CA1223510A - Analytical element having improved spreading zone and method of use - Google Patents

Abstract

ANALYTICAL ELEMENT HAVING IMPROVED
SPREADING ZONE AND METHOD OF USE

Abstract of the Disclosure A dry element capable of use for the analysis or transport of liquids comprises an isotropically porous zone which comprises a blush polymer having dispersed wherein a particulate material and a nonpolymeric heterocyclic, aliphatic or carbocyclic quaternary ammonium compound comprising at least 12 carbon atoms therein. This compound is present in the porous zone in an amount of at least about 2 weight percent, based on the dry weight of the blush polymer. The porous zone of this element has improved cohesive strength which improves its resistance to abrasion. The element can be used to determine an analyte in an aqueous liquid, such as biological fluids.

Description

us ANALYTICAL ELEMENT HAVING IMPROVED
SPREADING ZONE AND METHOD OF USE
Field of the Invention This invention relates to dry element useful for the chemical analysis or transport of water, foodstuffs and biological liquids. In particular, it relates to dry analytical elements which hove improved m~nufactursbility. This invention also relates to a method of using such elements for determination of an annihilate.
Background of the Invention Chemical analysis of water, foodstuffs like milk and biological fluids such us serum end urine it often desirable or necessary. Various ~n~lytic~l element to facilitate such ~n~lyse~ ore known.
Generally, ugh elements include regent thereinafter termed inter~ctlve composition), for substance under analysis (hereinafter termed annihilate). The interactive composition, upon contact with a liquid simple containing the ~n~lyte, effects a detectable change in response to the presence of the analyze. For example, such 8 detectable change can be the formation or dls~ppear~nce (e.g.
reduction of 8 detectable species, e.g. dye. Such 8 change con be determined us it occurs (i.e. rote Assay), or filter a certain time (i.e. endpoint Sue).
Recently, much work ha been directed toward developing dry analytical elements useful in diagnostic chemical analysis of biological fluids which provide highly quantitative results quickly end conveniently. For example, U.S. Potent 3,992,158 ~35

-2 -(issued November 16, 1976 to Przybylowicz et at) describes integral analytical elements which are a significant advance in the clinical chemistry art.
These elements generally contain an isotropic ally porous spreading zone, also known as a fluid metering zone, which uniformly distributes a liquid sample throughout the zone. A variety of useful spreading zones are described in that reverence. In one embodiment, the spreading zone is composed of a lo "blush" polymer which has a particulate material dispersed therein. Such elements are manufactured using conventional coating and drying techniques, and subsequently slitting or chopping the dried layer.
However, during such mechanical handling operations, it has been observed that the dry blush polymer layers are sensitive to abrasion, and objectionable flaking or dust is generated particularly during slitting and chopping operations. Not only is the dust an environmental hazard, but this abrasion sensitivity often leads to unacceptable product when the dried spreading layer has ragged edges or crumbles during slitting. Attempts have been made to improve spreading layer abrasion resistance by increasing the blush polymer coverage or by reducing I the amount of non-solvent (defined hereinbelow) used during the coating operations. These efforts, however, have had limited success; and undesirably resulted in a loss of zone porosity or void volume and attendant long liquid spreading times.
U.S. Patent 3,014,810 (issued December 26, 1961 to Dybalski et at) relates Jo pigment compositions which can be incorporated in nonporous rubbers and plastics Jo provide nonporous finished products having improved hardness, tensile strength and other physical properties. The pigment compositions described therein include a mixture of two amine surface active agents in a specified ratio. These surEactan~s allegedly reduce pigment agglomeration during storage and shipping and improve pigment dispersion in the hard rubbers and plastics.
Toe first agent is a ~uaternary ammonium compound and the second is a tertiary amine. The mixture of sur~ace-active agents described therein is not suitable for use in analytical elements containing various chemical reagents because the tertiary amine of the mixture is highly reactive with respect to reagents commonly employed in clinical chemistry analyses. There is no suggestion in this reference of any means for improving the cohesive strength of porous materials such as porous spreading layers used to transport liquids.
Hence, there is a need in the art for a porous spreading zone material which has improved cohesive strength and abrasion resistance but which retains desirable spreading properties and is inert to chemical reagents which may be incorporated therein.
Summary of the Invention The present invention provides an improved dry element which overcomes the abrasion problem observed in known elements. In particular, the elements ox this invention comprise an isotropic ally porous zone comprising a blush polymer which has improved cohesive strength, and hence improved abrasion resistance. These elements are much more easily manufactured as they have less tendency to flake or chip during slitting and chopping operations. Because of the improved abrasion resistance, reduction of hazardous dust and defective product produced during manufacture are achieved.
These improvements are achieved by incorporating one or more of a particular class of qua ternary ammonium compounds within the porous zone of the element.

