The completion of programs of study is an essential requirement for the development a new product, whether it be chemical products—subject to rules and regulations —medicines—subject to the rules and regulations regulatory agencies: EMA, FDA, ICH standards—or medical devices—subject to ISO standards. In the department we have extensive experience in the development of new compounds, which is associated with a high success rate in products that reach the clinical phase.

At our center, we offer a wide range ofin vivo programs of study conducted under good laboratory practices GLPs), covering the battery of programs of study by regulatory agencies for the development of any new product prior to its subsequent launch on the market.

In addition, we offer services in the phases prior to development , also known as screening phases, aimed at selecting the best candidates for subsequent assessment . This section the implementation of programs of study , which can be geared toward different objectives: decision-making when selecting or discarding candidates, planning programs of study timelines in preclinical phases to ensure compliance with all regulatory agency requirements, selecting model , or conducting programs of study , among others. In summary, personalized advice to ensure that development is carried out within the framework and to avoid subsequent cost overruns generated by failures in the design phases.

Our qualified staff specializes in work various in vivo experimental systems (rodents and non-rodents) and has access to modern housing facilities that comply with all requirements for animal housing and handling. Our animal welfare experts ensure the animals' health and well-being.

We currently offer the following programs of study :

• Dose escalation or acute toxicity study

• Repeated dose toxicity study -14 or 28 days-

• Repeated dose toxicity study - 90 days, 6 or 9 months -

In addition to the above, we design programs of study the needs of our clients in all phases of development drug development .

Dose escalation or acute toxicity study

The first programs of study be carried out, when there is no bibliography , are dose programs of study (or acute programs of study ), which form part of what are known programs of study general programs of study . In the case of development drugs and medical devices, in agreement the ICH Harmonized Tripartite Guideline M3 (R2)[1] and rule 10993-1[2] respectively, these programs of study the maximum dose to be used in programs of study repeated-dose toxicity programs of study to be determined.

Repeated dose toxicity study -14 or 28 days-

In the evaluation assessment the toxic characteristics of a substance, after an initial dose escalation (or acute toxicity) study, an assessment the product-toxic effect dose relationship is required in two animal species (one rodent and one non-rodent) after repeated dose administration. The issue 14 or 28 days—will depend on the intended use, exhibition clinical dosage, as set out in the current regulations.

Repeated dose toxicity study - 90 days, 6 or 9 months -

In the evaluation assessment the toxic characteristics of a substance, after an initial dose escalation (or acute toxicity) study, an assessment the product-toxic effect dose relationship is required in two animal species (one rodent and one non-rodent) after repeated dose administration. The issue 90 days, 6 or 9 months—will depend on the intended use, exhibition clinical dosage, as set out in the current regulations.

In agreement current legislation, for all products applied to the skin (area ), programs of study must be carried out to rule out possible adverse effects of the product on the body. Specifically, at least one single-dose skin tolerance study and one repeated-dose skin tolerance study that also provides general data are required. Finally, since the product may accidentally come into contact with the eyes, it is necessary to conduct a single-dose ocular tolerance study.

The goal is to detect any possible corrosive or irritant effect of the drug or essay product, as well as to determine Degree to which Degree affect the experimental system. It is therefore necessary to evaluate any skin lesions it may cause and check whether these are likely to be reversible.

To achieve these objectives, our center complies with the legal requirements demanded by regulatory agencies to carry out programs of study tolerance programs of study , including the following:

• Single and repeated dose skin tolerance study

• Single-dose and repeated-dose ocular tolerance study

• Study of parenteral tolerance at single and repeated doses

• Single-dose and repeated-dose vaginal tolerance study

• Single- and repeated-dose oral tolerance study

• Single- and repeated-dose nasal tolerance study

• Local Lymph essay : DA (LLNA:DA)

essay local lymph node essay is a essay allows potential skin sensitizers to be identified by determining the cell proliferation produced in the local lymph node. This makes it possible to quantify the local response generated by the body to the application of a essay product essay the skin.

• Intradermal reactivity test

This study is particularly effective in providing in-depth knowledge and assessing the potential of a product to cause a sensitization (or hypersensitivity) response after intradermal administration. Thanks to the technical specificity of the test, it is possible to determine with maximum precision the irritant effects that a given product may cause on the skin, making this essay test in the development a new product.

This department the possibility of obtaining biological samples from in vivo experimental systems for subsequent analysis. Our laboratories are officially certified by the Government of Navarra, accrediting compliance with the good laboratory practices GLPs) required by Spanish legislation.

In addition, the laboratory collaborates with other Departments sections of the University to expand the range of analysis options available after obtaining biological samples. 

