DETERMINATION OF ACTIVE INGREDIENT IN TAMSULOSIN DRUG BY USING HPLC

This study was aimed to development and modification the method of analysis for active ingredient in Tamsulosin drug by using high performance liquid chromatography (HPLC) also determined shelf life and storage conditions for Tamsulosin. Chromatographic conditions utilized stationary phase C18 (250*4.6) mobile phase 0.05Nmixture of di hydrogen ortho phosphate and Acetonitrile 55:45 , preserve the flow rate near (1ml/min) and length of wave has been 275nm, The retention time found to be 10 minute for the HPLC process . The Tamsulosin assay result was found to be 99.93% . The calibration curve linearity analysis results showed a strong linear relationship in the concentration range (10-200ppm) and the correlation coefficient, slope and intercept value were 0.9933,11796,190017, respectively The percentage recovery was found 99%. LOD value was found to 0.00053 µg/ml and LOQ value was found to 0.0016 µg/ml , Precision was found to be 99.49% Robustness was found to 99.69%. Our proposed procedure confirmed a group of merits such as Linear ,accurate ,precise ,and robust , could be recommended for determination of Tamsulosin.


‫العراقية‬ ‫الزراعية‬ ‫العلوم‬ ‫مجلة‬
In the prostate, prostatic capsule, prostatic urethra and bladder neck ,Tamsulosin is a selective antagonist of alpha1-A and alpha1-B adrenoceptors (10). At least three different subtypes of alpha adrenoceptor was identified as alpha-1A alpha-1B and alpha-1D heir distribution varies between tissue and organs of humans (16,17) . It is sold under the trade name Urimax, Veltam and Flomax is a medication used to treat symptomatic benign prostatic hyperplasia (BPH) (19) because, it is stimulate the muscle of bladder so lead to the urine pass out easily (13) also help with the passage of kidney stones, and for urinary retention along with other measures (7). About 70 % of the alpha1receptors is alpha-1A type. Blocking these receptors allows smooth muscles in the neck of the bladder to relax (23,5) . A common problem that affects about one third of men over 50 years of age is benign prostatic hyperplasia (BPH). In the United States, Over 14 million men have BPH symptoms (8).
is a non-malignant prostate enlargement, a wedge-shaped gland that, when it arises from the bladder, covers the male urethra. this enlargement is a natural consequence of aging, but it can be related to symptoms that may be distressing, but rarely life-threatening. firstly affects the inner parts of prostate. Enlargement of prostate gland causes a gradual squeezing of urethra, sometimes it causes difficulty in micturition and may create other urinary problems (9). drugs contains two main contents active ingredient and inactive ingredient. tamsulosin contain active ingredient is 0.4 mg that responsible for its effect and give curative benefit (29) . it is enables urine to pass easily , help the passage of kidney stones and decrease smooth muscle tension (8) while inactive ingredient it is substance that include in the composition and manufacture but not responsible for its effect and not curative benefit .the role give specific shape and texture such as titanium dioxide, ferric oxide and microcrystalline cellulose (4,11). In Pharmaceutical product stability testing is a complex series of procedures requiring significant costs, time consumption and scientific knowledge in order to build consistency, effectiveness and protection in the formulation of drugs (5) .Stability of a pharmaceutical produce may be characterized as the a capacity of a particular Ingredients Formulation stay within its microbiological ,physical, chemical, toxicological, protective and information requirements in a particular System of Container/Closure (24,27) .That is in other words, within the limits of the extent to which a substance retains defined, during its stocking and usage time, The same one characteristics and properties existed at the moment that it was packaged . The stability test thus tests the influence of ecological factors on the consistency of medicinal substance or a Producer used to predict the shelf life of the Determine proper conditions for storage and suggest labeling Instructions. Furthermore, The data produced during the stability test is an important prerequisite for any medication to be regulatory approved (12,14) One from the very significant parameters of pharmaceutical products is stability studies. The production of small molecule drug products is also important., mostly in the determination of stability properties, considering the value of the physical condition of the substance. Stability checking It is used to estimate shelf life for formulated items, to assess proper storage conditions (1) Stability Prior to chromatography investigations, many analyses quickly decompose for instance, Extraction, When preparation the sample solutions, While storing the prepared vials, Clean-up washing, phase transition (15). Stability is taken care of by the consistency test (26) Various analytical techniques they've been documented in the Survey of Literature for the study of Tamsulosin in drugs such as UV-Spectroscopy (6) Reversed-Phase High, Performance ,Liquid Chromatography (RP-HPLC) (20 (21,25) , spectrophotometry (20) , Thin layer chromatography ( TLC) (22) liquid chromatography mass spectrometry( LC -MS) (2) and UPLC method (28) . those methods have some disadvantages, such as the need for more time, an expert person, and a high cost (3). Therefore, it is of great significance to develop rapid and straightforward methods for tamsulosin determination ,in this approach our job was to establish a suiTable procedure for the determination of tamsulosin .the present work deal with HPLC stability for tamsulosin because it was sensitivity, accuracy and reproducibility both in bulk and in other dosage types.

