## 3 Results and Discussion
### 2-AP Contents in Raw and Cooked Rice  
The results of 2-AP contents in raw and cooked rice of ten different traditional and improved varieties grown in the wet season of 2020 are shown in Figure 1. In raw rice samples, 2-AP content was the highest in Pusa 1652 (Improved Kala Namak), followed by Kala Namak-2 and Kala Jeera (Figure 1a). The 2-AP level in the Pusa 1652 (Improved Kala Namak) raw rice sample was almost double of that in Kala Namak-2. No significant differences were found in the 2-AP levels of Pusa Basmati-1, Pusa Basmati-1509, Jeera-32, and Govind Bhog (Figure 1a). The rice samples of Kala Namak-1 and Kala Nuniya had 2-AP levels below the level of detection, similar to that observed in the non-aromatic rice variety, IR-64. Considering that Kala Namak is well-known aromatic rice, the aromatic property of the Kala Namak 1 sample similar to a non-aromatic rice with respect to 2-AP raises doubt on the authenticity of this sample used in our study.  
Figure 1b shows the 2-AP levels in the cooked rice samples. The cooked rice samples exhibited approximately 20-to-60-fold increase in the amount of 2-AP compared to the corresponding raw rice samples, demonstrating that 2-AP may be significantly increased during cooking. Akin to the trend observed in raw rice samples, the cooked sample of Pusa 1652 (Improved Kala Namak) had the highest amount of 2-AP compared to all other cooked samples. For the rest of the samples also, the pattern of 2-AP content was almost similar, as seen in their raw samples. The observed fold increase in the 2-AP content in cooked rice of Pusa 1652 (Improved Kala Namak) (63-fold) and Kala Namak-2 (57-fold) rice samples was higher than the Pusa Basmati-1509 (35-fold), Pusa Basmati-1 (31-fold), Govind Bhog (31-fold), Jeera-32 (25-fold), and Kala Jeera (18-fold). The Pusa 1652 (Improved Kala Namak) line was derived from a cross of two aromatic donors, traditional Kalamamak and Pusa 1176, and further backcrossed to Kalamamak. Pusa 1176 was derived from Bindli Mutant 68 (BM68) X ARC line. The BM 68 is a mutant of Bindli, a traditional tall short grain aromatic rice line. For non-aromatic rice, IR 64 and the two mild-aromatic 2 AP rice samples, Kala Nuniya and Kalanamak 1, the fold increase in the 2-AP content was not detected.  
The 2-AP contents in raw and cooked rice samples of the above varieties were also estimated in the stored material that were grown and harvested in 2019 (Figure S4). The observed 2-AP levels in the raw and cooked rice samples of 2019 were lower than those harvested in 2020. Interestingly, the 2-AP levels in the raw rice samples of all varieties studied were below the level of detection (Figure S4a). However, statistically significant variations in 2-AP levels were found in the cooked rice (Figure S4b). The cooked sample of Pusa 1652 (Improved Kala Namak) from the year 2019 also had the highest 2-AP content,  
Figure 1: (**a**) 2-acetyl-1-pyrroline (2-AP) content in raw, and (**b**) cooked rice samples of ten different traditional and improved varieties grown in the wet-season of 2020. The 2-AP content below the level of detection is indicated as hash (#). Different letters indicate significant differences at \(p<0.05\), as determined by [PERSON]’s test. Pusa-1652 (Improved Kala Namak) is abbreviated as Pusa-1652 (IKN).  
followed by Kala Namak-2 and Kala Jeera. The observed fold increase in 2-AP content due to cooking of 2019 samples was also lower than the increase observed in freshly harvested rice samples of 2020. Similar to 2020 harvested samples, the non-aromatic and the mid-aromatic genotypes harvested in 2019 did not show fold increase in the 2-AP content. For aromatic genotypes, the observed fold increase in the 2-AP content was in the range of 10.8-29.9. Collectively, these findings indicate that significant loss occurs in 2-AP content during storage, as reported previously [9; 21]. Additionally, compared to raw rice, the analysis of cooked rice samples could be crucial to understanding the aromatic nature of rice.