• Pharmacokinetic study

programs of study (PK) programs of study make it possible to determine the evolution of the concentration of essay product essay a biological matrix, from its administration to its elimination. In this way, they provide information about its absorption, distribution, metabolism, and excretion in the body and allow its profile to be characterized.

• Toxicokinetic study

In the evaluation assessment the toxic characteristics of a substance, together with repeated dose programs of study , a toxicokinetic (TK) study is required to correlate exhibition the product with toxicological findings, contributing to the assessment the relevance of these findings for clinical safety and providing information on the potential for drug accumulation.

essay the comet

This essay used to evaluate the genotoxic capacity of a compound. Genotoxicity in this case is detected based on issue DNA breaks that the compound under study may cause. In our department, two models of this test are used: the standard test and the test performed in combination with the FPG enzyme, which also allows the detection instructions and alkylated instructions . This is a growing method, capable of detecting genotoxicity very accurately and effectively.

As business in the implementation of this technique, the department different options for carrying out this essay subject . There are several variants of the same, which allow the test to be performed both on cells obtained in vitro and on cells obtained in vivo, with the possibility in the latter variant of using cells from fresh or frozen tissues.

essay

This is an essay of detecting the genotoxic effects of a compound on a given subject through the presence of micronuclei (MN), small independent nuclei additional to the main cell nucleus, related to the presence of structural or numerical chromosomal aberrations.

This technique is designed to be performed on both cells obtained in vitro and cells obtained in vivo.

A very important part of characterizing a drug is knowing its potential genotoxic effects. A drug's ability to cause genetic damage can lead to serious health problems, even causing the development diseases with very serious prognoses.

Therefore, it is essential to detect this subject behavior in a compound as early as possible during the earliest stages of its development. The analysis of profile includes, in addition to conducting the tests required by regulatory agencies, the interpretation of results, involvement and advice in the design phases design to initiation, and strategies to follow in the event that in vitro tests detect genotoxic activity of the drug.

At our facilities, we have a wide range ofin vitro and in vivo tests for the comprehensive assessment the genotoxicity of a substance or drug, including the following:

1. AMES test: toxicity and mutagenicity test

2. MiniAMES test

3. SOS/umu test

4. essay the comet

5. essay

Ames test

It is essential to know the effects that a substance may have on genetic material before placing it on the market. The AMES test is the ideal in vitro essay for carrying out this assessment, as it allows us to determine the general toxicity and mutagenicity of a substance on five different bacterial strains. Our department two different tests, which together generate comprehensive and robust information on the profile of the substance under study.

AMES test: Toxicity test. This is an essay of providing very useful information about the concentrations at which a substance is capable of causing genetic damage to bacteria. In addition, it will provide information on the solubility of the product, avoiding subsequent problems for other tests.

AMES test: Mutagenicity test. In this second variant, the goal essay to gain in-depth knowledge of the mutagenic capacity of a substance, allowing us to know whether this mutagenic effect is due to the product or to one of its metabolites.

In our laboratory also developed the "enhanced" AMES test for subject impurities: toxicity and mutagenicity test. A variant of the Ames test required by regulatory authorities.

MiniAMES test

This essay a reduced version of essay AMES essay , allowing it to be used as test or product selection test , as it requires less essay product, reagents, and cells.

As with the AMES test, two independent MiniAMES tests are carried out at our facilities, one for general toxicity and one for mutagenicity. Combining the results of both provides comprehensive and highly useful information when studying the profile of one or more substances.

MiniAMES test: Toxicity test. This essay will allow essay to determine the dose at which a given substance is toxic to cells, as well as information regarding its solubility. This will prevent subsequent problems for other tests.

MiniAMES test: Mutagenicity test. This essay on gaining in-depth knowledge of the mutagenic capacity of substances, allowing us to determine whether this mutagenic effect is due to the product itself or to one of its metabolites.

SOS/umu test

The SOS/umu test is an in vitro essay based on the bacterium Salmonella Typhimurium (strain TA1535/pSK1002) that detects DNA damage by inducing the SOS repair system. When a genotoxic lesion occurs, the umuC reporter gene is activated, whose activity is easily measured through a colorimetric reaction, allowing for rapid and automated screening of chemical compounds.

Although it does not have full regulatory validity on its own, i.e., it does not replace the Ames test in the data packages required by agencies such as the EMA or the FDA, it is widely accepted and used as a highly efficient tool in the early stages development and in water quality monitoring (under ISO standards).

essay the comet

This essay used to evaluate the genotoxic capacity of a compound. Genotoxicity in this case is detected based on issue DNA breaks that the compound under study may cause. In our department, two models of this test are used: the standard test and the test performed in combination with the FPG enzyme, which also allows the detection instructions and alkylated instructions . This is a growing method, capable of detecting genotoxicity very accurately and effectively.