~35~) Therefore, in accordance with this invention 9 a dry element for the analysis or transport of an aqueous liquid comprises an isotropic ally porous zone which comprises a blush polymer having dispersed therein a particulate material and a non polymeric heterocyclic, aliphatic or carbocyclic qua ternary ammonium compound comprising at least 12 carbon atoms therein. This compound is present in the zone in an amount of at least about 2 weight percent, based on the dry weight of the blush polymer.
In preferred embodiments, these elements are dry analytical elements for determination of an analyze, and also contain an interactive composition for the analyze.
This invention also provides a method for the determination of an analyze in an aqueous liquid using the dry analytical element of this invention.
This method comprises the steps of:
A. physically contacting a sample of the liquid together with an interactive composition for the analyze and an element described hereinabove to provide a detectable change; and B. measuring the detectable change.
I Detailed Descry on of the Invention The elements of this invention comprise an isotropic ally porous zone. This zone is often termed a spreading or metering zone in the art because of its ability to spread, meter or transport applied liquid samples rapidly therein. Isotropic porosity means that the zone is substantially porous to aqueous liquid in all directions within the zone. It will be understood that the degree of porosity can be variable within the zonk. In the elements of this invention, it is useful to have a void volume of at least about 25 percent Do the total zone volume, and void volumes of 50 to I percent may be desirable in certain instances. As can be appreciated, void volume within the zone can be controlled, for example, by the selection ox consent materials, or by varying the solvents or drying conditions during preparation as described in more detail hereinbelow.
The isotropic ally porous zone can be in any suitable location in the element. It can be a self-supporting matrix so that the zone alone, or in lo combination with other zones, is structurally strong enough to be self-supporting. Preferably, however, the zone is carried on a suitable substrate (or support hereinafter) and is the outermost layer on that support so that the liquid sample to be assayed comes into contact with this porous zone before any other part of the element. For example, an element can comprise one or more zones or layers which perform functions other than spreading but which element has the described porous layer as an outermost spreading layer to bring a sample of an aqueous liquid into contact with those other zones or layers. Alternatively, the element can have one or more other spreading or metering zones, and the particular blush polymer porous zone described herein can be between these zones or between the support and all of the other zones. In such embodiments, another spreading zone is the outermost zone in the element.
The isotropic ally porous zone essential to the present invention comprises a "blush" polymer.
Such polymers are generally formed on a support by dissolving a polymer in a mixture of two liquids, one of which is a lower boiling, good solvent for the polymer and the other which is a higher boiling non-solvent or pool solvent for the polymer. Such a polymer solution is then coated on the support and drip under controlled conditions. The lower boiling solvent evaporates more readily and the coaxing becomes enriched in the liquid which it the poor solvent As evaporation proceed under proper conditions, the polymer forms an i~otropic~lly porous layer I that term it used herein. Many different polymer can be used, tingly or in combination, for preparing Luke polymer porous zone, including polycarbonate~, polyamides, polyureth~nes end cellulose esters. Cellulose acetate and polyurethane are preferred polymers in the practice of this invention either tingly or in a mixture. The blush polymer (including mixtures of polymers) is enroll present in the porous zone of this invention at a coverage of from about 2 to about 40, and preferably from about 5 to bout 20, gym Solvent mixture for preparing blush polymer are well known in the art. Further detail for preparing blush polymer are provided in the art, e.g. in U.S. Potent

3,~92,158 (noted hereinabove).
The blush polymer porous zone of the eye-mints of this invention has dispersed therein one or more fine particulate material. Various type of particulate mutter, all desirably chemically inert to the ~nalytes sod the interactive compositions used in the Sue end generally in the form of a fine pow-don, ore useful. Pigments, such as titanium dioxide,b~rium sulfite, zinc oxide, lead oxide, etc. ore use-fur with titanium dioxide End barium sulfate being preferred. Other deslrRble particulate materials in-elude silicate such us di~tom~ceous earth, micro-crystalline colloidal materials derived from n~tur~lor synthetic polymers, and resinous or glue Bud.
Details of other useful particulate Metro no methods of incorporating such in the blush polymer porous zone Are provided, for example, in U.S. Potent 3,9~2,158 (noted hereinabove. The coverage of the ~Z3S~(~

