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PubMed1989

An evaluation of two configurations of tricalcium phosphate for treating craniotomies.

Biodegradable beta-tricalcium phosphate disks (TCP) of two configurations were inserted into 15-mm-diameter craniotomy wounds and nontreated control sites were evaluated in 60 rabbits. There were no adverse tissue reactions and no apparent difference in the clinical appearance of the 12- and 24-week implanted disks. By 36 weeks and continuing to 48 weeks, the omnidirectional TCP (OTCP) implants were degrading more rapidly than the unidirectional TCP (UTCP) implants, with degradation progressing centripetally and replacement by woven bone and maturing lamellar bone. Host-implant interface of both TCP configurations was a bone bond without interposed soft tissue. TCP disks may be clinically useful for craniotomy repair.

Animals↗
PubMed1990

Marrow cell induced osteogenesis in porous hydroxyapatite and tricalcium phosphate: a comparative histomorphometric study of ectopic bone formation.

To investigate the bone formation ability of porous hydroxyapatite (HA) and tricalcium phosphate (TCP), ceramic discs were implanted with or without rat marrow cells into subcutaneous sites in syngeneic rats. The discs of HA and TCP had identical microstructures: pore size was 190-230 microns, porosity was 50-60%, and they were fully interconnected. Implants without marrow cells (discs themselves) did not show bone formation, whereas implants with marrow cells showed bone formation in the pores of the ceramics. The bone formation of both HA and TCP occurred initially on the surface of the ceramic and progressed towards the center of the pore. The de novo bone was quantitated from decalcified serial sections of the implants. One month after implantation with marrow cells, the percentage fractions of the pore area filled with bone for implanted HA and TCP were 16.9 and 15.1, respectively. At 2 months after implantation with marrow cells, the fractions of bone were 34.3 and 30.9, respectively. These results indicate that both HA and TCP ceramics can show comparable osteogenic ability in the presence of marrow cells.

Animals↗
PubMed1994

Inflammatory response and degradation of three types of calcium phosphate ceramic in a non-osseous environment.

The cellular response and inflammatory reaction to three types of calcium phosphate ceramic spheres, namely fluorapatite (FA), hydroxyapatite (HA), and beta-tricalcium phosphate (TCP), were investigated. The degradation at ultrastructural level of these ceramics was also studied. The mouse peritoneal cavity was used as a model compartment for this study. To minimize the influence of a wound reaction, the spheres were implanted by injection. Intraperitoneal injection of latex beads of 6.4 and 25.7 microns showed that an inflammatory reaction is influenced by the size of injected particles; therefore, ceramic spheres with the same average diameter (11.3 microns) were used. After 2 and 4 days and 1, 2, 4, 8, 12, and 24 weeks, the spheres were harvested. All materials gave rise to a short, mild inflammatory reaction. The ceramic spheres aggregated rapidly, and foreign body granulomas were formed within 2 weeks. Degradation of all materials was observed in this study, and degradation products were frequently seen both intracellularly and extracellularly. After 2 weeks TCP and HA showed the formation of crystals at their surface and between individual grains of the spheres. This was most prominently seen with TCP. At 4 weeks, HA and TCP granulomas showed the formation of extracellular, globular deposits between macrophages or multinucleated giant cells and fragments of the ceramic. They consisted of an organic matrix containing apatite-like crystals and iron, and had an average diameter of 10 microns. The number of deposits was the highest for TCP. For FA, calcium phosphate precipitation and the formation of deposits was not observed before the 8-week interval and remained at a low level.

Animals↗
PubMed2004

A new bone-inducing biodegradable porous beta-tricalcium phosphate.

A new type of degradable biomaterial with bone-inducing capacity was made by combining porous beta-tricalcium phosphate (beta-TCP) with a delivery system for recombinant human bone morphogenetic protein-2 (rhBMP-2). The BMP delivery system consisted of a block copolymer composed of poly-D,L-lactic acid with random insertion of p-dioxanone and polyethylene glycol (PLA-DX-PEG), a known biocompatible and biodegradable material. The efficacy of this biomaterial in terms of its bone-inducing capacity was examined by ectopic bone formation in the dorsal muscles of the mouse. In the beta-TCP implants coated with the PLA-DX-PEG polymer containing more than 0.0025% (w/w) of rhBMP-2, new ectopic bone tissues with marrow were consistently found on the surface of implants. The radiographic density of beta-TCP was diminished in a time-dependent manner. On histological examination, numerous multinucleated osteoclasts with positive tartrate-resistant acid-phosphatase (TRAP) staining were noted on the surface of the beta-TCP. These experimental results indicate that beta-TCP implants coated with synthetic rhBMP-2 delivery system might provide effective artificial bone-graft substitutes with osteoinductive capacity and biodegradable properties. In addition, this type of biomaterial may require less rhBMP-2 to induce significant new bone mass.