As business in the implementation of this technique, the department different options for carrying out this essay subject . There are several variants of the same, which allow the test to be performed both on cells obtained in vitro and on cells obtained in vivo, with the possibility in the latter variant of using cells from fresh or frozen tissues.

essay

This is an essay of detecting the genotoxic effects of a compound on a given subject through the presence of micronuclei (MN), small independent nuclei additional to the main cell nucleus, related to the presence of structural or numerical chromosomal aberrations.

This technique is designed to be performed on both cells obtained in vitro and cells obtained in vivo.

The cytotoxic potential of a substance is defined as the ability of a product to cause an alteration in basic cellular functions, causing damage to the cell that affects its viability. In the development a new drug, it is essential to have knowledge its profile , as this allows us to predict the effects that the compound will have during its subsequent development and/or clinical development .

Conducting tests capable of detecting this cytotoxicity and assessing cell viability is essential when developing a new product, as it provides highly useful information for researchers interested in the basic toxicity of a substance. It is possible to characterize this profile in different cell models, with tests being carried out on primary cultures, tissue explants, or established cell lines.

The department Toxicology at the University of Navarra has specialized for more than two decades in conducting a wide variety of programs of study and cell viability programs of study , as well as providing advice both during the design phase and subsequently during the assessment the results obtained. The different programs of study out include the following:

1. MTT reduction essay

2. essay neutral red uptake

3. essay

4. essay cell viability estimation by exclusion of trypan blue dye

5. essay

6. essay ATP bioluminescence

7. LDH release cytotoxicity essay

8. ROS production (essay )

MTT reduction essay

The (3-4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) essay is one of the most widely used worldwide and one of the issue assays in research MTT is a salt capable of reacting with mitochondrial dehydrogenases to form soluble blue formazan crystals, whose absorbance can be measured and is proportional to the number of viable cells.

This allows it to have several applications:

• As essay cytotoxicity essay , to evaluate cell death after exhibition cells to an essay product.

• For any essay requires knowledge of the issue percentage of cells that grow under a given condition

• To study cell proliferation

essay capturing neutral red

The essay capturing the dye 2-amino-3-ethyl-7-diethyl amino phenacine hydrochloride, also known as neutral red, is based on measuring the absorbance of this dye in cells, where it binds to lysosomes thanks to its ability to cross the plasma membrane. This measurement is directly proportional to issue living cells present, providing a reliable indication of the cell damage or death caused by a previously essay product when compared with the controls.

This makes it applicable to different programs of study:

• As essay cytotoxicity essay , to evaluate cell death after exhibition cells to an essay product.

• For any essay requires knowledge of the issue percentage of cells that grow under a given condition

• To study cell proliferation

cell proliferation essay

This is an essay on comparing the survival capacity of cells treated with a specific essay product essay that of normal cells. Its basic principle and the short time required to obtain results make this method one of the most effective for evaluating the profile of a compound.

essay cell viability estimation by exclusion of trypan blue dye

The dye essay is frequently used to determine the proportion or issue viable cells present in a cell suspension. This is particularly useful when determining and quantifying how a particular essay compound acts essay a subject . Trypan blue dye is only able to penetrate dead or damaged cells, making it possible to calculate cell viability after treatment with a given substance.

essay

Determining the clonal efficiency of cells after they have been subjected to a specific essay product is a direct measure of their reproductive integrity. The essay seeks to identify a product's potential to eradicate the ability of cells to divide indefinitely, which is commonly associated with tumor cells.

This makes the essay a tool effective tool when it comes to:

• Knowing the issue percentage of cells that, after a certain treatment with an essay product, survive and are capable of multiplying to form visible colonies of cells.

• Knowing the clonal efficiency of a given cell line

• Study cell proliferation

essay ATP bioluminescence

This essay based on the measurement of luminescence emitted in the process catalyzed by the enzyme luciferase, which reacts with oxygen in the presence of ATP, emitting said luminescence. When ATP is the limiting factor, the measurement of luminescence emitted is proportional to the amount of ATP generated by the cells, which is directly associated with issue cells present.

This essay this essay several potential applications:

• As essay cytotoxicity essay , to evaluate cell death after exhibition cells to an essay product.

• To study cell proliferation

• As an indirect measure of bacteria, fungi, yeasts, and other microorganisms in food products, beverages, water, cosmetics, or other products.