particulate material in the porous zone is generally from about 20 to about 250, and preferably from about 40 Jo 120, g/m2.
In the elements of this invention, the weight ratio of particulate material to blush polymer is from about 2:1 to about 20:1, and preferably from about 6:1 to about 15:1.
The advantage of improved cohesive strength exhibited by the elements of this invention is lo obtained by the incorporation of one or more non polymeric he~erocyclic, aliphatic or carbocyclic qua ternary ammonium compounds, each compound comprising at least 12 carbon atoms therein in the porous spreading zone. Such a compound is present in the spreading zone in an amount of at least about 2, preferably from about 8 to about 30, and more preferably from about 10 to about 25, weight percent, based on the dry weight of the blush polymer in the zone. As used herein, the term "non polymeric" means that the compounds are not composed of repeating qua ternary ammonium cat ionic moieties and generally have a molecular weight of less than about 4000, and preferably less than about 2000.
As used herein, the term "heterocyclic"
qua ternary ammonium compound refers to an organic cat ionic ammonium compound having at least one heterocyclic moiety. The cat ionic charge can be either in the heterocyclic moiety or in another portion of the molecule. the heterocyclic moiety can be aromatic or non aromatic, and can contain such heteroatoms as nitrogen, oxygen, sulfur, and selenium atoms. Generally, the heterocyclic moiety has from 5 to 20 atoms in its backbone.
The term 1'alipha~ic" qua ternary ammonium compound refers to an organic cat ionic ammonium compound which contains four aliphatic, or open-chain, moieties attached to the qua ternary ~235~

ammonium atom. The aliphatic moieties contain from 1 to 30 carbon atoms and can have oxygen atoms interspaced along the chain, provided such compound comprises at least 12 carbon atoms. Preferably, at least one of the aliphatic moieties has at least 6 carbon atoms, and Gore preferably, at least one moiety comprises at least 12 carbon atoms.
The term "carbocyclic" qua ternary ammonium compound refers to an organic cat ionic ammonium compound which contains at least one carbocyclic moiety attached to the qua ternary ammonium atom.
Such carbocyclic groups include cycloalkyls generally of 5 to 20 carbon atoms, cycloalkenyls generally of 5 to 20 carbon atoms, cycloalkynyls generally of 5 to 20 carbon atoms, and Aurelius generally of 6 to 14 carbon atoms, it the cyclic backbone.
In preferred embodiments of this invention, heterocyclic, aliphatic and carbocyclic qua ternary ammonium compounds can be represented by the structures via and b):
Ian , R2 It.,-- R2- "
I' I+
Al- N - R3 X- or Al_ = N _ = R3 I

wherein _ = represents a single or double pond;
Al, R2, R3 and R., are independently substituted or unsubstituted alkyd, preferably of 1 to 30 carbon atoms (e.g. methyl, ethyl, chloroethyl, isopropyl, decal, dodecyl, alkyd groups substituted with an alkylcarbonamido group, such as Cl7H35CONH-, alkyd groups substituted with a fluorocarbonsulfonamido group, such as C~F~7S02NH-, etc.);

MU
g (alkylene-oxy~ H
wherein the alkaline comprises 2 to 6 carbon atoms and is substituted or unsubstituted, and n is an integer of 1 to 50 (erg e~hyleneoxy, propoxy, etc.);
substituted or unsubstituted aureole, preferably of 6 to 14 carbon atoms in the aromatic backbone (ego phenol, xylyl, naph~hyl, p-methoxyphenyl, eta.);
substituted or unsubstituted cycloalkyl, preferably of 5 to 20 carbon atoms in the carbocyclic ring (e.g.
cyclopentyl, cyclohexyl, etc.); substituted or unsubstituted alkaryl, preferably of 7 to 30 carbon atoms in the backbone (e.g. bouncily, 3-propylphenyl, etc.); or a fluorocarbon group, preferably having from 1 to 30 carbon atoms (e.g. perfluorohexyl, perfluorododecyl, etc.); provided that Al, R2, R3 and R4 together comprise at least 12 carbon atoms.
Alternatively, I and R2 taken together with the nitrogen atom, can form a qua ternary heterocyclic ring, e.g. pyrrolidinium, piperidinium and the like, to form a heterocyclic amine having two pendant groups, R3 and R4, attached to the qua ternary ammonium atom. This heterocycllc ring generally contains a total of 5 to 20 carbon atoms and heteroatoms (as defined hereinabove) in the rung backbone. The ring can be substituted with any of a number of moieties known to one skilled in the art. In this heterocyclic compound, R3 and R4 are as defined hereinabove but comprise at least 12 carbon atoms.
In another embodiment illustrated by structure IBM Al, R2 and R3, taken together with the nitrogen atom, can form a qua ternary heterocyclic ring 9 e.g. pyridinium, quinolinium, pyrimidinium and the like to provide a heterocyclic amine having a single pendant group ~2~35~

R4 . R4 it a defined above ~190 but compare at least 12 carbon atomic This heterocyclic amine generally contains 5 to 20 carbon atom and heteroatom~ (fig defined hereinabove) in the ring backbone (e.g. pyridinium, quinolinium, acridinium, ben20thi~zolium, benzoxazolium, etc.). Preferably, R4 it alkyd (as defined above) of 12 to 30 carbon atom, X to any suitable monovalent anion, such a halide, nitrate, phosphate, sulfate and the live.
In a more preferred embodiment, Al it substituted or un~ub~tituted alkyd of 1 to 6 carbon atoms; R2 it substituted or unsubstituted alkyd of 1 to 6 carbon atom or lo (alkylene-oxy OH
wherein ~lkylene has 2 to 4 carbon atoms end it cub3tituted or unsubstituted; and % it halide or nitrite.
The following lit of useful qusternary ammonium compounds is not exhaustive of compound useful in the practice ox this invention, but it provide A representative sampling of useful compound. Useful compounds include:
Compound Generic or trade name + - bromide, Of Dodecyltrimethyl ammonium CH3(CH~ N(C~3)3 chloride;
+ Bra Hexadecyltrimethyl CH3(CH2)1sN( 3)3 ammonium bromide;
CH2~1sCH3 By ex~decylpyridlnium -Chinook Of chloride;