Absorbable Implants↗
PubMed2005

In vitro evaluation of biocompatibility of beta-tricalcium phosphate-reinforced high-density polyethylene; an orthopedic composite.

Beta-tricalcium phosphate-reinforced high-density polyethylene (beta-TCP/HDPE) is a new biomaterial which was made as a copy of bone composition with the aim of replacement of bony tissues. The composite samples were prepared using medical grade TCP powder and granular polyethylene. The raw materials were first compounded and the resulting composite preforms were compression molded into desired shape. The biocompatibility of composite samples with different volume fractions of TCP (20, 30, and 40 vol %) was assessed by proliferation, alkaline phosphatase (ALP), and cell adhesion assays using G-292 osteoblast cells. Cell-material interaction on the surface of the composites was observed by scanning electron microscopy (SEM). The effect of beta-TCP/HDPE on the behavior of G-292 cells was compared with those of a composite and a negative control samples. Results showed the composite samples had a higher proliferation rate of G-292 cells in the presence of composite samples as compared to the composite control sample after 3, 7, and 14 days of incubation period. ALP production after incubation in the presence of composite samples was seen to peak on the day 7. The number of adhered cells on the composite samples was higher than the numbers adhered on composite and negative control samples after the above incubation periods. Morphology investigation of adhered cells by SEM indicated a normal morphology and also many of the cells were in the process of cell division. The above results indicate that beta-TCP/HDPE samples are biocompatible, nontoxic, and in some cases show an increase in the proliferation rate of the cells, ALP production, and cell adhesion as compared to the control counterparts.

Alkaline Phosphatase↗
PubMed2005

Comparison of hydroxyapatite and beta tricalcium phosphate as bone substitutes after excision of bone tumors.

Long-term results are reported in 23 patients and short-term results in 30 patients presenting with bone tumors treated by curettage or resection followed by implantation of hydroxyapatite (HA) or highly purified beta-tricalcium phosphate (beta-TCP), respectively. Mean follow-up was 97 and 26 months in cases involving HA implantation and beta-TCP implantation, respectively. Radiographs revealed HA incorporation into host bone in all but two cases; moreover, no obvious evidence of HA biodegradation was observed. A single patient exhibited late deformity following implantation of HA. All grafted beta-TCP was, at least partially, absorbed and replaced by newly formed bone. The mean period required for the disappearance of radiolucent zones between the ceramics and host bone was 17 weeks in HA and 9.7 weeks in beta-TCP. Highly purified beta-TCP appears to be advantageous relative to HA for surgical intervention in bone tumors consequent to the nature of remodeling and superior osteoconductivity.

Adolescent↗
PubMed2007

Comparison of osteoblast-like cell responses to calcium silicate and tricalcium phosphate ceramics in vitro.

Calcium silicate ceramics have been proposed as new bone repair biomaterials, since they have proved to be bioactive, degradable, and biocompatible. Beta-tricalcium phosphate ceramic is a well-known degradable material for bone repair. This study compared the effects of CaSiO3 (alpha-, and beta-CaSiO3) and beta-Ca3(PO4)2 (beta-TCP) ceramics on the early stages of rat osteoblast-like cell attachment, proliferation, and differentiation. Osteoblast-like cells were cultured directly on CaSiO3 (alpha-, and beta-CaSiO3) and beta-TCP ceramics. Attachment of a greater number of cells was observed on CaSiO3 (alpha-, and beta-CaSiO3) ceramics compared with beta-TCP ceramics after incubation for 6 h. SEM observations showed an intimate contact between cells and the substrates, significant cells adhesion, and that the cells spread and grew on the surfaces of all the materials. In addition, the proliferation rate and alkaline phosphatase (ALP) activity of the cells on the CaSiO3 (alpha-, and beta-CaSiO3) ceramics were improved when compared with the beta-TCP ceramics. In the presence of CaSiO3, elevated levels of calcium and silicon in the culture medium were observed throughout the 7-day culture period. In conclusion, the results of the present study revealed that CaSiO3 ceramics showed greater ability to support cell attachment, proliferation, and differentiation than beta-TCP ceramic.