• As essay enzymes that produce or degrade ATP

LDH release cytotoxicity essay

This essay based on measuring the amount of lactate dehydrogenase (LDH), a very stable cytosolic enzyme that only appears in the culture medium after the membrane of dead or toxic-affected cells has been lysed. This makes it possible to determine even the slightest cytotoxicity of an essay product, since the amount of LDH in the medium will be directly proportional to issue damaged cells.

There are several applications for this essay:

• Assess the cytotoxicity of a given compound or other agent, making it possible to detect it at very early stages of its development

• Calculate the issue of cells that grow or die under a given condition.

• Evaluate cytotoxicity mediated by other cells

ROS production (essay )

This in vitro essay focuses on measuring Reactive Oxygen Species (ROS) present in the culture medium. ROS are known to perform various functions within the cell, so any alteration in the latter will vary the concentration of these compounds. Cell treatment with a specific essay product essay produce cell toxicity, which can be detected by measuring these ROS, making this technique a tool useful tool in assessment cytotoxicity and cell viability.

When developing a new compound, it is very common to need to know the effect it may have on the skin. Potential skin toxicity must be detected in the earliest stages of development , for which there are several tests and trials capable of determining and quantifying this toxicity. At our facilities, we have the most advanced techniques for this purpose, including skin irritation and corrosion tests, and skin phototoxicity tests.

The University of Navarra's commitment to integrating the 3Rs principle has led our department search for new in vitro experimental systems capable of accurately reproducing the conditions of in vivo experimental models. This has made it possible to perform in vitro tests that were previously carried out in other models, providing very useful and reliable information without the need to use in vivo systems.

Skin irritation and corrosion tests

The use of in vitro models is applicable in many cases for skin programs of study . Our laboratories offer the possibility of conducting skin programs of study and corrosion programs of study the experimental system known as EPIDERM or Reconstructed Human Epidermis (RHE), model reliably reproduces the human epidermis.

• The skin essay goal the skin irritation potential of both a specific product and a mixture, in order to classify them agreement United Nations Globally Harmonized System of Classification and Labeling of Chemicals (UN GHS).

• The dermal essay , meanwhile, focuses on assessment dermal corrosive potential of both specific substances and mixtures, in order to classify them according to the United Nations Globally Harmonized System of Classification and Labeling of Chemicals (UN GHS).

Dermal phototoxicity tests

Studying the phototoxic potential of a substance is essential for predicting the possible negative effects it may have on the skin. Our department carry out essay using the EPIDERM in vitro model.

The department the University of Navarra has a laboratory in the analysis of biological samples. Our staff has extensive experience in performing various biological analyses, all in accordance with Spanish legislation on good laboratory practices GLPs). Our specialists have access to the best facilities, equipped with the latest systems and measuring equipment for performing various tests, including biochemical and hematological analyses, coagulation tests, and urine analyses.

Biochemical analysis

Our equipment allows biochemical analysis of serum and/or plasma samples, providing information on the following list of parameters:

Hematological analysis

Our laboratory has certified equipment for measuring various parameters in whole blood, including:

Coagulation tests

In our laboratories, it is also possible to perform plasma coagulation tests, which can determine a series of different parameters, as shown below:

• PT (prothrombin time)

• APTT (Activated Partial Thromboplastin Time)

• Fibrinogen (factor I)

Our facilities are equipped with the most specialized equipment for performing urine analysis, making it possible to obtain information on the following parameters:

There essay also the possibility of performing ELISA (Enzyme-Linked Immunosorbent Assay) tests. This Biochemistry technique Biochemistry the detection and quantification of substances—such as proteins, antibodies, or drugs—through the use of specific antibodies and an enzymatic reaction that generates a measurable signal. ELISA tests are offered, with approach special approach the quantification of inflammatory cytokines in different biological matrices.

We analyze, among other markers, IL1α and IL6 in culture medium in reconstituted human programs of study ; and in serum or plasma we determine IFNγ, TNFα, and IL6 in monkeys; IgE, IL12, and IL6 in rats; as well as IL6, TNFα, and IFNγ in mice. We also offer the possibility of expanding the analytical panel and adapting it to the specific needs of each project.

The activity of a drug against different groups of microorganisms (bacteria, molds, yeasts) can be determined by the effect it has on the metabolism of microorganisms (destruction or inhibition of growth). This activity is defined as the intrinsic ability of the compound to interfere with essential processes of the microbial cell (such as cell wall synthesis, DNA replication, protein synthesis, or membrane integrity), causing growth inhibition (bacteriostatic or fungistatic effect) or death of the microorganism (bactericidal or fungicidal effect).

