Nitrates present one of the major groundwater contaminants in the world. In the groundwater of Zagreb aquifer they are characterized as one of the five main groups of contaminants. The most important thing associated with decrease of contaminant concentration is the definition of its origin. Various geochemical and statistical methods have been used for the determination of nitrates origin and related geochemical processes in the Zagreb aquifer. Within geochemical methods, water stable isotopes and nitrate stable isotopes were calculated on sampled river and groundwater. Sava River and groundwater was sampled in the period from November 2015 to October 2016. Sampling was preliminarily performed on 56 observation wells, after which 30 observation wells were selected for monitoring of nitrate stable isotopes in time. Molar ratios of nitrate and chlorides, nitrate and sodium, and nitrate and potassium were calculated for both historical groundwater chemical analysis and for chemical analysis conducted on sampled river and groundwater. Correlation and multivariate statistical analysis have been used within statistical methods. All of the statistical analysis were done on historical groundwater quality data as well as on data measured on sampled river and groundwater. Historical data analysis of nitrate concentrations showed that there are two main areas of Zagreb aquifer where elevated nitrate concentrations have been identified. One area is located on the left bank of the Sava River, in the dominantly urban area, while other is located on the right bank, in the dominantly rural area, where agricultural activity is also present. Water stable isotopes showed that groundwater is isotopically most similar to Sava River and precipitation from Ljubljana, and least to the precipitation that falls in the area of Zagreb aquifer. Interpretation of nitrates stable isotopes showed that nitrate origin within the sampling period was organic. Wastewater and manure have been recognized as main sources of nitrate contamination. Comparison of δ15N-NO3- with nitrate concentrations, as well the values of calculated enrichment factors, suggest that denitrification is a negligible process that takes place in alluvial part of Zagreb aquifer. Results also suggest that nitrogen in Zagreb aquifer is in large extent introduced in the form of ammonium ions, after which is transformed to nitrates by nitrification. This assumption is supported by the calculated interval of δ18O-NO3- characteristic for nitrification of organic nitrogen compounds. Correlation analysis showed that chloride and sodium concentrations present very important parameters in study of nitrate origin. Factor and cluster analysis conducted on chemical analysis data showed that chlorides, sulphates, sodium and potassium present variables that can be used as main indicators of anthropogenic influence on groundwater quality in the Zagreb aquifer. The use of cluster analysis allowed the definition of observation wells where nitrates concentrations were mostly related to wastewater or manure. Four clusters of observation wells were defined based on nitrate, chlorides, sulphates, sodium and potassium concentrations, values of stable nitrate isotopes and molar ratios of nitrates to all other used ions. For observation wells in three clusters wastewater was defined as main source of contamination. In that sense the study of the molar ratios, especially nitrate and potassium, was found to be crucial. Groundwater from observation wells were manure was identified as main source of contamination had the highest molar ratios and lowest δ15N-NO3- values. It has been concluded that investigation of molar ratios, in combination with nitrate stable isotopes, can be very effectively used in estimation of nitrate origin in groundwater of Zagreb aquifer. In addition, correlation and multivariate statistical analysis have shown that nitrates present parameter whose stability and transformation in the groundwater of Zagreb aquifer is basically dependent on the dissolved oxygen concentrations. The nitrate trend analysis established the existence of statistically significant decreasing trends of nitrate concentrations, except in Petruševec and Sašnjak and Žitnjak well field influence area. It has been shown that evaluation of nitrate trends in different levels, in combination with confidence and prediction intervals, can provide more realistic insight in areas where nitrates contamination occurs. Also, it has been concluded that linear regression presents appropriate method for estimation of nitrate trends.