Alkaline Phosphatase↗
PubMed1985

Comparative study of porous hydroxyapatite and tricalcium phosphate as bone substitute.

This study was performed to quantitate the bone ingrowth and biodegradability of porous calcium phosphate. Two kinds of coralline hydroxyapatite (CHA), along with sintered tricalcium phosphate (TCP), were evaluated. All implants had totally interconnecting pores. These pores measured 260-600 mu for CHA-Goniopora (CHAG), 190-230 microns for CHA-Porites (CHAP), and 100-300 microns for TCP. Cylindrical implants (3 mm in diameter by 8 mm in length) were implanted into the diaphyses of rabbit tibias for 3 to 24 weeks. The quantity of regenerated bone, remaining implant, nonmineralized space, and the middle two quarters of the cortical area were measured by a computerized operator-assisted analysis for bone histomorphometry. At 3 weeks after implantation, 17.3% of CHAG, 11.3% of CHAP, and 7.0% of TCP were infiltrated by regenerated bone. At 24 weeks, 56.1% of CHAG, 52.7% of CHAP, and 44.7% of TCP were occupied by lamellar-type bone. Implant degradation was noted to be 46.4% for TCP and 27.5% for CHAP. In contrast, CHAG did not show appreciable degradation until 24 weeks.

Animals↗
PubMed2006

Sorption of chlorophenoxy propionic acids by organoclay complexes.

The organoclays Wyoming montmorillonite (SWy-1, Mon), W. R. Grace vermiculite (Ver), and Tung-Wei (Tw) soil montmorillonite were prepared to sorb or remove chlorophenoxy propionic acids (CPA) pollutants such as 2,4-dichlorophenoxy acid (2,4-DP), 2,4,6-tri chlorophenoxy acid (2,4,6-TCP), and pentachlorophenoxy propionic acid (PCP). The objective of this study was to evaluate the behavior of organoclays that have sorbed three types of CPA and to assess the applicability to basaltic-developed iron-rich calcareous soil. This type of soil contains high amounts of montmorillonite. The mean-layer charge per formula unit of Mon, Ver, and Tw clays, characterized by the alkylammonium method, was 0.27, 0.70, and 0.52 mol(c)/O(10)(OH)(2), respectively. The sorption isotherms of 2,4-DP, 2,4,6-TCP, and PCP sorbed by hexadecyltrimethyl-ammonium (HDTMA organoclay, 150% cation-exchange capacity) were evaluated. The data on sorption of CPAs fit well with the Freundlich isotherm equation. In general, the sorption of three types of CPA by the organoclays showed a linear relationship, with a high correlation coefficient (r > 0.935). The K(f) value of PCP was the same as that of 2,4,6-TCP sorbed by the organoclays and was higher than that of 2,4-DP. However, the 1/n(f) values of the three CPAs sorbed by the organoclays did not show significant differences. In general, maximum CPA sorption occurred at a pH of approximately 3, but very significant differences were observed for the 2,4,6-TCP and PCP sorbed by the organoclays. However, there was no significant difference in pH for 2,4-DP sorption. This should be related to the solubility and pKa of each CPA. The experimental results showed that solubility of 2,4-DP, 2,4,6-TCP, and PCP is a function of pH, ionic strength, and temperature.

Adsorption↗
PubMed1994

Reproductive toxicity of butylated triphenyl phosphate and tricresyl phosphate fluids in F344 rats.