H B Didodecyldimethyl ~CH3(CH~ MCKEE 3)2 ammonlum bromide;

~35~

I I -CHINCH) 2 Of Benzyldimethylphenyl ammonium chloride;
Jo \.
I!
Jo/

[CH3(CH2)s]4N Of- Tetrahexyl ammonium chloride;
+

SHEA (Shelley 7N(CH3) 2 Of- Stearyldimethylbenzyl 1 ammonium chloride;
ITCH

I!

+
H[OCH(CH3)CH2]n~CH(CH3)CH2N(C2Hs)2 Of-OH Polypropoxy qua ternary 3 ammonium chlorides wherein n is 8, 25 or 41;
Cyastat Snowily (American O C+3 Cyanamid, Wayne, New Jersey);
CH3(CH2)l6CNH(CH2)3NCH2CH20H NO-SHEA
Flurried (EM Company, Cl~Fl7SO2NHCH2CH2N(CH3)3 I- St. Paul, Minnesota);
+

~R')2N(CH3) 2 Of- Dicocodimethyl ammonium chloride (Armak Industrial Chemicals, Chicago, Illinois) which is a mixture of compounds wherein R' is alkyd of 12, 14 or 16 carbon atoms; and (CH2CH20~H
R" - N ;- SHEA Of- Polyethoxyl~ted qua ternary l ammonium chlorides.
(CH2CH20)yH

:~235~

wherein R" is alkyd of 8 to 18 carbon atoms, x and y are independently integers of 2 to 50.
The elements of this invention can include an interactive composition although the presence of such a composition is not required in the element for practice of this invention (e.g. it could be added in the liquid sample or as a separate sample). These compositions can be a single chemical compound or a combination of chemical compounds or reagents which can interact with the analyze, with a reaction decomposition product of the analyze, or with each other, upon contact with the analyze to produce a detectable change. Such interaction is meant to refer to chemical reactivity, catalytic activity as in the formation of an enzyme-substrate complex, antigen-antibody reaction and any other form of chemical or physical interaction that can produce or promote within the element, such as in a reagent or spreading zone, a change which is radio metrically detectable, that is by suitable measurement of light or any other energy form. For example, the change can be release of a preformed detectable species, the formation of a detectable species or the disappearance or reduction in amount of a detectable I species. The change produced can be correlated to the mount of analyze in the liquid sample.
The particular interactive composition distributed within the element will depend on the analysis of choice. The composition useful for a riven analysis would be within the skill of a worker in the clinical chemistry art. The elements of this invention can be adapted, for example, for the analysis of ground water, foodstuffs, and biological fluids, such as blood, plasma 9 serum, cerebral spinal fluid, urine and the like. Analyses of analyzes, such as glucose, lactate, triglycerides, tool protein, albumin, uric acid; bilirubin, creative Cannes, aimless, alkaline phosphates, lactate dehydrogenase, ala nine aminotransferase (AT) and aspartame aminotransferase (AT) are but representative of the potential uses of the elements of this invention.
In one embodiment of this invention, the element is used to determine aspartame aminotransferase (AT) or ala nine aminotransferase (AT). The interactive composition for AT elements can include, for lo example, in quantities known by one skilled in the oft, L-aspar~ate, ~-ketoglutarate, NASH
(nicotinamide adenine dinucleotide), lactate dehydrogenase, palate dehydrogenase and pyridoxal phosphate. Alternatively, other interactive compositions known for determining AT can be used.
n interactive composition useful in AT
elements includes, for example, in quantities known by one skilled in the art, L-alanine, NASH, ~-ketoglutarate, lactate dehydrogenase and pyridoxal phosphate Alternatively, other interactive compositions known for determining AT
can be used, if desired.
The interactive composition can be placed in any suitable location in the elements of this invention. Where the element consists of a single porous spreading zone the composition is in that zone in its entirety. However, where the element consists of multiple Jones, the composition can be in any of the zones Swig. a reagent zone), or the Jo components of the composition can be distributed among two or more zones in the element.
The dry analytical elements of this invention have at least one porous spreading zone, i.e. the blush polymer zone described hereinabove.
This zone can also be a spreading/reagent zone if it contains the interactive composition or Moe Component thereof. This zone can be a ~.Z~3~

