermal block.
Particularly, a strong need for systems capable of cycling a sample through a range of temperatures, i.e. thermal cyclers, became apparent with the advent of the Polymerase Chain Reaction (PCR), a technique which revolutionized the field of health care and molecular diagnostics.
PCR enables isolation of genomic material, sequencing and the detection of genetic diseases, recombinant DNA techniques, genetic fingerprinting and paternity testing. Viral DNA can likewise be detected by PCR and the amount of virus ("viral load") in a patient can be quantified by PCR-based DNA quantitation techniques or quantitative PCR.
Because the amount of product produced by PCR roughly correlates to the amount of starting material, PCR can be used to estimate the amount of a given sequence that is present in a sample and because of the high sensitivity, virus detection may be possible soon after infection and even before the onset of disease symptoms, thus giving a significant lead in treatment. Quantitative PCR is also useful for determining gene expression levels. In cells, each gene is expressed through the production of messenger RNA (mRNA), which is then used to create a protein corresponding to the gene. The amount of mRNA in the cell for a given gene reflects how active that gene is. By using reverse transcription to produce DNA complementary to the mRNA (called cDNA) and subsequently using PCR to amplify these molecules, the amount of DNA produced for each gives a rough measure of the underlying expression for that gene.
Real-time PCR is a special form of quantitative PCR. By this technique it is possible to simultaneously amplify and quantify a specific part of a given DNA molecule. The DNA is quantified after or during each round of amplification. Two common methods of quantification are the use of fluorescent dyes that intercalate with double-strand DNA, and modified DNA oligonucleotide probes that generate fluorescence at a certain point during the cycle.
PCR specificity and yield as well as throughput are directly related to the ability of the thermal-cycling system to rapidly and accurately arrive at and maintain reaction temperatures for an array of samples in parallel, e.g. in a multiwell plate in contact with a metal thermal block. Heating and cooling is normally achieved by means of temperature regulating units such as thermoelectric coolers (TECs) also called Peltier elements as well as a heat sink. One problem in the prior art is that differences in sample temperature may be generated by non-uniformity of temperature from place to place within the sample block. Temperature gradients may exist within the material of the block, causing some samples to have different temperatures than others at particular times in the cycle. Further, since there are delays in transferring heat from the sample block to the sample, those delays may differ across the sample block. These differences in temperature and delays in heat transfer, commonly referred to as well-to-well inhomogeneity, may cause the yield of the PCR process to differ from sample vial to sample vial. To perform the PCR process successfully and efficiently, and to enable quantitative PCR, these time delays and temperature errors must be minimized to the greatest extent possible.
One state of the art instrument currently available on the market, is the LightCycler® 480 Real-Time PCR System from Roche Diagnostics. This instrument reduces the problem above thanks to a special architecture of the thermal block unit, which comprises also a so-called Therma-Base™ unit, located beneath the Peltier elements, for improved heat transfer and distribution to all samples within a multiwell plate. The heat sink below the Therma-Base® unit features a maximized inner surface area to facilitate rapid heat absorption.
In U.S. Pat. No. 7,133,726B1, it is proposed instead to use a perimeter trench for the heat sink and a perimeter heater around the metal thermal block to reduce edge losses as well as a pin at the center of the assembly establishing a thermal path from the sample block to the heat sink in order to compensate for thermal gradients.
A problem in the state of the art is however represented by the inefficient control of the thermal block unit. Data measured within the thermal block unit, e.g. temperature values, are sent to a controller unit of an instrument and the instrument controls the thermal block unit. An instrument or thermal block test is typically carried out only when the instrument is turned on. One disadvantage is that only a limited number of data are processed, thus making it difficult to react promptly to errors and/or failures and/or any deviation from the normal or expected functioning of the temperature regulating units. Also, data transfer may be unreliable due to the possible influence of the electric connections, e.g. the electric resistance of the cables itself, cracks or line interruptions between thermal block unit and instrument.