<p>Lettuce is a vegetable species with a high commercial value and a short production cycle, requiring precise management to ensure profitable production. For this, the nutritional providing is an important factor, highlining the nitrogen, a macronutrient considered essential for the proper development of plants due to its participation in the composition of the main plant molecules, such as chlorophyll. Non-destructive strategies to monitor nutritional balance are essential to avoid nutrient deficiencies or excesses during the lettuce cycle. Agriculture 4.0 introduces new technological methods to achieve this goal. This study aimed to verify the potential of using a portable spectrophotometer to estimate nitrogen in lettuce shoots. The goal was to provide farmers with a non-destructive, precise, quick, cheap, and waste-free method for nitrogen monitoring. This method was compared against destructive laboratory analysis for shoot nitrogen and chlorophyll and the widely used SPAD chlorophyll meter. For this, a greenhouse experiment was conducted with lettuce cropped under N levels input, corresponding to 0, 25, 50, 75, 100 and 125% of the lettuce recommendation. The estimation for N in the lettuce shoot was performed by destructive analysis: (i) shoot N; (ii) chlorophyll and non-destructive analysis: (iii) chlorophyll meter SPAD and (iv) a portable spectrophotometer. Non-destructive measurements were performed 3 times at: 20, 23 and 26&#xa0;days after transplanting (DAT). The lettuce harvest occurred 28 DAT, to determine N shoot uptake and chlorophyll, using a destructive way, and lettuce production. The estimation of N shoot in lettuce using the portable spectrophotometer, a non-destructive tool, showed a high correlation to the standard destructive laboratory analysis and the chlorophyll meter (SPAD). The data showed a high potential, for the N monitoring shoot in lettuce, using the proposed portable spectrophotometer, on farm.</p>