elf-supp~rting carrier matrix, but preferably it it carried on a separate support. Such a support it a film or sheet made of any suitable dimen~lon~lly table, and preferably transparent (lye. rAdiQtlon tran~mi~ive) materiel which transmit electromagnetic radiation of a wavelength between about 200 and about 900 no. Useful support materiels include polystyrene, poultry [e.g. polyethylene terephthalate)], polyc&rbonates, cellulose esters, lo etc.
The element can have a inlay zone or a plurality of zones (including spreading, reagent, subbing, buffer, reflective, barrier, etc. zones), Rome or ~11 of which can cont~ln reagent. These Jones are in fluid contact with each other, meaning that fluids con pow between superposed region of adjacent zones. Stated in another manner, fluid contact refer to the ability to import component of a fluid between the zone in fluid contact.
Preferably, the zones are separate costed layers, although one or more zones can be in single layer, or one so more separate layers can be in a jingle zone, of an element. Dry element format end materiel sure known in the art and described for example in U.S. Patents 3,992,159 (noted herein-above; 4,042,335 (idea August 16, 1977 to Clement), 4,144,306 (issued March 13, 1979 to Pharaoh); 4,132~528 (idea January 2, 1979 to ken berry et at); 4,~58,001 (idea March 24, 1981 to Pierce et 81~; 4,292,272 (issued September 29, 1981 to Kita~ma et at); 4,430,436 (issued February 7, 1984 to Comma et Al); Japanese Potent Publication 57(1982)-101760; and U.S. Patent

4,450,232 (i sued May 22, 1984 to Sanford et I

Particularly useful elements for the determination of the transferases, AT and ALTO aye illustrated in the examples hereinbelow.
The analytical method of this invention can be automated or manual. In general, an analyze in an aqueous liquid is determined by taking an element from a supply roll, slide packet or other source and physically contacting it with a sample of the liquid. Such contact can be accomplished in any suitable manner, e.g. dipping or immersing the element into the sample or, preferably, by spotting the element by hand or machine with a drop (e.g.
about 1-20 AL) of the sample by pipette or another suitable dispensing means After sample application, the element is exposed to any conditioning, such as incubation, heating or the like, that may be desirable to quicken or otherwise facilitate obtaining any test result.
The interactive composition will chemically react with any analyze present in the sample and produce a detectable change (as described hereinabove which can be measured at an end-point or as a rate change with suitable detection equipment and techniques.
Such equipment includes conventional reflection, transmission or fluorescence spectrophotometers which are well known in the art.
he following examples illustrate the practice of this invention. In preparing the dry analytical elements, the components were obtained from the following sources: polyurethane resin ~Estane'n 5715) from B. F. Goodrich (Cleveland, Ohio); Triton'n ~05 surfactant from Room & Hays (Philadelphia, Pennsylvania); Brian 98 surfactan~
f~Qm ICY Americas, Inc. (Wilmington, Delaware);
sodium ~-ketoglutarate from Sigma Chemicals (St. Louis Missouri); sodium L-aspartate from ION
Nutritional Biochemical Cleveland, Ohio); lactate dehydrogenase, palate dehydrogenase and pyridoxal-5-phosphate from Boehringer Minim (Indianapolis Indiana); NASH from P-L Biochemical (Milwaukee, Wisconsin); polypropoxy qua ternary ammonium chlorides as EMCOL~ CC36 and CC42 from Witch Chemical Corp. (New York, Jew York);
amylopectin starch from National Starch Chemical Corp. (Buffalo, New York); Drimarine Red Z2B dye from Sundays Corp. (Hanover, New Jersey), and the remainder from Eastman Organic Chemicals (Rochester, New York or from inlays sources.
The cohesive strength test used in the examples measures the distance, in millimeters, at which a sapphire stylus scribing the surface of the blush polymer spreading layer first produces a "flaking" of the layer so that the layer begins crumbling.
The test was curried out in the following manner: dried blush polymer spreading layers were conditioned at about 20C and So% relative humidity for 1 hour. A 12 x 18 cm sample of the layer was then positioned under the stylus and the stylus was lowered to the surface of the spreading layer. The sample was mechanically moved under the lowered stylus for about 158 mm. The stylus arm was then raised and cleaned. This scribing procedure was repeated 5 times for each sample, each time at a different location on the sample. The distance of scribing at which "flaking" first occurred was then measured for each scribing and the six measurements for each sample were averaged. "Flaking" refers to the breaking apart of the spreading layer and formation of slakes or pieces thereof.
I

Examples 1 3 Elements With Spreading Layers Having Improved Cohesion Elements containing a blush polymer spreading layer were prepared on a polyethylene terephthalate) support. The spreading layer comprised the components noted in Table I
hereinbelow. Control elements outside the scope of this invention were also prepared. In each case, the element prepared according to the present invention having a blush polymer spreading layer composed of a qua ternary ammonium compound described herein exhibited significant improvement in cohesive strength over the Control element.