This activity can be measured in vitro using different methodologies and can be compared with that produced by reference letter substances that have demonstrated their action against certain microorganisms according to international standards (CLSI, EUCAST).

• Antibiograms: these are microbiological tests that determine the sensitivity or resistance of a bacterium or fungus to different antibiotics/antifungals. This analysis is essential in the field of clinical microbiology to guide the appropriate treatment of bacterial infections. These are agar diffusion methods in which discs impregnated with the substance to be tested are placed on a culture plate inoculated with the microorganism to be tested. The growth inhibition halo around the disc is measured.

• Turbidimeter methods: these are performed by microdilution in a plate, where decreasing concentrations of the product are tested in a liquid medium to determine the Minimum Inhibitory Concentration (MIC) and the Minimum Bactericidal Concentration (MBC) and/or Minimum Fungicidal Concentration (MFC). The MIC is the leave concentration leave an antimicrobial agent (expressed in μg/mL) that inhibits the growth of a microorganism (under defined in vitro conditions), while the MBC or MFC is the leave concentration leave an antimicrobial agent capable of killing 99.9% of the initial microorganism population under in vitro conditions.

• Determination of MIC in solid medium: this consists of preparing agar plates containing decreasing concentrations of an antimicrobial agent. Once the medium has solidified, a standardized suspension of the microorganism is inoculated onto the surface of each plate. After incubation at the appropriate temperature and for the appropriate time, growth is evaluated at each concentration, and the MIC is determined as the lowest concentration of the product at which no visible growth of the microorganism is observed.

Antimicrobial preservatives are chemicals added to products (such as food, medicines, cosmetics, or industrial products) with the goal inhibiting the growth or destroying microorganisms (bacteria, fungi, yeasts, molds, etc.) that could alter their quality, safety, or stability. The mechanisms of action on microorganisms are diverse (alteration of the cell membrane, inhibition of enzymes essential for their metabolism, interference with DNA replication, etc.) and the result the inhibition of the growth of microorganisms or a significant reduction in their numbers within a given time. 

This study consists of inoculating the essay product essay a known quantity of microorganisms and determining their viability at intervals of time at a given temperature. The preservatives will be considered effective if, under the test conditions, there is a significant decrease and/or no increase (depending on the case) in issue microorganisms. 

These programs of study intended to guarantee the microbiological quality of medicines that have been stored for varying periods of time under specific temperature and humidity conditions (compliance with the microbiological criteria established by the Pharmacopoeias). These programs of study :

• Validation of microbiological control methods: this involves demonstrating that the microbiological technique applied allows the target microorganism to be recovered or detected in the presence of the pharmaceutical product, eliminating any possible inhibition that could be caused by the active ingredient and the resulting false negatives. We have a collection of reference letter strains reference letter fortify the samples and compare the recovery of microorganisms in the presence and absence of the drug. 

• Microbiological control in medicines: performing the requested microbiological tests using the previously validated control method.

The microbiological tests carried out in the programs of study are as follows:

• total aerobic microorganism count (TAMC) 

• total mold and yeast count (TYMC)

• count of bile-tolerant Gram-negative bacteria 

• research detection) of Escherichia coli

• research detection) of bile-tolerant Gram-negative bacteria 

• research detection) of Pseudomonas aeruginosa

• research detection) of Staphylococcus aureus

• research detection) of Salmonella spp

The laboratory also offers other microbiological tests on pharmaceutical products, following the guidelines of the Pharmacopoeias.

• Validation of culture media (growth promotion and inhibitory properties): this consists of verifying that the media used in the microbiological control of pharmaceutical products are capable of promoting the growth of target microorganisms (or, where applicable, inhibiting them), thus ensuring that the medium is suitable for use. Known concentrations of reference letter strains are used reference letter the recovery (and, where applicable, inhibition) of microorganisms in the medium to be validated is compared with the culture medium used as a control (previously validated).

• Microbiological control of water for injectables: this involves checking that the water used in injectable formulations complies with the requirements of the Pharmacopoeia, in order to guarantee the microbiological quality of the final product and comply with requirements Manufacturing requirements .

• essay : the goal to confirm the total absence of bacteria, molds, or yeasts in medical or pharmaceutical products, which according to the Pharmacopoeia must be sterile. Before carrying out the essay , it is necessary to validate the procedure (by carrying out fortifications with reference letter strains) in order to demonstrate that, if present, we are able to detect microorganisms, eliminating any possible inhibition that could be caused by the active ingredient and the resulting false negatives. The essay can essay performed by filtration or direct inoculation, the former being the option of choice whenever possible.