The effects of tricresyl phosphate (TCP) and butylated triphenyl phosphate (BTP)-based hydraulic fluid on reproduction were studied in F344 rats using a modification of the National Toxicology Program's Continuous Breeding Protocol. Groups of breeding pairs received single daily oral doses of an equal volume of either 0, 0.6, 1.0 g BTP/kg or 0.4 TCP/kg in sesame oil or 1.7 g neat BTP/kg for up to 135 days. A naive control group allowed to breed, but not dosed or handled daily, demonstrated that daily dosing and handling of the rats had no effect on reproduction. The fertility index and number of litters born were significantly decreased in rats exposed to 1.0 and 1.7 g BTP/kg and 0.4 g TCP/kg. The number of pups per litter was significantly decreased in the TCP group. A crossover mating experiment using 0.4 g TCP/kg/day and 1.0 g BTP/kg/day groups, each mated with vehicle controls, demonstrated that TCP caused 100% infertility in male rats but did not affect reproduction in females. BTP caused a significant decline in reproduction in female rats characterized by low mating and fertility indices, decreased number of litters, and abnormal estrous cycles. Fertility was decreased in the BTP-dosed male rats. Both sexes of rats in the crossover experiment with TCP and BTP had significant decreases in terminal body weights and increases in adrenal gland and liver weights. Only TCP-dosed male rats had significantly decreased testicular and epididymal weights.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1997

Kinetics of Dissolution of beta-Tricalcium Phosphate

The rate of dissolution of beta-tricalcium phosphate (beta-Ca3 (PO4 )2 ; beta-TCP) has been measured in the solution system Ca(OH)2 -H3 PO4 -NaOH-HNO3 -H2 O. The effects of different parameters such as pH, temperature, time, and saturation have been investigated. At zero saturation, the logarithm of the dissolution rate is a linear function of the pH (log(j 0 ) = 2.02 - 0.82 pH; r 2 = 0.993; in mmol/m2 s), indicating diffusion-limited dissolution. A simple calculation of the theoretical rate of a diffusion-controlled process showed that our data are consistent with theory. Moreover, the activation energy for this process is low (E act = 3.9 kcal/mol) also suggesting that the beta-TCP dissolution is controlled by diffusion processes. At increased saturation, the initial beta-TCP dissolution rate decreases much faster than that predicted assuming a diffusion-controlled model. However, this latter model gives a good prediction of the results if it is assumed that beta-TCP dissolution is controlled by the dissolution of an interfacial layer of hydroxyapatite (Ca5 (PO4 )3 OH; HAp): log(j ) = -1.47 + 1.34 log(1 - S HAp ); r 2 = 0.959. The beta-TCP dissolution rate decreases very sharply with time. This effect increases at higher pH or saturation. Several explanations are proposed and discussed, even though none is conclusive.

Journal Article↗
PubMed1993

In vitro production of toxin-coregulated pili by Vibrio cholerae El Tor.

Toxin-coregulated pili (TCP) have been shown to be a virulence determinant and protective antigen for Vibrio cholerae strains of classical biotype, but their role in infection by strains of the alternative El Tor biotype remains uncertain. In an attempt to demonstrate TCP production by El Tor vibrios we have over-expressed the El Tor tcpA gene in Escherichia coli, in order to prepare a biotype-specific anti-TcpA serum. This reagent proved to be a very sensitive indicator of TcpA production in immunoblotting studies, but failed to detect polymerized pili on the bacterial surface by immuno-electron microscopy (IEM). However, results with an analogous reagent which detects classical TcpA suggested that antisera to unprocessed TcpA do not efficiently recognize epitopes on native proteins. Accordingly we prepared a serum against a cell envelope fraction rich in processed El Tor TcpA. After extensive absorption this reagent reacted almost exclusively with TcpA by immunoblotting; when used in IEM, it allowed visualization of typical TCP bundles on the surfaces of each of five El Tor strains known to produce TcpA in vitro. We have previously reported that the El Tor strain O17 does not synthesize TcpA during growth in vitro, but that an O17 clone carrying a cosmid of classical origin expresses surface TCP. Using the biotype-specific anti-TcpA reagents in immunoblotting studies it has been possible to detect the product of the host chromosomal tcpA gene in such constructs; transcription of this gene was confirmed using biotype-specific tcpA probes. IEM revealed that El Tor TcpA was present in the TCP bundles associated with the O17 cosmid clones. Further studies suggest that regulation of the genes encoding TcpA and cholera toxin varies between different strains of El Tor biotype.

Antibodies, Bacterial↗
PubMed1996

The toxin-coregulated pilus is a colonization factor and protective antigen of Vibrio cholerae El Tor.