- -18~ I

o , Jo , e o Us o co P e Oily .,1 t:: o F, Jo a; o o O rl o e . . I u Al H 0 0 I
a Jo ,~: x a O Jo V rl H ad 01 I

O
pa o o us C`J O Us o zoo owe zoo Jo ,1 3 o G
o ~,~ I a) Al rl ¢
I J J C) I 0 Owe O J- O I 5 o I r X us rl O Eel o o us o Jo O El F O
or C O C
O 1` or I 1 0 X OJC~-rl I a) Us X t-) c) X 1 h a Jo JO Jo I
I in ox c to to to to rJ
o to I o to I to to o Err 5 o o lid e O
I; I or I O'er r-l to O a) 3 X h O Ed or a to us h to or A r-l a or I I O I
O, to Ed X En O
Jo C to a) Lull O Jo 01~ ¢ I
C Eye O 1-1 0 Sol 0 I
Jo Go a F I h Cud Jo , o , 04 Jo rJ
S I O X o a o Jo I

Example 4 Element for Determination of Aspartame Aminotransferase (AT) This is an example comparing an element useful for determining AT prepared according to this invention to a conventional element. The element of this invention had the format and components noted hereinbelow. The Control element was similarly made except N-hexadecylpyridinium bromide was omitted from the spreading/reagent layer.

Barium sulfite g/m2 Cellulose astute g/m2 Briton X-405 surfactant0.6-6 g/m2 Spreading/ Estane'A 5715 polyurethane 0.5-5 g/m2 Reagent Sodium ~-ketoglutarate0.1-1 g/m2 Lowry Sodium aspartate1.5-15 g/m2 N-hexadecylpyridinium bromide 0.3-3 g/m2 _ Subbing Polyvinyl pyrrolidone) 0.15-1.5 g/m2 lever Gelatin (hardened) 3-30 g/m Briton X-405 surfactant 0.15-1.5 g/m2 Reagent Tris(hydroxymethyl)amino Layer methane 1.5-15 g/m2 Lactate dehydrogenase300-3000 U/m2 Palate dehydrogenase300-3000 U/m2 NASH 0.1-1 g/m2 Pyridoxal-5 phosphate g/m2 _ / Polyethylene terephthalate) /// / / /
I/ Support / Jo / /

These elements were evaluated for spreading/rea~en~ layer cohesive strength by the procedure described hereinabove. The results presented in Table II hereinbelow, show that the element of this invention exhibited substantial improvement in cohesive strength of the spreadingtreagent layer over the Control element.

Table II
Cohesive Strength Element (mm to flake) Improvement Control 32 --Example 4 60 88%

Both the Control element and the element of this invention were used to determine AT in serum samples, The incorporation of the qua ternary ammonium compound in the spreading/reagent layer of the element of this invention did not adversely affect the determination of AT.
Example 5 Element fur Determination of Ala nine Aminotranserase (AT) This is a comparative example like Example 4. An element for determining AT was prepared having the format and components described for the AT element except that L-alanine was substituted for sodium aspartame in the spreading/reagent layer, and palate dehydrogenase was omitted from the reagent layer. A Control element was similarly prepared with the N-hexadecylpyridinium bromide omitted from the spreading/reagent layer. The results, presented in Table III hereinbelow, show that the element of this invention exhibited substantial improvement in cohesive strength of the spreading/reagent layer over the Control element.
Table III
_ Cohesive Strength Element(mm to flake % Improvement Control 57 --Example 5 1~0 75%

35~

Both the Control element and the element of this invention were used to determine AT in serum samples. The incorporation of the qua ternary ammonium compound in the spreading/reagent layer of the element of this invention did not adversely affect the determination of ALTO
Example 6 Element for Determination of Serum Aimless This is an example comparing an element of this invention useful for determination of aimless in a serum sample to a Control element prepared without a qua ternary ammonium compound in the spreading/reagent layer. The element of this invention had the format and components shown hereinbelow. The Control element was similarly prepared but the EMCOL~-CC36 compound was omitted from the spreading/reagent layer.

Titanium dioxide 20-200 g/m2 Cellulose acetate 3-30 g/m Bridge surfactant 0.2-2 g/m2 Briton X-405 surfactant 0,4-4 g/m2 Spreading/ Estate 5715 polyurethane 0.5-5 g/m2 Reagent KH2PO~ buffer 0.6-6 g/m2 Layer Nope buffer 0.4_4 g/m2 Dyed amylopectin* 1-10 g/m2 EMCOL~-CC36 0.3_3 g/m2 Subbing Polyvinyl pyrrolidone) 0.15-1.5 g/m2 Layer Gelatin (hardened) 0.5 10 g/m2 Poly(styrene-co-N-vinyl-benzyl-N-benzyl-N,N-Reagent dimethylammonium chloride-Layer co-divinyl Bunsen g/m2 Nope (pi 6~8) buffer 0.05-1 g/m2 Briton X 405 surfactant 0.05_0.3 g/m2 .