Limit of detection( LOD) and Limit of quantitation (LOQ) LOD :The limit of detection of an analytical
person technique in a sample is the smallest quantity of analyte it is possible to detect this as an exact value, but not necessarily quantitated. LOQ: The limit of quantitation of an individual analytical method is the smallest In a sample, the quantity of analyte that could be quantitatively measured With enough precision and accuracy. The sensitivity of the analytical methodology was measured by means of the lowest detection limit and the lowest quantitation limit. The LOD and LOQ were calculated using the mathematical equation given in the Table 2.

Precision
Standard deviation or relative standard deviation is a measure of precision. The sample solution containing 50% was analyzed at for three times. Accuracy Accuracy analysis at three stages of normal addition from the advertised formulation confirmed the accuracy of the process.. % recovery has been reported for Tamsulosin. The rang of the procedure justifies a recovery of between 98% and 102%.

Robustness
By measuring the impact of small but intentional variations in the chromatographic conditions, the robustness was examined. The implementation of minor improvements, such as changes in the flow rate. The impact of these modifications on the findings have been investigated.

RESULTS AND DISCATION Method of Assay :
Tamsulosin was the target of an assay. The protocol was followed in preparing the standard and sample solutions. chromatographic conditions used clarify in Table 1. the standard solution was injected in 2 replicates and the sample was injected in 2 replicates into the HPLC port. The assay results are listed in Table 3. and representative chromatograms are shown in Figures 1. and 2.   show calibration curve between concentration and peak area, Table 4. Show result the linearity of Tamsulosin from calibration curve in Figure 3. the results was found to be correlation coefficients (R 2 ) 0.9933 , slop =11796 and intercept =19007. Table 4. Linearity of tamsulosin dru

Method validation
In accordance with the International Conference on, the established method was validated for checking of assay procedures for its precision, accuracy, linearity, and robustness  Table 5 .  µg/mL was injected three times ,The precision was determined by %RSD ( Relative standard deviation ) was found to be 1 less than 2 % .The results of system precision is presented in Table 6.  Figure 13. Precision of tamsulosin LOD and LOQ LOD denotes the lowest analyte concentration that can be detected, while LOQ denotes the lowest analyte concentration that can be quantified.. LOD and LOQ can be calculated by slope and standard deviation from calibration curve after applying equation in Table (2) the value was found LOD 0.00053 µg/ml and LOQ 0.0016µg/ml usually LOD high concentration LOQ . Robustness: Robustness is a measure of method ability to remain unaffected by small changes such as change in flow rate from 1min/ml to 1.2 min/ml .No significant different data by change in flow right ,The robustness determined by RSD %was found 1.6 less than 2 % so, this is method was robust The results of robustness of the method for flow rate is presented in Table 7.

Figure 14. Robustness of tamsulosin Stability study
Can be defined is a complex set of procedures due to great coast and continue long time may be 3 or 6 months to get quality , effective and safety for drugs . It is very important because determined shelf life after opening for Tamsulosin. In pharmaceutical Industries the research and development dept. work on the develop of the products and after finishing the products they put trail batch samples for each new product in the stability devise for period 3 months to make sure the new formula or new products is stable or not stable .The must do analysis for the same sample weekly ,monthly and check the result .if the sample no change in physical or chemical properties that mean is the sample formula is stable .The stability study result for 3 months continuously give you indication about the formula .by observing the result in Table 8. we find assay is 99% this means that is within limits and no change in description that mean is the formula is good and accepted and can transfer the Tamsulosin to the planning department or production department to product and sale in the market. From the results the shelf life after opening of Tamsulosin is 3 months .