We have previously shown that insertional inactivation of tcpA, the gene encoding the major pilin subunit of the toxin-coregulated pilus (TCP), renders Vibrio cholerae O1 strains of El Tor biotype virtually avirulent in the infant mouse cholera model (IMCM). We now report that more refined mutants, bearing an in-frame deletion in tcpA, show a similar dramatic attenuation in vivo. In mixed-infection competition experiments the ratio of wild-type:mutant vibrios increased c. 10(3)-10(5) fold during a period of in vivo growth. An attempt to complement the delta tcpA mutants by providing a functional El Tor tcpA gene in trans was only partially successful. Sera raised against El Tor TcpA were able to passively protect infant mice against challenge with TCP-positive strains of homologous biotype and were also protective against isolates of the novel O139 serovar. These sera failed to protect against challenge with a strain of classical biotype, nor could antibodies to classical TCP confer immunity to El Tor challenge. We conclude that TCP is a critical colonization factor of V. cholerae O1 El Tor and that antibodies to TCP are sufficient to confer protection against such strains in the IMCM. Our data suggest that the biotype-specific epitopes carried by TcpA are of greater vaccine significance than those epitopes common to both proteins.

Animals↗
PubMed2002

Host-dependent recombination of a Tomato bushy stunt virus coat protein mutant yields truncated capsid subunits that form virus-like complexes which benefit systemic spread.

This study examined the contribution of the Tomato bushy stunt virus (TBSV) coat protein (CP) and its corresponding RNA to systemic infection of plants. Compared to results obtained with a mutant lacking the 5'-half of the CP gene, the presence of those CP-RNA sequences in another mutant benefited TBSV infection on Nicotiana benthamiana even though wild-type CP expression was eliminated by introduction of a small out-of-frame deletion. RT-PCR of viral RNA associated with rapid infections established by this CP frameshift deletion mutant revealed that in planta recombination had provided the progeny with the ability to express a truncated CP (tCP) with a block of N-proximal 30 residues deleted from the 66 amino acid RNA-binding domain. Subsequent biochemical characterizations revealed the presence of large ribonucleoprotein complexes that were shown to contain viral RNA as well as the approximately 38-kDa tCP. Electron microscopic examination of purified complexes showed particle-like structures that were nonuniform in size and shape compared to wild-type TBSV particles. Inoculation of pepper with the tCP-containing ribonucleoprotein complexes resulted in a rapid systemic infection similar to that caused by wild-type TBSV. In contrast, infections established in pepper by the original CP frameshift deletion mutant transcripts were restricted to inoculated leaves and did not yield recombinants capable of systemically infecting this host. In summary, TBSV possesses the flexibility to form alternative virion-like structures even if a substantial portion of the RNA-binding domain is deleted from the CP; mutants producing the tCP-containing particle-like structures are more effective for virus spread than those devoid of CP expression; and recombination events to produce the alternative tCP-RNA complexes are host-dependent.

Capsid↗
PubMed1984

Microbial biodegradation of 2,4,5-trichlorophenoxyacetic acid and chlorophenols.

We have succeeded in isolating a pure culture of Pseudomonas cepacia, AC1100, from a chemostat enrichment culture experiment that is capable of growing on 2,4,5-trichlorophenoxyacetic acid as its sole source of carbon and energy. AC1100 is not only capable of degrading 2,4,5-T but is also able to completely or partially dehalogenate a wide variety of halophenols. The regulation of the dehalogenating ability of AC1100 has been investigated which demonstrates that the enzyme(s) which allow the conversion of 2,4,5-T to 2,4,5-TCP are constitutive, while the enzymes that allow the degradation of 2,4,5-TCP are inducible by 2,4,5-TCP (or some metabolite of 2,4,5-TCP) but not by 2,4,5-T or other halophenols which can serve as substrates. Moreover, the 2,4,5-TCP degradative pathway is repressed by the presence of an abundant alternative carbon source. The detailed pathway of 2,4,5-T degradation by AC1100 is currently under study. Although field tests have yet to be conducted, laboratory experiments have demonstrated rapid and complete degradation of 2,4,5-T from contaminated soil. Soil previously contaminated with as much as 5,000 micrograms of 2,4,5-T/g of soil could be detoxified by AC1100 treatment, allowing the growth of plants sensitive to less than 10 micrograms 2,4,5-T/g of soil. Moreover soil contaminated with as much as 20,000 micrograms of 2,4,5-T/g of soil showed greater than 90% degradation after six weekly AC1100 treatments. After 2,4,5-T has been substantially degraded in contaminated soil the titer of AC1100 rapidly falls to nearly undetectable levels, which indicates that no serious ecological disturbance is likely to result from the application of AC1100. It appears possible that the treatment of contaminated areas with appropriate microorganisms may allow essentially a total restoration of the original soil condition.