/ / / Polyethylene terephthalate) / // / /
//// Support / / / / / /

~2~35~(~

* This is a dye starch complex prepared from amylopectin and Drimarine Red Z2B.
These elements were evaluated for cohesive strength of the blush polymer spreading/reagent layer according to the procedure described hereinabove.
The results of the tests, shown in Table IV
hereinbelow, indicate that the element of this invention had substantially improved cohesive strength over the Control element.
Table IV
Cohesion Element (mm to flake) % Improvement Control 9 Example 6 27 300%

The invention has been described in detail with particular reference to preferred embodiments thereof, but it will be understood that variations and modifications can be effected within the spirit and scope of the invention.

I

Claims (20)

I claim:

1. A dry element for the analysis or transport of an aqueous liquid, said element comprising an isotropically porous zone which comprises a blush polymer having dispersed therein a particulate material, and a nonpolymeric heterocyclic, aliphatic or carbocyclic quaternary ammonium compound having at least 12 carbon atoms therein, which compound is present in said zone in an amount of at least about 2 weight percent, based on the dry weight of said blush polymer.

2. The element of claim 1 wherein the weight ratio of said particulate material to said blush polymer is from about 2:1 to about 20:1.

3. The element of claim 1 wherein said quaternary ammonium compound is represented by the structures (Ia and b):
or wherein ? represents a single or double bond;
R1, R2, R3 and R4 are independently alkyl of 1 to 30 carbon atoms, ?alkylene-oxy?n-H
wherein said alkylene comprises 2 to 6 carbon atoms and n is an integer of 1 to 50; aryl of 6 to 14 carbon atoms, cycloalkyl of 5 to 20 carbon atoms, alkaryl of 7 to 30 carbon atoms, or a fluorocarbon group of 1 to 30 carbon atoms; provided that R1, R2, R3 and R4 together comprise at least 12 carbon atoms; or R1 and R2, taken together with the nitrogen atom, form a quaternary heterocyclic ring and R3 and R4 are as defined above and comprise together at least 12 carbon atoms; or R1, R2 and R3, taken together with the nitrogen atom, form a quaternary heterocyclic ring and R4 is as defined above and comprises at least 12 carbon atoms; and X- is a monovalent anion.

4. The element of claim 3 wherein R1 is alkyl of 1 to 6 carbon atoms, R2 is alkyl of 1 to 6 carbon atoms or ?alkylene-oxy?n-H wherein said alkylene comprises 2 to 4 carbon atoms; and X- is halide or nitrate.

5. A dry analytical element for determination of an analyte in an aqueous liquid, said element comprising:
an interactive composition for said analyte;
and a support having thereon an isotropically porous zone which comprises a blush polymer having dispersed therein a particulate material, and a nonpolymaric heterocyclic, aliphatic or carbocyclic quaternary ammonium compound having at least 12 carbon atoms therein, which compound is present in said zone in an amount of at least about 2 weight percent, based on the dry weight of said blush polymer.

6. The element of claim 5 wherein said quaternary ammonium compound is represented by the structures (Ia and b):

or wherein ? represents a single or double bond;
R1, R2, R3 and R4 are independently alkyl of 1 to 30 carbon atoms, ?alkylene-oxy?n-H
wherein said alkylene comprises 2 to 6 carbon atoms and n is an integer of 1 to 50; aryl of 6 to 14 carbon atoms, cycloalkyl of 5 to 20 carbon atoms, alkaryl of 7 to 30 carbon atoms, or a fluorocarbon group of 1 to 30 carbon atoms; provided that R1, R2, R3 and R4 together comprise at least 12 carbon atoms; or R1 and R2, taken together with the nitrogen atom, form a quaternary heterocyclic ring and R3 and R4 are as defined above and comprise together at least 12 carbon atoms; or R1, R2 and R3, taken together with the nitrogen atom, form a quaternary heterocyclic ring and R4 is as defined above and comprises at least 12 carbon atoms; and X- is a monovalent anion.

7. The element of claim 6 wherein R1 is alkyl of 1 to 6 carbon atoms, R2 is alkyl of 1 to 6 carbon atoms or ?alkylene-oxy?n-H wherein said alkylene comprises 2 to 4 carbon atoms; and X- is halide or nitrate.

8. The element of claim 6 wherein said quaternary ammonium compound is a heterocyclic amine represented by the structure (Ib) wherein R1, R2 and R3, taken together with the nitrogen atom, complete a 5 to 20-membered quaternary heterocyclic ring.

9. The element of claim 6 wherein said quaternary ammonium compound is selected from the group consisting of:
CH3(CH2)8?(CH3)3 Br-; CH3(CH2)11?(CH3)3 Cl-;

CH3(CH2)15?(CH3)3 Br-;

?CH2?15CH3 Br-; Cl-;

[CH3(CH2)11]2?(CH3)2 Br-; Cl-;

[CH3(CH2)5]4? Cl-; ;

H[OCH(CH3)CH2]nOCH(CH3)CH2(C2H5)2 Cl-wherein n is 8, 25 or 41;
;

C8F17SO2NHCH2CH2?(CH3)3 I-; (R')2?(CH3)2 Cl-wherein R' is alkyl of 12, 14 or 16 carbon atoms; and Cl-wherein R" is alkyl of 8 to 18 carbon atoms, x and y are independently integers of 2 to 50.

10. The element of claim 9 wherein said quaternary ammonium compound is ?CH2?15CH3 Br-, or H[OCH(CH3)CH2]nOCH(CH3)CH2(C2H5)2 Cl-wherein n is 8 or 25.

11. The element of claim 5 wherein said quaternary ammonium compound is present in said porous zone in an amount of from about 8 to about 30 weight percent, based on the dry weight of said blush polymer.

12. A dry multizone analytical element for determination of an analyte in an aqueous liquid, said element comprising an interactive composition for said analyte, and a support having thereon, in order and in fluid contact, a reagent zone containing at least one component of said composition, and an isotropically porous spreading zone which comprises a blush polymer having dispersed therein a particulate material, and a nonpolymeric heterocyclic, aliphatic or carbocyclic quaternary ammonium compound which is present in said zone in an amount of at least about 2 weight percent, based on the dry weight of said blush polymer;
wherein said quaternary ammonium compound is represented by the structures (Ia and b):

or wherein ? represents a single or double bond;
R1, R2, R3 and R4 are independently alkyl of 1 to 30 carbon atoms, ?alkylene-oxy?n-H
wherein said alkylene comprises 2 to 6 carbon atoms and n is an integer of 1 to 50; aryl of 6 to 14 carbon atoms, cycloalkyl of 5 to 20 carbon atoms, alkaryl of 7 to 30 carbon atoms, or a fluorocarbon group of 1 to 30 carbon atoms; provided that R1, R2, R3 and R4 together comprise at least 12 carbon atoms; or R1 and R2, taken together with the nitrogen atom, form a quaternary heterocyclic ring and R3 and R4 are as defined above and comprise together at least 12 carbon atoms; or R1, R2 and R3, taken together with the nitrogen atom, form a quaternary heterocyclic ring and R4 is as defined above and comprises at least 12 carbon atoms; and X- is a monovalent anion.

13. The element of claim 12 wherein said particulate material is titanium dioxide or barium sulfate.

14. The element of claim 13 wherein said spreading layer comprises cellulose acetate.

15. The element of claim 12 wherein said analyte is aspartate aminotransferase and said interactive composition comprises NADH, L-aspartate, .alpha.-ketoglutarate, lactate dehydrogenase, malate dehydrogenase and pyridoxal phosphate, or said analyte is alanine aminotransferase and said interactive composition comprises NADH, L-alanine, .alpha.-ketoglutarate, lactate dehydrogenase and pyridoxal phosphate.

16. A method for the determination of an analyte in an aqueous liquid, said method comprising the steps of:
A. physically contacting a sample of said liquid together with an interactive composition for said analyte and a dry analytical element to provide a detectable change, said element comprising an isotropically porous zone which comprises a blush polymer having dispersed therein a particulate material, and a nonpolymeric heterocyclic, aliphatic or carbocyclic quaternary ammonium compound having at least 12 carbon atoms therein, which compound is present in said zone in an amount of at least about 2 weight percent, based on the dry weight of said blush polymer; and B. measuring said detectable change.

17. The method of claim 16 wherein said quaternary ammonium compound is represented by the structures (Ia and b):

X- or X-wherein ? represents a single or double bond;
R1, R2, R3 and R4 are independently alkyl of 1 to 30 carbon atoms, ?alkylene-oxy?n-H

wherein said alkylene comprises 2 to 6 carbon atoms and n is an integer of 1 to 50; aryl of 6 to 14 carbon atoms, cycloalkyl of 5 to 20 carbon atoms, alkaryl of 7 to 30 carbon atoms, or a fluorocarbon group of 1 to 30 carbon atoms; provided that R1, R2, R3 and R4 together comprise at least 12 carbon atoms; or R1 and R2, taken together with the nitrogen atom, form a quaternary heterocyclic ring and R3 and R4 are as defined above and comprise together at least 12 carbon atoms; or R1, R2 and R3, taken together with the nitrogen atom, form a quaternary heterocyclic ring and R4 is as defined above and comprises at least 12 carbon atoms; and X- is a monovalent anion.

18. The method of claim 17 wherein said detectable change is in the rate of formation of a detectable species.

19. The method of claim 17 wherein said detectable change is in the rate of disappearance of a detectable species.

20. The method of claim 17 wherein said interactive composition is in said element.