2,4,5-Trichlorophenoxyacetic Acid↗
PubMed1994

Comparisons of the actions of high and low doses of the MAO inhibitor tranylcypromine on 5-HT2 binding sites in rat cortex.

Tranylcypromine (TCP) is a commercially available antidepressant drug, and recent literature reports suggest that high doses of this drug may be particularly effective in treating refractory depression. Down-regulation of 5-HT2 receptors in rat cortex is an effect produced after chronic administration of several antidepressants, and we have conducted a chronic study comparing low- and high-dose TCP in this regard. Male Sprague-Dawley rats were administered TCP (0.5 or 2.5 mg/kg/day) or vehicle (distilled water) via Alzet minipumps implanted subcutaneously in the dorsal thoracic area. Groups of rats were killed 4, 10 or 28 days after pump implantation and whole cortex was dissected out and utilized for preparation of a membrane fraction. Binding studies were performed with this fraction using 3H-ketanserin as the radioligand. Down-regulation (decrease in Bmax) of the 5-HT2 binding site was observed in high-dose animals after 10 and 28 days but not after 4 days. Low-dose TCP had no effect on 5-HT2 densities at any time interval. The affinity of 3H-ketanserin for the 5-HT2 site was not affected by either dose at any time interval. These results suggest that down-regulation of the 5-HT2 site may contribute to the efficacy of high-dose TCP in the treatment of refractory depression.

Animals↗
PubMed1981

Behaviour of IOP and pupil size after topical tranylcypromine in the rabbit eye.

Repeated topical administration of tranylcypromine (TCP) to adult rabbit eyes causes a transient rise in intraocular pressure (IOP) for one half hour and a dilatation of the pupil for more than two hours. These effects are blocked by phenoxybenzamin but not by timolol. Indomethacin blocks the rise in pressure but not the pupillary dilatation. Chemical denervation blocks both effects. The rise in IOP is not followed by a hypotensive phase. It is suggested that TCP has a dual effect on the eye. Firstly, TCP as a releaser of endogenous nor-epinephrine (nE) and as a monoamine oxydase inhibitor will induce accumulation of nE at the receptor-side which in turn probably stimulates the synthesis of PGs to raise IOP. Secondly, TCP as a prostacyclin synthase inhibitor may be responsible for the absence of a hypotensive response. This is strengthened by the finding that TCP inhibits the hypotensive response to nE but potentiates the dilatation of the pupil.

Animals↗
PubMed1975

Biodegradation of alpha-hexachlorocyclohexane. V. Characterization of the major urinary metabolites.

1. Free 2,4,5- and 2,4,6-trichlorophenol (TCP) have been identified by co-crystallization with authentic carrier as constituents of the urine of rats given 3-H-labelled alpha-hexachlorocyclohexane (alpha-HCH). The two TCP's constituted 7% and 63%, respectively, of the free phenol fraction which was found to account for less than 5% of all urinary metabolites. 2. 75% of the label contained in the urine of rats collected for four weeks after an i.p. dose of 14-C-alpha-HCH could be extracted into an organic solvent after alkaline and acid hydrolysis. The radioactive material thus extracted was examined by TLC and GLC. It is shown to consist mainly of chlorophenols and chlorothiophenols. 3. The extracts' major constituent was 2,4,6-TCP. Its amount was determined by GLC and was found to account for, on average, 45% of the total urinary metabolites. In conjunction with other evidence, this is considered to establish 2,4,6-TCP as the major product of alpha-HCH-biodegradation in the rat. 4. Chromatographic evidence is presented for 2,3,4,5-tetrachlorophenol and 2,4,5-TCP being other components of the chlorophenol fraction isolated from hydrolysed urine. 5. The amount of label associated with the chlorothiophenol fraction suggests a pathway involving endogenous thiol to be of significance in alpha-HCH-biodegradation in rats. Pretreatment with the drug itself increased the proportion of label associated with the fraction.

Animals↗